United States      Region III      Region III       EPA 903-R-10-002
Environmental Protection  Chesapeake Bay    Water Protection    CBP/TRS 301-10
Agency        Program Office     Division       May 2010

In coordination with the Office of Water/Office of Science and Technology, Washington, D.C., and the states of Delaware,
Maryland, New York, Pennsylvania, Virginia and West Virginia and the District of Columbia
 Ambient Water Quality Criteria
   for Dissolved Oxygen, Water
 Clarity and Chlorophyll a for the
  Chesapeake Bay and Its Tidal
    Tributaries: 2010 Technical
 Support for Criteria Assessment
       Protocols Addendum

                May 2010

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  Ambient Water Quality Criteria for Dissolved
Oxygen, Water Clarity and Chlorophyll a for the
 Chesapeake Bay and Its Tidal Tributaries: 2010
   Technical Support for Criteria Assessment
               Protocols Addendum
                       May 2010
             U.S. Environmental Protection Agency
                       Region III
               Chesapeake Bay Program Office
                   Annapolis, Maryland

                          and

                       Region III
                 Water Protection Division
                 Philadelphia, Pennsylvania

                   in coordination with

                     Office of Water
               Office of Science and Technology
                    Washington, D.C.

                          and

                      The states of
               Delaware, Maryland, New York
                 Pennsylvania, Virginia and
           West Virginia and the District of Columbia

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                                       Contents
Acknowledgements	3

   I.     Introduction	5
          Literature cited	6
   II.     Designated Use Boundaries: Episodic Pycnocline Application and
          Expanded Designated Uses	 8
          Background	 8
          Revising a Procedural Anomaly in the Designated Use Delineation	9
             Identification of a Procedural Anomaly	9
             Episodic Pycnocline Criteria Assessment Protocol Modification	10
          Expanded Application of Deep-Water and Deep-Channel Designated Uses	11
             Review of Designated Use Definitions	11
             Mesohaline Segments Expanded Designated Uses	12
          Literature Cited	14

   III.    Biologically-based Reference Curve: Revisions to the Methodology and
          Applications	15
          Background	15
          Issues with Dissolved Oxygen Criteria Assessment with the Available  Previously
          Published Biologically-based Reference Curves	16
          Updates to Dissolved Oxygen Biologically-based reference Curve Derivation
          Methodology	17
             Restrict datasetto data collected beginning in 1996	18
             Use sequential 3-year time periods rather than single years	18
             Screening criteria sample size n>10	19
             Screening criteria standard deviation < 1.0	19
             Definition of healthy benthic macroinvertebrate reference community
             conditions	20
             Use Grand Score in computations involving fixed station data	20
          Summary of Recommendations	21
          Application of a Reference Curve for Open-Water 30-day Mean Dissolved Oxygen
          Criteria: Summer Season	23
          Application of a Reference Curve for Deep-Water Mean Dissolved Oxygen
          Criteria	23
          Application of a Reference Curve for Deep-Channel Instantaneous Minimum
          Dissolved  Oxygen Criterion	24
             Comparisons of Degraded Reference Benthic Communities with the Published
             Deep Channel Reference Curve    	25
             Rationale for Acceptable Exceedances of the Deep Channel  Instantaneous
             Minimum Dissolved Oxygen Criterion	26
          Assessment of Summer Season Dissolved Oxygen Criteria	27

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      Literature Cited	28

IV.   Revisions to the Chlorophyll a Criteria Assessment Methodology	31
      Background	31
      Review of the Current Chlorophyll a Criteria Attainment Assessment   Procedure:
      Method and Assumptions	32
      Chlorophyll a: Data Skewness, Log Transformation and the Seasonal Mean
      Calculation	34
          Log normal character of chlorophyll a data	34
          James River focused analyses of log transformed chlorophyll a data for
          normality	35
      Chlorophyll a criteria Assessment Protocol Refinements using Log-
      Transformations	36
      Implications of the Revised Assessment Protocol	37
      Literature Cited	38
Acronyms	41

Appendices

   A. IBI Sample Size and Standard Deviations on IBI Scoring When Screening
   Segments for Reference Community characterization	42

   B. Shape of the Biologically-based Reference Curve	46

   C. Derivation of the Deep-Water Biologically-Based Reference Curve	49

   D. History of EPA Guidance Regarding the Deep-Channel Reference Curve	52

   E. James River Chlorophyll a Data Normality Analysis Checking Normality of Log-
   Transformed Chlorophyll a Data	54

   F. SAS Computer Code for James River VA Chlorophyll a Normality Tests, Spring and
   Summer Season	57

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                           Acknowledgements
This fifth addendum to the EPA April 2003 publication of Ambient Water Quality Criteria for
Dissolved Oxygen,  Water Clarity and Chlorophyll a for the Chesapeake Bay and Its Tidal
Tributaries  (Regional Criteria Guidance)  was  developed  and  documented  through  the
collaborative efforts of members of the Chesapeake Bay Program's (CBP) Criteria Assessment
Protocols Workgroup and the Water Quality Goal Implementation Team.

PRINCIPAL AND CONTRIBUTING AUTHORS

The  following are  principal  and  contributing authors  of this addendum: Peter Tango, U.S.
Geological  Survey/Chesapeake  Bay Program Office; Jeni Keisman,  University  of Maryland
Center for Environmental  Science/Chesapeake Bay Program Office; Richard Batiuk, U.S. EPA
Region  3  Chesapeake Bay Program  Office;  Elgin Perry,  Statistical Consultant,  and Jackie
Johnson, Interstate Commission  on the Potomac River Basin/Chesapeake Bay Program Office.

CRITERIA ASSESSMENT PROTOCOL WORKGROUP
Peter Tango, Chair, U.S. Geological  Survey/Chesapeake Bay Program Office; Cheryl Atkinson,
U.S. EPA Region  3, Water  Protection  Division; Harry  Augustine,  Virginia Department of
Environmental Quality; Mark Barath, U.S.  EPA Region 3, Water Protection Division;  Tom
Barron, Pennsylvania  Department of Environmental Protection; Stephen Cioccia,  Virginia
Department  of  Environmental  Quality;  Richard  Eskin,   Maryland   Department   of  the
Environment; Sherm Garrison,  Maryland Department  of Natural Resources;  Darryl Glover,
Virginia  Department of  Environmental  Quality; Rick  Hoffman,  Virginia  Department of
Environmental  Quality;   Jackie Johnson,  Interstate Commission  on  the  Potomac  River
Basin/Chesapeake Bay Program Office;    Jeni Keisman, University  of Maryland Center for
Environmental Science/Chesapeake  Bay  Program  Office;  Susan McDowell, United  States
Environmental Protection Agency Region  III;  Larry  Merrill,  United  States Environmental
Protection Agency; Bruce Michael,  Maryland Department of  Natural Resources; Ken Moore,
Virginia Institute of Marine Science;  Shah Nawaz, District Department of the Environment;
Jennifer Palmore, Virginia Department  of Environmental  Quality;  Tom Parham, Maryland
Department of Natural Resources; Elgin Perry, Statistics Consultant; Charlie Poukish, Maryland
Department  of  the  Environment;  Tish  Robertson,  Virginia Department of Environmental
Quality; Matt Rowe, Maryland Department of the Environment; John  Schneider, Delaware
Department  of Natural Resources and Environmental Control; Gary  Shenk, United  States
Environmental Protection  Agency;  Donald  Smith,  Virginia Department of Environmental
Quality; Scott Stoner, New York State Department of Environmental Conservation; Matt Stover,
Maryland  Department  of  the Environment;  Bryant  Thomas,  Virginia  Department of
Environmental Quality;  Mark  Trice, Maryland Department of Natural Resources; David
Wolanski, Delaware Department of Natural Resources and Environmental Control.

WATER QUALITY GOAL IMPLEMENTATION TEAM
Dave  Hansen,   Co-Chair,  University  of  Delaware;  Robert  Koroncai,   Co-Chair,  U.S.
Environmental Protection  Agency Region  3 Water Protection Division; Katherine Antos,
Coordinator, Chesapeake Bay Program Office, U.S. Environmental Protection Agency; Victoria

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Kilbert, Staff, Chesapeake Research Consortium; Rachel Streusand, Staff, Chesapeake Research
Consortium; Rich Batiuk, U.S. Environmental Protection Agency Region 3 Chesapeake Bay
Program  Office; Steve Bieber,  Metropolitan Washington Council of Governments;  Patricia
Buckley,  Pennsylvania  Department  of  Environmental  Protection;  Collin Burrell,  District
Department of the Environment; Monir Chowdhury, District Department of the Environment;
Frank Coale,  University of Maryland; Lee Currey, Maryland Department of the Environment;
James Davis-Martin, Virginia Department of Conservation and Recreation; Chris Day,  U.S.
Environmental  Protection  Agency Region  3; Ron  Entringer,  New  York  Department  of
Environmental  Conservation; Richard  Eskin,  Maryland  Department  of the Environment;
Normand Goulet, Northern Virginia Regional Commission; Krista Grigg U.S. Navy; Mike Haire,
U.S. Environmental Protection Agency Office of Water;  Jeffrey Halka, Maryland Geological
Survey; Carlton Haywood, Interstate Commission on the Potomac River Basin; Dave Heicher,
Susquehanna  River  Basin Commission; Rick Hill, Virginia Department of Conservation and
Recreation; Beth Horsey, Maryland Department of Agriculture: Ruth Izraeli, U.S. Environmental
Protection Agency Region 2; Bill Keeling, Virginia Department of Conservation and Recreation;
John Kennedy,  Virginia Department  of Environmental Quality; Teresa Koon, West Virginia
Department of Environmental Protection; Felix Locicero, U.S. Environmental Protection Agency
Region 2; Charles Martin,  Virginia  Department of Environmental Quality;  Bruce  Michael,
Maryland  Department  of Natural Resources; Matt Monroe,  West  Virginia  Department  of
Agriculture;  Dave  Montali,  West Virginia  Department of Environmental Protection; russel
Morgan, U.S. Department of Agriculture Natural Resource Conservation Service; Matt Mullin,
Chesapeake  Bay Commission;  Kenn Pattison, Pennsylvania  Department of Environmental
Protection; Russ Perkinson, Virginia Department of Conservation and Recreation; Alan Pollock,
Virginia Department of Environmental Quality; Marel Raub, Chesapeake bay Commision; John
Rhoderick,  Maryland Department of Agriculture;  John Schneider, Delaware Department  of
Natural  Resources  and Environmental  Control;  Mohsin  Siddique,  D.C.  Water and Sewer
Authority; Jennifer  Sincock, U.S. Environmental Protection Agency Region 3; Randolph Sovic,
West Virginia Department of Environmental Protection; Tanya Spano, Metropolitan Washington
Council of Governments; Ann Swanson, Chesapeake Bay Commission; Jennifer Volk, Delaware
Department of Natural Resources and Environmental Control; Robert  Yowell, Pennsylvania
Department of Environmental Protection.

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                                     CHAPTER 1


                                 Introduction

In April 2003, the U.S. Environmental Protection Agency (EPA) published the Ambient Water
Quality Criteria for Dissolved Oxygen,  Water Clarity and Chlorophyll afar the Chesapeake Bay
and Its Tidal Tributaries which was the foundation document defining Chesapeake Bay water
quality criteria and  recommended implementation procedures for monitoring and assessment
(U.S.  EPA 2003a).   In October 2003, EPA published the  Technical Support Document far
Identification of Chesapeake Bay Designated Uses and Attainability which defined the five tidal
water designated uses to be protected through the published Bay water quality criteria (U.S. EPA
2003b):

    •   Migratory fish spawning and nursery habitat;
    •   Open-water fish and shellfish habitat;
    •   Deep-water seasonal fish and shellfish habitat;
    •   Deep-channel seasonal refuge habitat; and
    •   Shallow-water bay grass habitat.

A total of six addendum  documents have been  published by EPA since April 2003. Three
addenda  were published  documenting detailed  refinements to the  criteria  attainment  and
assessment procedures (U.S. EPA 2004a, 2007a, 2008) previously published in the original April
2003  Chesapeake Bay water  quality  criteria document (U.S. EPA  2003a). One addendum
published Chesapeake Bay numerical chlorophyll  a  criteria  (U.S.  EPA  2007b).  Another
addendum addressed detailed issues involving further delineation of tidal water designated uses
(U.S.  EPA 2004b) building from the original October 2003 tidal water designated uses document
(U.S.  EPA 2003b). Finally,  one addendum addressed refinements to the  Chesapeake  Bay
Program analytical segmentation schemes (U.S. EPA 2005) building from the original U.S. EPA
2004 document (U.S. EPA 2004c).

The detailed procedures for assessing attainment of the Chesapeake Bay water quality criteria
continued to be  advanced through  the  collective  EPA, States and District of Columbia
partnership efforts. These partners continue to develop and apply procedures that incorporate the
most   advanced   state-of-the-science,   magnitude,   frequency,  duration,  space  and  time
considerations with, as available,  biologically-based  reference  conditions  and cumulative
frequency distributions. As a rule, the best test of any new method or procedure is putting  it to
application with  partner involvement and stakeholder input. Through  the work of its  Criteria
Assessment Protocols Workgroup, the Chesapeake Bay Program partnership has an established
forum for resolving  issues, factoring in new scientific findings, and ensuring implementation of
consistent  bay-wide criteria  assessment  procedure development   and implementation.  The
Workgroup draws upon the talents and input from  state, federal, river basin  commission and
academic  partners as well  as local  government and municipal stakeholders.  This EPA 2010
Chesapeake Bay Criteria addendum provides previously undocumented features of the present
procedures as well as refinements and clarifications to the previously  published Chesapeake Bay
water quality criteria assessment procedures.

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Chapter 2 documents refinements to the procedures for defining Chesapeake Bay designated uses
and expands the application of the deep-water seasonal fish and shellfish designated use to two
Chesapeake Bay segments in Maryland's tidal waters.

Chapter 3 documents refinements  and additions to the  previously published procedures  for
deriving biologically-based reference  curves and  recommendations  for  their application  for
Chesapeake Bay dissolved oxygen criteria assessments.

Chapter 4 documents refinements and  provides recommendations for the procedures assessing
the previously published numerical Chesapeake Bay chlorophyll a criteria.

Appendices to these three  chapters provide more detailed documentation on derivation of the
recommended refined criteria assessment procedures.

This document represents the fifth formal addendum to the original 2003 Chesapeake Bay water
quality criteria document. As such readers should regard the sections in this document as new or
replacement chapters and appendices to the original published Bay Criteria report (U.S.  2003a).
The  criteria assessment procedures published  in this addendum also  replace and  otherwise
supersede similar criteria assessment procedures published in the 2004, 2007 and 2008 addenda
(U.S. EPA 2003a, 2004a, 2007a, 2007b, 2008). Publication of future addenda by EPA on behalf
of the Chesapeake Bay Program watershed jurisdictional partners is likely as continued scientific
research and management  applications reveal new insights and knowledge that should be
incorporated into  revisions  of state water quality standards  regulations  in upcoming triennial
reviews.
                                LITERATURE CITED

U.S. Environmental Protection Agency. 2003a. Ambient Water Quality Criteria for Dissolved
Oxygen, Water Clarity and Chlorophyll a for  the Chesapeake Bay and Its  Tidal Tributaries
(Regional Criteria Guidance). April 2003.  EPA 903-R-03-002. Region III Chesapeake  Bay
Program Office, Annapolis, MD.

U.S. Environmental Protection Agency. 2003b. Technical Support Document for Identification of
Chesapeake Bay Designated Uses and Attainability. October 2003. EPA 903-R-03-004. Region
III Chesapeake Bay Program Office, Annapolis, MD.

U.S. Environmental Protection Agency. 2004a. Ambient Water Quality Criteria for Dissolved
Oxygen, Water Clarity and Chlorophyll a for the Chesapeake Bay and Its Tidal Tributaries -
2004 Addendum.  October 2004. EPA 903-R-04-005. Region III Chesapeake Bay  Program
Office, Annapolis, MD.

U.S. Environmental Protection Agency. 2004b. Technical Support Document for Identification of
Chesapeake Bay Designated Uses and Attainability - 2004 Addendum. October 2004. EPA 903-
R-04-006. Region III Chesapeake Bay Program Office, Annapolis, MD.

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U.S.  Environmental  Protection  Agency.   2004c.  Chesapeake Bay  Program Analytical
Segmentation Scheme: Revisions, Decisions and Rationales 1983-2003. October 2004. EPA 903-
R-04-008. Region III Chesapeake Bay Program Office, Annapolis, MD.

U.S.  Environmental  Protection   Agency.  2005.  Chesapeake  Bay  Program Analytical
Segmentation Scheme: Revisions, Decisions and Rationales  1983-2003.  2005 Addendum.
December 2005.  EPA 903-R-05-004. Region III Chesapeake Bay Program Office, Annapolis,
MD.

U.S. Environmental Protection Agency. 2007a. Ambient Water  Quality Criteria for Dissolved
Oxygen, Water Clarity and Chlorophyll a for the Chesapeake Bay and Its Tidal Tributaries -
2007 Addendum.  July 2007. EPA 903-R-07-003. Region III Chesapeake Bay Program Office,
Annapolis, MD.

U.S. Environmental Protection Agency. 2007b. Ambient Water  Quality Criteria for Dissolved
Oxygen, Water Clarity and Chlorophyll a for the Chesapeake Bay and Its Tidal Tributaries -
Chlorophyll a Addendum. October 2007. EPA 903-R-07-005.  Region III Chesapeake Bay
Program Office, Annapolis, MD.

U.S. Environmental Protection Agency.  2008. Ambient Water Quality Criteria for Dissolved
Oxygen,  Water  Clarity  and  Chlorophyll a  for  the  Chesapeake  Bay and  Its  Tidal
Tributaries-2008 Technical Support for Criteria Assessment Protocols Addendum. September
2008. EPA 903-R-08-001. Region III Chesapeake Bay Program Office, Annapolis, MD.

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                                    CHAPTER 2


     Designated  Use Boundaries:  Episodic Pycnocline

         Application  and Expanded  Designated Uses

                                  BACKGROUND

In the 2003  Ambient Water Quality  Criteria for Dissolved Oxygen,  Water Clarity  and
Chlorophyll a for Chesapeake Bay and Its  Tidal Tributaries, EPA  defined five tidal water
habitats as designated uses providing the context for setting protective Chesapeake Bay water
quality criteria  (U.S. EPA 2003). Detailed  dissolved  oxygen criteria were  established for
Chesapeake Bay and its tidal tributaries  and embayments  tailored  to each  designated use
accounting for its variations in space and time.  EPA has published and Delaware, Maryland,
Virginia and the District of Columbia have adopted into their state's water quality standards
regulations dissolved oxygen criteria protective of the  published migratory spawning, open-
water, deep-water and deep-channel designated uses. These dissolved oxygen criteria include 30-
day,  7-day and 1-day means along with  instantaneous  minima as needed to  protect  various
species and life stages within the designated uses (U.S. EPA 2003).

Since the Chesapeake Bay dissolved oxygen  criteria were published  in 2003, refinements and
updates to the criteria attainment assessment methodologies have been published. Most recently,
the refined and expanded dissolved oxygen criteria assessment methodologies documented in
Chapter 3 and  associated  appendices of the Ambient Water  Quality Criteria for Dissolved
Oxygen, Water Clarity and Chlorophyll a for the  Chesapeake Bay and its Tidal Tributaries -
2008  Technical Support for Criteria  Protocols Addendum, replaced  the   methodologies
previously published by EPA (U.S. EPA 2008).

Critical to the dissolved oxygen criteria assessments are the pycnocline delineations defining the
timing and vertical position of the open-water,  deep-water and  deep-channel  designated use
boundaries. The standardized method for calculating upper and lower boundaries of pycnoclines
was originally published in Ambient Water Quality Criteria for Dissolved Oxygen, Water Clarity
and Chlorophyll a for the Chesapeake Bay and its Tidal Tributaries - 2004 Addendum (U.S.
EPA 2004a). U.S. EPA (2008), on pages  15-18 together with its Appendix  A, provide a review
of, and step by step details associated with, calculating upper and lower pycnoclines which, in
turn,  delineate  the  vertical boundaries  for  the  open-water,  deep-water and  deep-channel
designated uses.

The following outline lays out the assessment protocol steps for the 30-day  mean criteria (open-
water and deep-water dissolved oxygen criteria (see  U.S. EPA 2008, Appendix A for details):

                1) Compiling and formatting the data set
                2) Interpolation of water quality monitoring data
                       2.1 Vertical interpolation
                       2.2 Horizontal interpolation
                       2.3 30-day average interpolation by month

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                        2.4 Apportioning results by designated use
                        2.5 Water quality criteria assessment, attainment and violations

Step 2.4 above, carried forward the Step 4-Pointwise Compliance considerations of a statistical
decision-making framework originally published in U.S. EPA 2007a Chapter II: Refinements to
Chesapeake Bay Water Quality Criteria Assessment Methodology (pp. 17-18) and revisited in
U.S. EPA 2008 (Appendix A). This section on pointwise compliance states:

       "While interpolation allows for standardization of many types of data, pointwise
       attainment allows  for standardization of many criteria. Because  attainment is
       determined at moments  in time and  points in space, it is possible  to vary the
       criterion in time and space.  If different levels of a water quality criterion are
       acceptable in different  seasons,  then the criterion can vary  seasonally.  It is
       possible to implement different criteria over space for a segment that  bridges, for
       example, oligohaline and mesohaline, salinity zones. It might even be possible to
       let the criterion be a continuous function  of some ancillary variable  such as
       temperature or salinity,  although this situation requires that such data exist for
       every interpolator cell. The  only requirement is that the  final  attainment
       determination be "yes" or "no" for each interpolator cell."

The implicit assumption of the Chesapeake Bay partners was that if no pycnoclines were found
for a particular sampling  event then the open-water designated use and its respective dissolved
oxygen criteria were being applied, i.e. that water column dynamics including "episodic
pycnoclines" were accounted for as part of the  criteria assessment computations. The U.S. EPA
Chesapeake Bay Program Office's  criteria assessment computer code,  however, applied  the
long-term average pycnocline depth(s) to those water quality monitoring cruise sampling events
when no pycnocline was found for those 13 segments, identified in U.S. EPA 2004 where deep-
water and/or deep channel designated uses applied during the  June-September time period.
Therefore, under special cases, on the basis of pre-determined characterization, there were errors
in designated use classification.

REVISING A PROCEDURAL ANOMALY IN THE DESIGNATED USE DELINEATION

Identification of a Procedural Anomaly

During 2009, a procedural anomaly was discovered between EPA published dissolved oxygen
criteria assessment protocols through 2008 for pycnocline delineation that defined the boundaries
for the open-water, deep-water  and/or  deep-channel designated uses  and  the  assessment
procedures  as  defined in the  criteria  assessment computer  code  developed by the EPA
Chesapeake Bay Program Office  and used by the  states  and the District.  The published
procedures  set forth that attainment is determined at moments in time and space given that the
designated uses, their boundaries and the applicable dissolved oxygen criteria will also vary in
time and space (U.S. EPA 2003). U.S. EPA (2008) published details  of the computations for
identifying  pycnoclines where they exist  on  a  water quality monitoring cruise-by-cruise basis.
EPA also identified 13 Chesapeake Bay segments where  deep-water and (or) deep-channel
designated uses applied during the June-September time period (U.S. EPA 2004, page 5, Figure

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II-2, and Table II-1 below). The remaining tidal segments in Chesapeake Bay were characterized
as having the open-water designated use year-round.
Table II-1 Chesapeake Bay segments with assigned designated uses.
 Designated
 Use
Segment Code
Location
  Deep Water
   and Deep
    Channel
CB3MH
CB4MH
CB5MH
CHSMH
EASMH
PATMH
POTMH
RPPMH (portion S of UTM Y = 4185000)
Chesapeake Bay Mainstem
Chesapeake Bay Mainstem
Chesapeake Bay Mainstem
Chester Paver
Eastern Bay
Patapsco River
Lower Potomac River
Lower Rappahannock River
  Deep Water
     Only
CB6PH (portion north of UTM Y 4145)
CB7PH (portion N/NW of UTM Y = UTM X + 3752745)
PAXMH
SBEMH
YRKPH
Chesapeake Bay Mainstem
Chesapeake Bay Mainstem
Lower Patuxent River
South Branch Elizabeth River
Lower York River
Source: U.S. EPA 2004

The  EPA Chesapeake Bay Program Office acknowledged this computation code improperly
imposed pycnocline presence at times and places where none was found. Such applications of a
long-term mean pycnocline instead of no  pycnocline were,  therefore,  incorrectly  applying
dissolved  oxygen criteria  assessments in such  situations.  The EPA published procedures,  as
described below, allow for the presence of episodic pycnoclines.

Episodic Pycnoclines Criteria Assessment Protocols Modification

The dissolved oxygen criteria assessment methodology is now clarified to specifically allow the
deep-water and deep-channel designated  uses to  occur "episodically" for those 13 segments that
have been identified as having deep-water and (or) deep-channel designated uses (see Table II-1
in U.S. EPA 2004) When a pycnocline  is observed during the tidal water quality monitoring
cruise within one of the 13 segments during June  1 through September 30, the deep-water and
(or) deep-channel designated uses exist  and their respective numeric dissolved oxygen criteria
are applied to those uses. When no pycnocline is  observed, the open-water designated use applies
to the entire water column. By definition, this approach eliminates the  default use of long term
pycnocline average when no pycnocline is observed.

Reassessment of previous dissolved oxygen assessments  by EPA and its state and District
partners showed only small changes in  Chesapeake Bay  dissolved oxygen  criteria attainment
results over time. Times and places where no pycnocline could be defined for summer season
among the 13 Chesapeake Bay segments with previously defined deep-water and deep-channel
designated uses were shown to be rare events.
                                                                                     10

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        EXPANDED APPLICATION OF DEEP-WATER AND DEEP-CHANNEL
                                 DESIGNATED USES

A total of 13 Chesapeake Bay segments characterized with deep-water and deep-channel
designated uses were published in U.S. EPA 2004 (Table II-l). In a number of segments
classified as having the open-water designated use only applied year-round, dissolved oxygen
criteria assessments through time provided evidence of persistent criteria non-attainment. In a
select set of these same Chesapeake Bay segments, results from numerous Chesapeake Bay
water quality/sediment transport model scenarios, simulating dissolved oxygen concentrations
across a wide range of nutrient load reductions, suggested lack of dissolved oxygen responses to
nutrient load reductions due to physical constraints to re-oxygenation. Segments not previously
classified with the deep-water and (or) deep-channel designated uses in mesohaline salinities but
showing both stratification (presence of a pycnocline) and persistent dissolved oxygen criteria
non-attainment were reviewed for possible expanded application  of deep-water and deep-channel
designated uses.

Review of Designated Use Definitions

The 2003 Ambient Water Quality Criteria for Dissolved Oxygen , Water Clarity and Chlorophyll
afar Chesapeake Bay and Its Tidal Tributaries highlights two relevant guidelines-stratification
(presence of pycnoclines) and evidence of a physical barrier restricting reoxygenation-for
determining the need to apply the deep-water and (or) deep-channel designated uses (U.S. EPA
2003). Specifically, the following are published definitions for determining when and where the
open-water, deep-water and (or) deep-channel designated uses apply within Chesapeake Bay
tidal waters:

       Open-Water Designated Use
       "If the presence of a pycnocline prevents oxygen replenishment, the open-water
       fish and shellfish designated use extends only as far as the upper boundary of the
       pycnocline. If a pycnocline exists but other physical circulation patterns (such as
       the inflow of oxygen-rich oceanic bottom waters) provide oxygen replenishment
       to the  deep waters, the open-water fish and shellfish designated use extends to the
       bottom water-sediment interface." U.S. EPA 2003, Appendix A, page A-6.

       (Also  see U.S.  EPA 2007, pages 37-38, Dissolved oxygen assessments in shallow
       versus open waters, for details regarding the open water designated use definition
       beyond vertical water column structure.)

       Deep-Water Designated Use
       "Tidally influenced waters located between the measured depths  of the upper and
       lower  boundaries of the pycnocline, where a measured pycnocline is present and
       presents a barrier to oxygen replenishment from June 1 to  September 30...the
       deep-water designated use extends from the upper boundary of the pycnocline
       down  to the  sediment/water interface  at the bottom, where a lower boundary of
       the pycnocline  is not calculated." U.S. EPA 2003, Appendix A, page A-6.
                                                                                     11

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       Deep-Channel Designated Use
       "Tidally influenced waters at depths greater than the measured lower boundary of
       the pycnocline in isolated deep channels." U.S. EPA 2003, Appendix A, page A-
       6.

Mesohaline Segments Expanded Designated Uses

Using the time period 1991-20001,  depth profiles of change in density and dissolved oxygen
concentrations from the Chesapeake Bay Water Quality Monitoring Program2 were reviewed for
both evidence of stratification and prevention of re-oxygenation. Chesapeake Bay segments in
the mesohaline salinity zone, not previously classified with deep-water and (or) deep-channel
designated uses, were evaluated for evidence of stratification and persistent dissolved oxygen
criteria  non-attainment under  a range of different Chesapeake Bay water quality/sediment
transport model loading scenarios. Ten segments meeting these characteristics were identified in
Maryland and Virginia's Chesapeake Bay tidal waters (Table II-2).

Table 11-2. Ten  Chesapeake Bay segments in  the mesohaline salinity zone of Maryland and
Virginia's Chesapeake  Bay  tidal  waters reviewed  for  possible expanded  designated use
classifications.
          Chesapeake Bay Segment
     Tidal Water Body
                 MAGMH
                  SOUMH
                  EBEMH
                 WBEMH
                  CRRMH
                  FSBMH
                  WICMH
                  SEVMH
                 WSTMH
                 YRKMH
      Magothy River
        South River
East Branch Elizabeth River
West Branch Elizabeth River
     Corrottoman River
        Fishing Bay
      Wicomico River
        Severn River
        West River
        York River
Only the South River (SOUMH) and Magothy River (MAGMH) segments met the deep-water
designated use definition originally described in U.S. EPA 2003 where a measured pycnocline
was present and  presented a barrier to  oxygen replenishment during the period  June 1 to
September 30.

In the South River segment, 39 of 43 depth profiles (91%) had an upper pycnocline and 19 of 43
depth profiles (44%) had a lower pycnocline. In the Magothy River, 16  of 40 depth profiles
(40%)  had an upper pycnocline and 0 of 40 depth profiles  (0%) had  a lower pycnocline.
Evaluation of the Chesapeake Bay water quality/sediment transport model scenario results for
both segments showed depression of dissolved oxygen concentrations with increasing depth
suggesting a physical mixing constraint on re-oxygenation due to stratification.
1 These years of Chesapeake Bay Water Quality Monitoring Program data were selected to be consistent with the
hydrologic period for management application of the Chesapeake Bay Water Quality/Sediment Transport Model.
2 www.chesapeakebay.net
                                                                                     12

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In the presence of a pycnocline, the deep-water designated use will also apply to the Magothy
River and South River mesohaline segments in the June 1 through September 30 time period.
The application of the deep-water designated use to these two segments is fully consistent with
previously published procedures which called for:

   1.   the physical exchange of higher oxygenated waters from the upper water-column is much
       reduced by density stratification, and
   2.   pycnocline waters are not reoxygenated by riverine or oceanic bottom waters

in order to apply the deep-water designated use (U.S. EPA 2003).

Previously, such  segments  including the deep-water designated use were only thought to be
"located principally in the river channel at the lower reaches of the major rivers and along the
spine  of the middle mainstem of the Bay"  (U.S. EPA 2003).  These analyses conducted in
support of the  development of this addendum have demonstrated the deep-water designated use
can occur in smaller  tidal  tributaries segments receiving  limited freshwater  flow from their
surrounding watershed.

Given the South River segment has a lower pycnocline and  19 of 43 depth profiles (44%) over
the 1991-2000 data record, consideration was given to whether a deep-channel designated use
should apply to this segment as well as a deep-water designated use. The published procedures
for delineating a deep-channel designated use included:

   1.   The very  deep water-column and adjacent bottom  surficial  sediment  habitats  located
       principally in the river channel at the lower reaches of the major river and along the spine
       of the middle mainstem of the bay;
   2.   At depths below which seasonal anoxic to severe hypoxic conditions routinely set in and
       persist for  extended periods of time under current conditions; and
   3.   At depths greater than the lower boundary of the pycnocline (U.S. EPA 2003).

The South River segment does not contain a  "very deep water-column" given a total maximum
depth  of 5 meters and the segment does not  have conditions where "seasonal anoxic to severe
hypoxic conditions routinely set  in and persist for extended periods of time under  current
conditions". Therefore, even in the presence of a lower pycnocline, a deep-channel designated
use will not be applied to this segment.

The initial review of stratification  and dissolved oxygen data from the eight remaining segments
identified in Table 11-2 did not provide immediate evidence of where stratification appeared to be
limiting oxygen replenishment. A more in-depth review of water column stratification conditions
and identification  of any needs for further adjustments to the applicable designated uses for the
remaining segments is planned for completion prior to the 2012 303(d) listing cycle.
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                                LITERATURE CITED

U.S. Environmental Protection Agency. 2003. Ambient Water Quality Criteria for Dissolved
Oxygen, Water Clarity and Chlorophyll a for  Chesapeake Bay and Its Tidal Tributaries April
2003 (Regional Criteria Guidance). EPA 903-R-03-002. Region III Chesapeake Bay Program
Office, Annapolis, Maryland.

U.S. Environmental Protection Agency. 2004.  Technical Support Document for Identification of
Chesapeake Bay Designated Uses and Attainability-2004 Addendum. October 2004. EPA 903-R-
04-006, Chesapeake Bay Program Office, Annapolis, MD.

U.S. Environmental Protection Agency. 2007. Ambient Water Quality Criteria for Dissolved
Oxygen, Water Clarity and Chlorophyll a for the Chesapeake Bay and Its Tidal Tributaries -
2007 Addendum. July 2007. EPA 903-R-07-003. Region III Chesapeake Bay Program Office,
Annapolis, MD.

U.S. Environmental Protection Agency. 2008. Ambient Water Quality Criteria for Dissolved
Oxygen, Water Clarity and Chlorophyll a for the Chesapeake Bay and its Tidal Tributaries -
2008 Technical Support for Criteria Protocols Addendum. September 2008. EPA 903-R-08-001.
Region III Chesapeake Bay Program Office, Annapolis, MD.
                                                                                   14

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                                    CHAPTER 3


 Biologically-based Reference Curves: Revisions to the

                   Methodology and Applications

                                 BACKGROUND

The published dissolved oxygen criteria assessment methodology currently used for assessing
Chesapeake Bay water quality criteria attainment involves the  use of cumulative frequency
distribution (CFD) curves in a two-dimensional space of percent time and percent space (U.S.
EPA 2003). Minimum concentrations of dissolved oxygen must be present to support species and
their various  life stages requiring protection. Dissolved  oxygen  criteria provide  threshold
conditions established for the designated uses such that water quality conditions that exceed this
threshold are considered impaired.

However, it is recognized that all water quality parameters are inherently variable in space and
time. There will be small regions that persistently exceed the threshold due to poor flushing or
other natural conditions.  The Chesapeake Bay dissolved oxygen criteria have several durations
reflecting the various tolerances of different life stages and effects (U.S. EPA 2003, 2008). Small
regions or time periods of degraded condition should not lead to a degraded assessment for the
segment (U.S. EPA 2003). Recognition that ephemeral exceedances of the threshold in both time
and space do not  represent persistent impairment  of the segment leads  to  an assessment
methodology  that allows these conditions  to be  classed  as acceptable while  conditions of
persistent and wide spread impaired condition will be flagged as  unacceptable. (E. Perry, Pers.
Comm. 2005).

During an independent scientific peer review of the EPA published  CFD procedures, reviewers
raised specific concerns  about the method for deriving the  biological reference curves  (STAC
2006).  At the time, there were no apparent solutions to resolve the concerns that were raised.
However,  during recent  application of  criteria  assessment procedures to model simulated
outputs, evaluation  of the  resultant model outputs put  the  spotlight  back  on  the  criteria
assessment process and the underlying biological reference curve methodology.

Work by the EPA Chesapeake Bay Program Office and its partners suggested that application of
the currently published application of the Benthic-Index of Biotic Integrity (B-IBI) (Weisberg et
al. 1997)  did not  accurately  distinguish between healthy and degraded communities  with
corresponding distinct sets of dissolved oxygen violations. EPA Chesapeake Bay Program Office
analysts and partners worked with recognized Chesapeake Bay benthic community experts2 to
revise the published methods  for identifying "healthy"  and "degraded" benthic communities.
During this process, it was determined that the B-IBI provides a robust delineation of healthy and
degraded benthic communities with corresponding distinct dissolved  oxygen violation rates.
! Dr. Dan Dauer, Old Dominion University and Dr. Roberto Llanso, Versar, Inc.


                                                                                   15

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Using the newly delineated "healthy" and "degraded" benthic communities, EPA Chesapeake
Bay Program Office analysts worked to produce a set of revised biological reference curves that
minimize the error in distinguishing between "healthy" and "degraded" segments. In this chapter
and  its  associated  appendices,  updates to  the  methodology  involving  development of
biologically-based reference curves with Chesapeake Bay benthic macroinvertebrate monitoring
program data are provided. Further, directions on application of reference curves for open-water,
deep-water and deep-channel designated uses are provided  for completing the Chesapeake Bay
dissolved oxygen criteria attainment computations.

     ISSUES WITH DISSOLVED OXYGEN CRITERIA ASSESSMENT WITH THE
    PREVIOUSLY PUBLISHED BIOLOGICALLY-BASED REFERENCE CURVES

The current published method for assessing dissolved oxygen (DO) impairments in Chesapeake
Bay incorporates the use of a cumulative  frequency distribution  as the final step of assessment
(U.S. EPA 2003).  In this step, a set of DO violation rates for a particular segment-designated use
(e.g. "CB4MH Deep  Water") are plotted as  a cumulative frequency distribution (CFD) and
compared to  a "biological reference curve" comprising a cumulative  frequency distribution of
"acceptable violation rates" of the DO criteria.  If the assessment curve  exceeds, at any point, the
reference CFD, then the given segment is considered "impaired (Figure  III-l).
                              10
                               9
                            CD  '

                            CD  6
a  s
   4
   3
   2
                               1
                                               CFD Curve
                 Area of Criteria
                  Exceedence
                                                 Area of Allowable
                                                     Criteria
                                                   Exceedence
                                 01   23456789  10
                                          Percent of Space
Figure III-l. Conceptual graph illustrating the CFD assessment procedure.  The red line is an
example of a hypothetical "healthy" assessment curve; the blue line is the hypothetical reference
curve.

It has been recognized, however, that by combining violation rates from all healthy areas into
one  biologically-based  reference  curve,  we create  a  curve that theoretically  represents
approximately the median of all curves included.  Thus, a large percentage  of the presumably
"acceptable" violation rate CFDs that were pooled in order to generate the  biologically-based
reference curve may fail an assessment conducted against that same biologically-based reference
curve. A more  detailed evaluation confirmed this concern. In Figure III-2 below, the CFD for
                                                                                     16

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CB3MH Deep Water 1987, a segment/designated use considered having a healthy B-IBI for that
year, whose acceptable violation rates were included in the generation of the biological reference
curve, fails assessment by that same biological reference curve.
                                  Curve of all acceotable violation rate CFDs
                                           B3MH 1987
                                          0.4        0.6

                                        Percent Space
                                                            0.8
Figure III-2.  An  example of a 30-day  mean  deep-water dissolved oxygen criteria  and the
violation expressed by a healthy segment (CB3MH  1987) curve used  in deriving the 30-day
mean criterion biologically-based reference curve.

Further analyses revealed that the biological reference curves used for the deep-water and deep-
channel dissolved oxygen criteria  attainment  assessments fail the majority of  supposedly
"healthy" segment-years used to construct those same curves.

As described in U.S. EPA 2003, the preferred methodology for defining the reference curve is to
determine levels of allowable  violation  based on the  demonstrated tolerance of the  living
resources  for  whose protection the  water quality  criteria were  designed. Benthic habitat
assessments were  conducted with the updated methodology,  which is described below, for
assessing the  appropriateness  of biologically-based  reference  curves  as indicators of  water
quality conditions.
    UPDATES TO DISSOLVED OXYGEN BIOLOGICALLY-BASED REFERENCE
                       CURVE DERIVATION METHODOLOGY

Based  on the  findings described above, the following revisions  are  recommended  to  the
methodology for categorizing benthic communities as "healthy" for the purposes of providing a
reference for allowable frequency of dissolved oxygen criteria exceedance. The intent of these
revisions is to improve the accuracy with which benthic communities are categorized as healthy.

Revisions to the previously published methodology for developing dissolved oxygen biological
reference curves include:
    1) Restriction of the reference dataset to data collected beginning in 1996;
    2) expansion of time period for  classifying benthic  community health  from  1 year to
       sequential 3-year time periods;
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   3)  restriction of reference segment-periods to those for which at least 10 observations are
       available;
   4)  refined definition of a "healthy" benthic community as one for which the mean B-IBI
       score is at least 3.0;
   5)  the standard deviation of the mean is less than 1.0; and
   6)  use Grand Score in computations involving fixed data.

The rationale underlying each of these six modifications is described in further detail below.

Restrict Dataset to Data Collected Beginning in 1996

Criteria violation results of dissolved oxygen criteria attainment assessments are compared  with
a reference CFD curve (e.g., standard 10% reference or biologically-based reference CFD curve),
representing allowable amounts of criteria exceedance in a healthy habitat.  When an appropriate
biological reference community is identified and sufficient  data  are deemed available, a
biological reference curve of acceptable  percent exceedance  is generated using  a CFD  of
violation rates for "healthy"  biological  communities in that designated use. A review of the
plotting methodology is provided in U.S. EPA 2008 (see Appendix A).

Historically, the benthic monitoring work of the Chesapeake Bay Benthic Monitoring Program
consisted  of fixed station monitoring  with sampling usually taking place in August  and
September (Chesapeake Bay Program 1989). The sampling design was primarily intended to
assess long-term trends in living resources over decadal, annual and seasonal time scales.

Derivation of the original dissolved oxygen biologically-based reference  curves relied on the
1985-2005 Chesapeake Bay benthic monitoring program dataset in order to take advantage of the
full two decades of monitoring results. However, data  collection  methods  have  undergone
revision during the 21 years of monitoring. In 1996, a stratified random sampling component was
added to the benthic monitoring program in  order to provide confidence limits on estimates of
impaired waters in  Chesapeake Bay. In order to ensure adequate spatial resolution of benthic
community health,  STAC  (2009),  in  accordance with  recognized  Chesapeake Bay benthic
community experts, recommended  truncating the  reference data set  to start in 1996 when the
updated sampling procedures were  established. The data period was extended one year to 2006
to include the most recently available data. The use of the 1996-2006 Chesapeake Bay benthic
monitoring program data set is an update to previously published  methods (U.S. EPA 2007,
Chapter 4). The recommended data  set represents a consistent period of improved assessments of
Chesapeake Bay health condition.

Use Sequential 3-year Time Periods Rather Than Single Years

The biologically-based reference curve derivation methodology, as outlined in U.S.  EPA 2007
(see Chapter 4), used single year assessments to determine the health of the benthic community
for the  purposes of identifying acceptable dissolved oxygen criteria exceedances.   However,
dissolved  oxygen criteria assessments are conducted on sequential 3-year time frames for  each
segment (U.S. EPA 2003); two year time steps are used in reporting for 303d listing cycles  (e.g.
the 2008 303d listing  cycle used 2004-2006  data, the 2010 303d listing cycle used  2006-2008
                                                                                     18

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data) while benthic community assessments are conducted annually with annual time steps for a
variety  of purposes (e.g.  indicator reporting  for the  Chesapeake  Bay  Barometer).  Using
sequential 3-year time periods to classify benthic community health, advancing the data in one
year time steps (e.g. 1996-1998, 1997-1999, etc.), brings the reference  community identification
method into better alignment with the dissolved oxygen criteria assessment protocols for which
reference communities are being identified. This modification addresses a concern raised by the
Chesapeake Bay Program's Scientific and Technical Advisory Committee (STAC) review of the
CFD approach (STAC 2006) which noted that sample sizes for reference and assessed conditions
should be made  similar to reduce the effect of sample size bias on the shape of the CFD. The
combination of a segment and sequential 3-year assessment time periods is hereafter referred  to
as a "segment period".

Screening criteria:  Sample Size > 10

Keller and Cavallaro (2008) reported that  listing decisions on the U.S. Clean Water Act 303a
listing impairments of surface waters by states were often based on insufficient data, or that data
were not sufficiently representative of temporal and spatial conditions  for the water body being
assessed. Llanso et  al. (2009), however, require a minimum sample size of n >  10 for habitat
health assessments using the Chesapeake Bay B-IBI. The EPA Chesapeake Bay Program Office
and its partners examined the effects of relaxing the data screening criteria to accept segment-
period combinations with sample size >  8 to increase  the number  of "healthy" segment-periods
available for reference community analysis.

The decision to eliminate  segment-periods with  fewer than  10  observations  was  based on
analyses by EPA Chesapeake Bay Program Office and its partners, which showed that fewer
than  10  observations  weakened the ability of the reference CFD to appropriately classify
segments. Llanso et al  (2009) confirmed Keller and  Cavallaro (2008)'s findings  regarding
sample size and  temporal and spatial distribution. They found that analysis of Chesapeake Bay
segments with less than 10 samples produced "inconclusive results relative to the  (U.S. EPA)
listing process."  In their review of the proposed methodology, STAC  (2009) determined that a
minimum sample size of 10 is reasonable and has been applied elsewhere (Alden et al.  2002).
Further details on the sample size analyses are available in Appendix A.

Screening Criteria: Standard Deviation < 1.0

The EPA Chesapeake Bay Program Office  and its partners examined the isolated  and combined
effects of relaxing the  data screening criteria to accept segment-periods with fewer samples (n >
8 instead of 10)  and/or expanding the standard deviation criteria surrounding the B-IBI  results
from <1.0 to < 1.2 in order to increase the number of "healthy"  segment-periods for analysis.
The relaxation of both the  sample size and the standard deviation criteria (see  Scenario D  in
Appendix A) increases the number of segment-periods classified as "healthy" from 10 to 16.
However, four of these additional CFD  curves extend into "degraded" CFD  space to a  degree
that calls into  question  the  accuracy  of  their classification as  healthy  (see Figure  A-3  in
Appendix A).  Defining healthy benthic  communities  for deriving a benthic community based
biological reference curve, therefore, relies on  sample size n > 10 with a  standard deviation <
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1.0. Further details of the sample size and standard deviation analyses are available in Appendix
A.

Definition of Healthy Benthic Macroinvertebrate Reference Community Conditions

The methodology described by U.S. EPA (2007, 2008) defined healthy segments (with respect to
benthic communities) as those with a minimum B-IBI score > 3.0.  However, no  sample size
restriction was introduced. As a result, a large segment could contain a single B-IBI score, and if
that single  score exceeded 3, then the segment was classified as healthy.  The likelihood of a
degraded segment containing 10 B-IBI scores (in  any given  3 years) all of which  are > 3.0 is
small.   Furthermore, benthic community experts  (Llanso  et al. 2009) have more commonly
defined a healthy community as one with a sample mean > 3.0, given an adequately large sample
size and small variance.  Thus, the EPA Chesapeake Bay Program Office  and its partners now
define  "healthy" benthic reference communities as those with an average B-IBI score > 3.0 and
standard deviation (SD) < 1.0.  STAC (2009) supported the use of "healthy"  benthic reference
communities defined by those with an average B-IBI score >  3.0, rather than a minimum, and a
standard deviation (SD) < 1.0,  (n > 10). A degraded benthic community is  defined as having an
average B-IBI score < 3.0 with a standard deviation < 1.0, (n > 10).

The methodological refinements described above led to findings that provide ongoing support of
the need for a hyperbolic curve that distributes allowable violations in CFD  space, as do both the
new  deep-water biologically-based reference  curve described below  and  the default   10%
reference curve described in U.S. EPA (2007).  A more in-depth discussion of the shape of the
reference curve with respect to "healthy" and "degraded" CFD-space can be found in Appendix
B.

STAC  (2009) recommendations suggested that based  on the assumptions of normality, the
standard deviation  criterion  applied  when  classifying  a  healthy  benthic  community  could
alternatively be expressed as "no more than 16% of the sample observations should have a  score
less than 2.0". This is a one-sided version of the screening criterion, and addresses concerns that
clearly  healthy segments with high  variance  could  be excluded  from the analyses.  EPA
Chesapeake Bay Program Office staff conducted an exploratory  analysis to classify benthic
communities using the following benthic community classification rules:

   1)  average B-IBI score > 3.0 with no more than 16% of sample observations < 2.0 (n > 10)
       defines a healthy benthic community, and
   2)  average B-IBI score < 3.0 with no more 16% of sample observations > 4.0 B-IBI score,
       (n > 10) defines a degraded benthic community.

Results using these  revised  classification  rules were consistent with the results  of the
biologically-based reference curve derivation methodology outlined in this chapter.

Use Grand Score in Computations Involving Fixed Station  Data

The 1996-2006 Chesapeake Bay B-IBI sample results consist of both fixed station  and random
station data. These data are combined in the analyses but have different scoring categories within
the CIMS database.
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Specifically, for the "fixed station" samples both "total_score" and "grand_score" records are
reported within the Chesapeake Bay Information Management System (CIMS)3. "Total_score"
records  are  replicate measurements of the same  community event;  the  average of these is
reported as the "grand_score." The Chesapeake Bay benthic  experts (R.  Llanso, Versar Inc.,
Pers. Comm. 2009) recommended using the "grand_score" in the Chesapeake Bay  dissolved
oxygen criteria assessment analyses to avoid errors not accounting for the  replicate results of a
sampling event. By comparison, random  station records in the CIMS database report only a
"total score" as the sampling event B-IBI measure; no "grand scores" will be found associated
with random station data records.

                        SUMMARY OF RECOMMENDATIONS

Based  on  the  findings  of  the  analyses   and  in  accordance  with  the  STAC  (2009)
recommendations,  Table III-l  summarizes the revisions  to  the  methodology  for identifying
dissolved oxygen biologically-based reference  curves for Chesapeake Bay water quality criteria
attainment assessments (Table III-l).
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Table  III-l.  Chesapeake  Bay  dissolved  oxygen  criteria  biologically-based reference  curve
derivation recommendations.
 U.S. EPA 2007, 2008 Addenda
2010 Addendum
 Obtain dataset of all benthic index of biotic integrity (B-IBI) scores for
 time period 1985-2005
Restrict dataset to data starting in
1996; for random station data use
'total score' and for fixed station
samples use "grand score" only.1
 For the relevant subset of Chesapeake Bay  segments with open-water
 (OW) and deep-water (DW) and/or deep-channel (DC) designated uses:

 Match benthic stations and scores in a dataset with monthly open-water,
 deep-water, and deep-channel designated use boundaries.

     •   Boundaries  are  derived using  the standardized,  automated
         method for identifying pycnocline boundaries  documented in
         U.S. EPA 2008.

 Pycnocline  boundaries are  then interpolated using the  interpolator
 (Visual  Basic program, Version 4.61, August 2006, Chesapeake  Bay
 Program Office, as referenced in U.S. EPA 2008, Appendix A, p36).

     •   Interpolator cells are matched with benthic station locations, and
         interpolated pycnocline boundaries are applied to each benthic
         station location.
Pycnocline boundaries are
interpolated using the episodic
pycnoclines approach defined in
Chapter 2, this addendum.
 Benthic stations  (and their associated B-IBI scores) are assigned to a
 designated use: OW, DW, or DC.
No modification recommended
 To define the biological reference community for each designated use, all
 individual segment-years for which the minimum B-IBI was > 3.0 are
 identified. (Minimum sample size within a segment-year is recognized as
 n=l.) These are denoted as 'healthy' segment-years.
 a.  Use 3-year rolling time periods
    rather than single years.2
 b.  Require a B-IBI score sample
    size n^ 10.3
 C.  "Healthy" reference
    communities are those with an
    average B-IBI score > 3.0 rather
    than a minimum, and standard
    deviation (SD) < l.O.4	
 For the 'healthy' segment-years, the monthly (in the case of OW and
 DW) or instantaneous (DC) dissolved oxygen criteria violation rates are
 obtained based on the water quality profiles of sampling data collected by
 the Chesapeake Bay long term water quality monitoring program.	
No modification recommended.
 These season- and designated use-specific Chesapeake Bay dissolved
 oxygen criteria violation rates (e.g. percentage of a segment-designated
 use volumes failing the DO criteria in a given month; thus 4 measures
 per summer for OW and DW - June thru September) are used to define
 "acceptable"  exceedances  of  the dissolved  oxygen criteria.  This
 definition of acceptable exceedances in space and time is  based on the
 logic  that  if a healthy  benthic  community  existed in the segment-
 designated use in that summer, then the degree of DO criteria violation
 that occurred did not lead to an impaired benthic community.	
No modification recommended.
Source: U.S. EPA 2003, 2007, 2008.
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1.  Restrict dataset to 1996-2006 time period. For the fixed station samples included in the analyses, use the "grand
   score" only from the CIMS data base with these samples.
2.  Use 3-year rolling time periods rather than single years. This brings the reference community ID method into
   better alignment with the DO criteria assessment method for which reference communities are being identified.
3.  Require a B-IBI score sample size > 10. This improves the spatial representation of the B-IBI score.
4.  "Healthy" reference communities are those with an average B-IBI score > 3.0, and a standard deviation (SD) <
   1.0, rather than a minimum.  Using the average is consistent with published methods used by Chesapeake
   benthic experts to assess benthic communities (e.g. Llanso et al. 2009).

   APPLICATION OF A REFERENCE CURVE FOR OPEN-WATER 30-DAY MEAN
                DISSOLVED OXYGEN CRITERIA: SUMMER SEASON

Reference curves for the 30-day mean open-water dissolved oxygen criterion (June 1-September
30 only) were based on criteria levels that would not impair biological communities (U.S. EPA
2003). Analyses conducted by the EPA Chesapeake Bay Program Office and its partners suggest
that the B-IBI does not provide an appropriate reference community  for assessment of open-
water dissolved oxygen criteria violations. Even with the latest improvements in the assessment
methodology to distinguish between  healthy  and  degraded benthic communities, Figure III-3
illustrates that the health of the benthic community is not an appropriate indicator of open-water
low dissolved oxygen conditions as  defined by the summer  season open-water 30-day mean
dissolved  oxygen  criterion.  This  result  is  demonstrated by the  cloudplot  (Figure III-3)
representing  Chesapeake Bay Program segments deemed "healthy" and "degraded" according
the updated assessment methodology.
Figure III-3. Open water "healthy" and "degraded" benthic communities are not distinguished
by violations of the summer open water DO 30-day mean criterion

APPLICATION OF A REFERENCE CURVE FOR DEEP-WATER MEAN DISSOLVED
                                 OXYGEN CRITERIA

Reference curves for the 30-day mean deep-water dissolved oxygen criteria (June 1-September
30 only) were based on criteria levels that would not impair biological communities (U.S. EPA
2003). Reference areas for derivation  of the original 2003 published deep-water biologically-
                                                                                       23

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based  reference curves  were identified using  a measure of benthic community  health - the
Chesapeake Bay B-IBI (Weisburg et al. 1997). Using the revised methodology outlined in this
addendum chapter, the EPA Chesapeake Bay  Program Office and its partners identified two
distinct sets of "healthy" and "degraded" (average B-IBI < 3.0, SD < 1.0) benthic communities,
with correspondingly distinct violation rates (Figure III-4). The EPA Chesapeake Bay Program
Office, in coordination with its partners, further determined that a reference curve constructed
from the  100th percentile  of healthy violation rates  (x) for each point in time (y) accurately
distinguished  between  healthy  and  degraded  benthic communities  with zero  error  in
classification.

A step-by-step guide to the derivation of this curve (Figure III-4), including the x-y coordinate
values for plotting the curve, is provided in Appendix C.
                                                                  -CB6PH_1996_1998
                                                                  -CB7PH_1996_1998
                                                                  -CB6PH_1997_1999
                                                                  -CB7PH_1997_1999
                                                                  -CB7PH_1998_2000
                                                                  -CB6PH_1999_2001
                                                                  -CB7PH_1999_2001
                                                                  -CB7PH_2000_2002
                                                                  -CB6PH_2004_2006
                                                                  -CB7PH_2004_2006
                                                                  -PAXMH19961998
                                                                  -POTMH19961998
                                                                  -PAXMH19971999
                                                                  -POTMH19971999
                                                                  -POTMH19982000
                                                                  - PAXMH19992001
                                                                  -POTMH19992001
                                                                  - PAXMH20002002
                                                                  - RPPMH20002002
                                                                  - PAXMH20022004
                                                                  - PAXMH20032005
                                                                  - PAXMH20042006
                                                                  - RPPMH20042006
                                                                  •100th percentile
Figure III-4. Dissolved  Oxygen deep-water criteria violation rates corresponding to healthy
(blue) and degraded  (red) benthic communities.  Recommended new deep-water biologically-
based reference curve represented by the 100th percentile of healthy violations is shown in black.

          APPLICATION OF A REFERENCE CURVE FOR DEEP-CHANNEL
           INSTANTANEOUS MINIMUM DISSOLVED OXYGEN CRITERIA

In the  case of  the  deep-channel  instantaneous minimum  dissolved oxygen  criterion,  the
application  of a biological  reference curve was recommended  in U.S. EPA 2007 (p.  43).
Appendix D provides a more thorough review of history of EPA guidance regarding application
of the deep-channel reference curve.

This  2007  recommendation  for application  of  a biologically-based reference curve  for
assessment  of the deep-channel dissolved oxygen criterion was based on the identification of a
                                                                                         24

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small number of deep-channel segment-periods  within which the benthic communities were
categorized as "healthy" and,  therefore, appropriate for use as a  biological reference.  These
benthic communities were categorized using the  methodology described on pp.  39-41 of U.S.
EPA 2007.

The revised methodology published in this addendum was  applied to derive a new deep-channel
biologically-based reference curve.  The revised method yielded no segment-periods meeting the
revised criteria outlined in Table III-l above.   This  suggests that the occurrence of healthy
benthic communities in the deep-channel  designated use are currently insufficient to identify a
corresponding set of "acceptable" violations of the instantaneous minimum dissolved oxygen
criteria to develop a biologically-based reference curve.

Comparisons of Degraded Reference Benthic Communities with the Published Deep-
Channel Reference Curve
While  no benthic communities could be categorized as "healthy" in the  most recent review, 25
"degraded"  reference  benthic  community segment-periods  were  identified.    The  EPA
Chesapeake Bay Program Office and its partners  conducted  and reviewed analyses that showed
that  all 25 segment-periods  (in  the  1996-2005 time period)  for  which deep-channel benthic
communities  were categorized as  "degraded" failed  a dissolved  oxygen criteria  assessment
conducted using the 10% default reference curve (Figure III-5).
                                     space
Figure III-5. CFD graph of deep-channel instantaneous minimum dissolved oxygen criterion
violation rates corresponding to benthic communities categorized as "degraded" (red lines) in
relation to the 10% default reference curve (blue line).

In the  absence  of a suitable reference community, a biological reference curve for the deep-
channel instantaneous minimum dissolved oxygen criterion can not be constructed at this time.
Under  these  circumstances,  "a default reference curve such as the normal distribution curve
                                                                                     25

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representing approximately  10  percent exceedance is appropriate  in this case to account  for
anticipated natural criteria exceedances" (U.S. EPA 2003; p.  173).

Rationale for Acceptable Exceedances of the Deep Channel Instantaneous Minimum
Dissolved Oxygen Criterion
EPA determined that there are  allowable criteria exceedances that would not adversely effect
protection of the designated use. As documented on p. 168 in U.S. EPA 2003:

     "The  recommended criteria  attainment assessment approach is  designed to
     protect the living resources as defined by the designated uses. The criteria levels
     themselves were largely based on  scientific studies  performed in laboratory
     settings or under controlled field conditions.  The criteria establish the level of a
     given habitat condition that  living  resources need for  survival. They do not
     account for many other environmental factors that could affect survival.

     Reference curves were developed to provide a scientific-based, direct measure
     of the 'allowable' criteria exceedances. These exceedances are defined to be
     those that last a short enough time or cover a small  enough area to have no
     adverse affects on the designated use. It is assumed that the designated uses can
     be attained even with some limited level of criteria exceedances and thus, the
     reference  curves  define those criteria exceedances deemed to be allowable—
     chronic in time but over small areas, or infrequent occurrences over large areas.
     Exceedances that occur over large areas of space and time would be expected to
     have  significant detrimental  effects on biological communities, which would
     imply nonattainment of designated uses."

As  reported in a recent  paper  on  the Chesapeake Bay dissolved  oxygen  criteria  by the key
members of the original  Chesapeake Bay dissolved oxygen  (DO) criteria  team (Batiuk et al.
2009):

     "Unlike chemical contaminants or  other more conventional  pollutants,  there
     were  no  clear, well  established  guidelines for deriving criteria for DO,
     particularly for estuarine waters inhabited by fresh-water and marine species.
     The goal in setting Chesapeake DO criteria was to use  the best science possible
     to define conditions that would improve or sustain the suitability of Chesapeake
     Bay habitats for finfish and invertebrates, with the states ultimately factoring in
     consideration of attainability in adopting the criteria as water quality standards.
     Thus, we developed criteria that would  greatly increase the spatial and temporal
     extent of Bay waters in which oxygen concentrations were not  major limitations
     to growth and survival of organisms dependent on particular Bay  habitats. We
     did not, however, derive criteria that would  require oxygen concentrations high
     enough at all times  and  in  all locations  such that  no organism would be
     negatively affected in  any  location  in   the  Bay.   The  states  and  U.S.
     Environmental Protection Agency (EPA) determined that such conditions would
     not be achievable either economically  nor technologically (U.S. EPA, 2003d)
     and may not, in fact, reflect pre-historical conditions of Chesapeake Bay, which
                                                                                      26

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     showed that low oxygen conditions, although not nearly as severe as today, may
     have been  a  historical feature in the deep channel of the bay (Cooper  and
     Brush, 1991; Karlsen et al., 2000; Adelson et al., 2001; Zimmerman and Canuel,
     2002; Bratton et al., 2003; Colman and Bratton, 2003; Cronin and Vann, 2003;
     Zheng et al., 2003)."

In support  of  the deep-channel instantaneous minimum criterion of 1 mg/L, U.S. EPA 2003
summarized findings published in peer-reviewed literature  sources indicating that  several
keystone benthic species "are resistant to dissolved oxygen concentrations as low as 0.6 mg/L,"
and that "extensive mortality is likely  only under persistent  exposure to very low  dissolved
oxygen concentrations at high summer temperatures" (p. 61).

In light of both (1) the recognition that low dissolved oxygen conditions are a 'pre-historical'
feature of these deep channel habitats, and (2) the observation that keystone benthic species of
these deep channel  habitats can tolerate small-scale occurrences of severe hypoxia (DO
concentrations below 1 mg/L), EPA believes that an allowance for a small, limited set of
exceedances in time and space is acceptable in assessment of the deep-channel designated use
dissolved oxygen criterion.

       ASSESSMENT OF SUMMER SEASON DISSOLVED OXYGEN CRITERIA

EPA recommends revising previously published dissolved oxygen criteria assessment guidance
as described in Table III-2, including applying the default 10% reference curve for assessment of
summer  season (June 1- September 30) 30-day  mean open-water and instantaneous minimum
deep-channel dissolved  oxygen criteria. The 30-day mean deep-water dissolved oxygen criterion
biologically-based  reference curve,  as  described in this addendum, is recommended  for use
assessing attainment of this criterion. Until EPA publishes methodologies for assessing the 7-day
mean,  1-day mean and instantaneous minimum open-water and deep-water dissolved  oxygen
criteria, respectively, the Agency recommends that the states and the District of Columbia rely
strictly on  the assessment of the 30-day mean  open-water  and deep-water dissolved  oxygen
criteria for listing decisions (U.S. EPA 2007). The previously published non-summer open-water
dissolved oxygen criteria reference curve remains unchanged as the 10% default reference curve
(U.S. EPA 2007, p.42).
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Table III-2. EPA recommended reference curves for conducting 303(d) list Chesapeake Bay
dissolved oxygen criteria assessments.	
 Season and Designated
          Use
U.S. EPA 2007 July Addendum
       Reference Curve
 U.S. EPA 2010 Addendum
     Reference Curve
Summer Open Water
(30-day mean)
Published biologically-based
reference curve.

Refer to U.S. EPA 2007
p. 41, Figure IV-2.	
Published default 10%
reference curve

Refer to U. S. EPA 2007, p 13,
Figure II-4 and Equation 1.
Non-summer Open Water
(30-day mean)
Published default 10% reference
curve.

Refer to U.S. EPA 2007,
p. 13, Figure II-4 and Equation 1.
Published default 10%
reference curve

Refer to U.S. EPA July 2007,
p. 13, Figure II-4 and Equation
1.
Summer Deep Water
(30-day mean)
Published biologically-based
reference curve.

Refer to U.S. EPA 2007,
p. 41, Figure IV-3.	
Revised Biologically-based
reference curve, Figure III.4,
and Appendix C. this
document.
Summer Deep Channel
(instantaneous minimum)
Published Biologically-based
reference curve.

Refer to U.S. EPA 2007
p. 42 Figure IV-4 and Appendix F
andG.
Published default 10%
reference curve

Refer to U. S. EPA 2007, p 13,
Figure II-4 and Equation 1.
Sources: U.S. EPA 2003, 2007.
                                LITERATURE CITED

Adelson, J. M., G. R. Helz and C. V. Miller. 2000. Reconstructing the rise of recent coastal
anoxia; molybdenum in Chesapeake Bay sediments. Geochemica et Cosmochemica Acta
65:237-252.

Alden III, R.A.,  D.M. Dauer, J.A. Ranasinghe, L.C. Scott, and RJ. Llanso.  2002.  Statistical
verification of the Chesapeake Bay benthic index of biotic integrity. Environmetrics 13:473-498.

Bratton, J. F., S. M. Colman, R. R. Seal and P. C. Baucom. 2003.In press. Eutrophication and
carbon sources in Chesapeake Bay over the last 2,700 years: Human impacts in context.
Geochimica et CosmochimicaActa.

Batiuk,  R.A., D.L. Breitburg, RJ. Diaz, T.M.  Cronin, D.H.  Secor  and G. Thursby. 2009.
Derivation of habitat-specific dissolved  oxygen criteria for Chesapeake  Bay  and its tidal
tributaries. J. Exp. Mar. Biol. Ecol. 381:8204-8215.
                                                                                   28

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Chesapeake Bay Program. 1989. Chesapeake Bay Basin Monitoring Program Atlas. Volume II.
Biological and Living Resource Monitoring Programs. CBP/TRS 35/89. Annapolis, Maryland.

Cooper, S. R. and G.  S. Brush. 1991. Long-term history of Chesapeake Bay anoxia. Science
254:992- 996.

Colman, S. M. and J.  F. Bratton. 2003. Anthropogenically induced changes in sediment and
biogenic silica fluxes in Chesapeake Bay. Geology 31(l):71-74.

Cronin, T. M. and C.  Vann. 2003. The sedimentary record of anthropogenic and climatic
influence  on the Patuxent estuary and Chesapeake Bay ecosystems. Estuaries 26 (2A).

Karlsen, A. W., T. M. Cronin, S. E. Ishman, D. A. Willard, R. Kerhin, C. W. Holmes and M.
Marot. 2000.  Historical trends in Chesapeake Bay dissolved oxygen based on benthic
foraminifera from sediment cores. Estuaries 23:488-508.

Keller, A.A and L. Cavallaro. 2008. Assessing the US Clean Water Act 303d listing process  for
determining impairment of a waterbody. J. Env. Manage. 86:699-711.

Llanso, R.J.,  D.M. Dauer and  J.H. Volstad. 2009. Assessing ecological  integrity for impaired
waters decisions in Chesapeake Bay, USA. Marine Pollution Bulletin 59:48-53

Scientific  Technical Advisory Committee. 2006. The Cumulative Frequency Diagram Method
for Determining Water Quality Attainment: Report of the Chesapeake Bay Program STAC Panel
to Review  of Chesapeake Bay Program Analytical Tools. STAC Publication 06-003. 74 pp.

Scientific  Technical  Advisory  Committee. 2009. Application or reference curves in dissolved
oxygen criteria assessment.  STAC Review and recommendations for  the  Chesapeake Bay
Program.  STAC Publ. 09-005. http://www.chesapeake.org/stac/Pubs/biorefcurvesreview.pdf

U.S. Environmental  Protection Agency.  2003. Ambient Water Quality Criteria for Dissolved
Oxygen, Water Clarity and Chlorophyll a for the Chesapeake Bay and Its  Tidal Tributaries
(Regional Criteria Guidance).  April 2003.  EPA 903-R-03-002. Region III  Chesapeake Bay
Program Office, Annapolis, MD.

U.S. Environmental  Protection Agency.  2007. Ambient Water Quality Criteria for Dissolved
Oxygen, Water Clarity and Chlorophyll a for the Chesapeake Bay and Its Tidal Tributaries -
2007 Addendum. July 2007.  EPA 903-R-07-003. Region III Chesapeake Bay Program Office,
Annapolis, MD.

U.S. Environmental  Protection Agency.  2008. Ambient Water Quality Criteria for Dissolved
Oxygen, Water Clarity and Chlorophyll a for the Chesapeake Bay and Its Tidal Tributaries-2008
Technical Support for Criteria  Assessment Protocols Addendum. September 2008. EPA 903-R-
08-001. Region III Chesapeake Bay Program Office, Annapolis, MD.
                                                                                    29

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Weisburg, S.B, J.A. Ranasinghe, D.M. Dauer, L.C. Schaffner, RJ. Diaz and J.B. Frithsen. 1997.
An estuarine benthic index of biotic integrity (B-IBI)  for Chesapeake Bay.  1997. Estuaries
20:149-158.

Zheng, Y.,Weinman, B., Cronin, T., Fleisher, M.Q., Anderson, R.F., 2003. A rapid procedure for
thorium, uranium, cadmium, and molybdenum in small sediment samples byinductively coupled
plasma-mass spectrometry: application in Chesapeake Bay.
Appl. Geochem. 18, 539-549.

Zimmerman, A.R., Canuel, E.A., 2002. Sediment geochemical records of eutrophication
in the mesohaline Chesapeake Bay. Limnol. Oceanogr. 47, 1084-1093.
                                                                                   30

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                                     CHAPTER 4


             Revisions to the Chlorophyll a Criteria

                       Assessment Methodology


                                   BACKGROUND

In the 2003  Ambient  Water  Quality  Criteria for Dissolved  Oxygen,  Water  Clarity and
Chlorophyll a for  Chesapeake  Bay  and Its Tidal  Tributaries, EPA  published  narrative
chlorophyll a criteria that states chlorophyll a

     "...shall not exceed levels that result in ecologically undesirable consequences-
     such as reduced water  clarity, low dissolved oxygen, food supply imbalances,
     proliferation of species deemed potentially harmful to aquatic life or humans or
     aesthetically objectionable  conditions - or otherwise  render tidal  waters as
     unsuitable for designated uses balanced aquatic plant life populations and against
     the overgrowth of nuisance, potentially harmful species" (U.S. EPA 2003).

From 2004 to 2006, Virginia and the District of Columbia adopted numerical chlorophyll a
criteria for application in the tidal James River  (Virginia) and across the District's jurisdictional
tidal waters. In the  2007 publication Ambient Water Quality Criteria for Dissolved Oxygen,
Water Clarity and Chlorophyll afar Chesapeake Bay and its Tidal Tributaries-2007 Addendum,
EPA published chlorophyll a criteria assessment procedures (U.S. EPA 2007a, p. 62). With the
establishment of numerical chlorophyll a concentration-based criteria promulgated by  the states
into  their water quality standards  regulations within Chesapeake Bay tidal waters, it was
necessary to establish a reference curve for use  in the published criteria attainment process (U.S.
EPA 2003).

A biologically-based reference curve with which to assess chlorophyll a criteria attainment in
Chesapeake Bay is  not yet available. A dataset has not been identified from which there is
confidence  that a biological reference  curve  can  be derived  (U.S. EPA 2007b).  The EPA
Chesapeake Bay Program Office and its partners, in consultation with regional experts in
phytoplankton and chlorophyll a monitoring and research, have explored the published work of
Buchanan et al. 2005 and Lacouture et al.  2006  conducted during  development of  the
phytoplankton index of biotic integrity (P-IBI).

In its current form, the published P-IBI work does not provide for a suitable representation of the
integrated seasonal biological  community conditions  necessary to inform appropriate seasonal
reference conditions for Chesapeake Bay chlorophyll a criteria attainment assessments. Benthic
macroinvertebrates, for example, have life spans that integrate temporally variable environmental
conditions over space, and the effects of multiple types of environmental  stress and habitat
alteration as used with the  B-IBI for Chesapeake Bay (Llanso et al. 2009). However, standing
crops of phytoplankton  communities  will respond  to  nutrient  perturbations in  10-14 days
(Heiskary and Walker 1995).  Tracking the P-IBI results indicates any given segment can  and

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does move in and out of degradation within a single spring or summer season. Thus, the P-IBI
does  identify  instances  of high  quality  conditions,  but  currently does  not  provide the
characteristics of a season-long "healthy" condition in terms of allowable exceedances that could
be used  to  support  derivation of a  biologically-based reference for  chlorophyll a criteria
assessments.

Further work is needed  to  specify a metric that can provide a priori identification of an
unimpaired  system  on  the relevant  timescale, from which  allowable exceedance of the
chlorophyll a criteria can then be inferred. EPA, therefore, recommends a  default 10% reference
curve for assessing the chlorophyll a criteria (U.S. EPA 2007a, Figure II-4, and Equation 1).

REVIEW  OF  THE CURRENT   CHLOROPHYLL  A  CRITERIA  ATTAINMENT
ASSESSMENT PROCEDURE: METHOD AND ASSUMPTIONS

In Table IV-1, the current Chesapeake Bay chlorophyll a  attainment assessment procedure  is
outlined for developing a seasonal mean for a Chesapeake Bay management segment to compare
with published numerical criteria (e.g.,  Virginia tidal James River and the District of Columbia's
tidal waters, U.S. EPA 2007a, Appendix C).

Table IV-1. Outline of the  previously  published  Chesapeake  Bay chlorophyll a criteria
attainment assessment methodology.
Outline of chlorophyll a attainment assessement steps
Chlorophyll a data used for scenario assessments comprise all
chlorophyll a values in the CIMS water quality database with layer
flagged "S" for surface.
Data are organized into individual "cruise" files for interpolation.
Individual cruise files are interpolated using the Chesapeake Bay
Interpolator (version 4.61), with the "In-transform" and the "2-D
Inverse-Distance Squared" options selected.
Interpolated chlorophyll a surfaces are averaged for an entire season (on
a cell-by-cell basis).
Seasonal means are assessed (cell-by-cell) against the criterion for the
relevant river segment-season. Assessment curves were compared
against a default reference curve. Non-attainment is calculated by
subtracting the area of the reference curve from the area under the
chlorophyll a criteria assessment curve*.
Comments
For Virginia chlorophyll a assessments,
use all publically available and
appropriate surface data, i.e., CIMS data
plus VIMS/HRSD DATAFLOW data.
(U.S. EPA 2008 p.30).

The Interpolator automatically back-
transforms chlorophyll a values in its
output files. (U.S. EPA 2008 p.30).
The current methodology calculates an
arithmetic mean on the back-transformed
chlorophyll a values.
* If the assessment curve exceeds, at any
point, the reference CFD, then the given
segment is considered "impaired".
Source: U.S. EPA 2008

To review the method details, U.S. EPA (2007a, p. 62) first states in the assessment procedure:

        "Assessments of seasonal mean chlorophyll a criteria should be based  on
        seasonal averages of interpolated data sets.  To calculate seasonal averages,
        each interpolated cruise within a season  should be  averaged on  a point-by-
        point basis in matching interpolator grid cells. Spatial violation rates should be
        calculated  for  each  seasonally  aggregated  interpolation in an  assessment
        period.  For example summer open water  seasonal  chlorophyll a  criteria
                                                                                     32

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         assessment of  a  three-year assessment  period,  three  seasonal  average
         interpolations representing each  season  (Year 1 summer, Year 2 summer,
         Year 3 summer) should be used."

In the publication Ambient  Water Quality Criteria for Dissolved Oxygen,  Water Clarity and
Chlorophyll afar Chesapeake Bay and its Tidal Tributaries-2008 Technical Support far Criteria
Assessment Protocols Addendum, EPA provided further details documenting the chlorophyll a
criteria assessment procedures (U.S. EPA 2008). Chapter 5 (U.S. EPA 2008, pp.  30-32) reviews
the chlorophyll a criteria  procedural steps to assess attainment while Appendix G (U.S. EPA
2008) provides a highly detailed  step-by-step process  for completing the chlorophyll a criteria
assessments.  The application of data transformations to the chlorophyll  a assessment data sets
occurs during analyses in the  process of calculating the seasonal  mean  (U.S.  EPA 2008).
Chapter  5, p. 30, Step 4 (U.S.  EPA  2008) highlights the  use of such a transformation on
chlorophyll a data and states:

     "Data  sets are imported into  the Chesapeake Bay  interpolator and transformed
     (natural log) prior to interpolation, as chlorophyll a measurements tend to follow a
     log-normal  distribution.  The  program defaults  for search  area  (25 km2) and
     maximum  sample size (4) are  used, and  the  '2D Inverse  Distance Squared'
     algorithm is chosen. The  Interpolator automatically back-transforms  interpolated
     estimates before creating the output files."

Table IV-1 above shows the next  step of computing a seasonal mean requires computation of an
arithmetic mean over time at each point in the spatial interpolations represented by the 30-day
means for the appropriate chlorophyll a criteria assessment season.

First, while  the mean is often used to report central tendency, for skewed  data the arithmetic
mean may not be in accord with the  notion of 'middle'. Skewed data make it unsuitable to
estimate  quantiles, proportions or means by normal distribution expectations (Gilbert 1987), i.e.
an arithmetic mean. Tett and Wallis (1995) cite Barnes (1952) as indicating it is common for the
variance  of measurements on phytoplankton to be dependent on the mean.  Sokal  and Rohlf
(1969) recommend logarithmic transformation of data exhibiting such characteristics.

The previously  published  protocols  for   assessing  Chesapeake Bay  chlorophyll  a  criteria
attainment were  inconsistent in  carrying out the seasonal  mean  computations  since spatial
interpolations are conducted on log transformed data while temporal averaging is conducted on
untransformed data (U.S. EPA 2003, 2007a, 2008). Bland and Altman (1996) recommend that
once data are transformed,  carrying out all calculations on the transform  scale and transform
back once one has calculated the confidence intervals of the sample mean.

Transformations on data provide the ability to approximate a statistical distribution based on the
analyses  to  be performed using  established inferential statistical  procedures.  When  there is
substantial skew  in the data it  is common to transform the data to  a symmetric distribution.
Analyses conducted with data approximating a normal distribution throughout the calculations
then support the use of a wide array of well known statistical inference procedures based on well
established statistics of the normal distribution.
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Second, there is an underlying assumption to the calculations conducted as defined in the U.S.
EPA 2007a and 2008 chlorophyll a criteria assessments that Chesapeake Bay chlorophyll data
show log-normal tendencies. Based on this assumption, analyses depend on log-transforming of
chlorophyll a data to provide a reasonable approximation of the normal distribution and support
the use  of normal distributional inference  procedures. There  is  use  of log-transformation
chlorophyll a data in the Chesapeake Bay criteria literature cited in U.S. EPA 2007b, and there is
a suggestion  for positive skewness for chlorophyll a data shown with a hypothetical chlorophyll
a data distribution (U.S. EPA  2007b).  However,  there is  little  background documenting the
statistical distributional characteristics of Chesapeake Bay chlorophyll a data within the Ambient
Water Quality Criteria for Dissolved Oxygen, Water Clarity and Chlorophyll a for Chesapeake
Bay and its Tidal Tributaries publication series (U.S. EPA 2003, 2007a, 2007b, 2008).

The following sections address: 1) peer-reviewed supporting literature regarding skewness and
non-normality issues of chlorophyll  a data; 2) log-normal transformation applications during
analyses of Chesapeake Bay and other chlorophyll a data;  and 3) recommended refinements to
the published criteria assessment procedures.  All these sections are  directed towards providing
consistency in  computing the  season  mean  of the 3-year assessments  in logarithmic-space,
thereby providing a sound estimate of central tendency for the final chlorophyll a assessment
measures with the seasonal mean criteria.

     CHLOROPHYLL A: DATA SKEWNESS, LOG TRANFORMATION AND THE
                         SEASONAL MEAN CALCULATION

Log-normal Character of Chlorophyll a Data

Support for the log-normal characteristics of chlorophyll a data have  been published in the peer-
reviewed scientific literature  across a  diversity  of  ecosystems.  Harris (1986,  Figure 9.7)
illustrates  seasonally  dependent log-normal  chlorophyll results for Hamilton Harbor  (Lake
Ontario). Vollenweider and Krekes (1980), as cited in Harris (1986), noted that algal biomass
data from lakes was log-normally distributed. Recent work on Colorado lakes (n = 20) showed
19  of 20 lakes  chlorophyll measurements were  well fit  with log-normal  transformations to
approximate the normal distribution4.

Within Chesapeake Bay, Jordan et al. (1991) describe correlations between watershed discharges
and chlorophyll  concentrations as complicated by non-normal distributions. Jordan et al. (1991)
used the Box-Cox method (Sokal and Rohlf 1981) to identify the best power transformation for
normalizing the data which  was a log transformation. Harding (1994) showed that frequency
distributions  of chlorophyll and nutrient concentrations in  Chesapeake Bay  data were skewed;
logarithmic transformations of the data produced normal distributions.

4http://www.chatfieldwatershedauthoritv.org/pdf/Characterizing%20Chlorophvll%20Distributions%20in%20Colora
do.pdf

Log-transforming  Chesapeake Bay  water quality  indicator data (including chlorophyll) was
integral to improvements of the Relative Status Indicator  during its  evolution (Olson 2009).
Initially, Olson  (2009)  reports that  positive skewed  data led to  unequal  data  distributions
                                                                                      34

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affecting the outputs resulting in too many areas characterized as "good" when they were clearly
unsatisfactory. Modifications applied to indicator calculations from 1998-2000, benchmark and
status data sets (3-year windows) were log-transformed  prior to analysis to address data
skewness issues negatively impacting equality of data distribution characterizations. It was thus
noted "that for water quality parameters the log and square root transformations are about equal
in effecting a normal distribution of the data, and more effective than inverse transformations or
using untransformed data" (Olson 2009).

U.S. EPA (2007b) extended  the published analyses of Harding (1994) and Harding  and  Perry
(1997) modeling historical chlorophyll a data using a Generalized Linear Model (GLM) for logic
(chlorophyll a). In deriving  reference chlorophyll a criteria thresholds  for  Chesapeake Bay,
thresholds were recommended as being  derived by a model  for the desired  mean level of
chlorophyll a in log space (U.S. EPA 2007b, page 17). Tables III-2 and III-3 in U.S. EPA 2007b,
page  18)   illustrate reference  condition recommendations in  log  transform  space   mean
chlorophyll and back  transformed  means. Recommendations for harmful algal bloom based
chlorophyll a criteria  in tidal fresh and oligohaline waters of Chesapeake Bay were further
dependent upon log-transformed chlorophyll a analyses in their development (U.S. EPA 2007b).

James River Focused Analyses of Log-transformed Chlorophyll a Data for Normality
Tidal James  River chlorophyll  a data (1991-2000, n =  828) were log-transformed; natural
logarithms  were used. A  Generalized  Linear  Modeling  (GLM)  approach was used to test
chlorophyll a data for  normality. Statistical Analysis Software (SAS) was used in the analysis.
Seven Chesapeake Bay  segments were included in the analysis: Mouth of Chesapeake Bay
(CB8PH),  Mouth to mid-Elizabeth River (ELIPH), Southern Branch Elizabeth River (SBEMH),
Mouth of the James  River  (JMSPH),  Lower  James River (JMSMH),  Middle James  River
(JMSOH)  and Upper James  River (JMSTF). Segments were grouped into one  of four groups
depending on similarity of their variances:

                    "JMSPH"                         thenSegGrp=l;
                    "JMSMH" "SBEMH"              then  SegGrp =  2;
                    "JMSOH" "CB8PH" "ELIPH"      then  SegGrp =  3; and
                    "JMSTF"                         then  SegGrp =  4.

             The GLM model was ln(chlorophyll)= year, segment. (Equation 1)

Data was analyzed by season. Spring was defined as March, April and May with summer defined
as July, August and September. Normality diagnostics were reviewed for the raw residuals.

For Spring and Summer seasons within the tidal James River, even without standardizing  for
heterogeneous variance, the In(chla) residuals from the GLM model results show a fairly close
approximation  to  a  normal distribution.  The normal   probability  plot  shows very  high
concordance between the expected  residuals and the observed  residuals except  for two outlier
points in the extreme tails of the sample. These outliers probably reflect a failure of the simple
model to capture some extreme event rather than a failure  of log normality.  The Shapiro-Wilk
statistic of 0.994 (spring) and 0.988  (summer) shows that the residuals are  very highly  correlated
with the expected residuals  for approximating a normal  distribution  (see Appendix C). The
                                                                                    35

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Shapiro-Wilks statistic ranges from 0 to 1 where 0 is farthest from normality and 1 is high
fidelity with a normal distribution. The normality test p-value suggests a statistically significant
departure from normality but this is not surprising with a sample size n  = 828. The Shapiro-
Wilks test is sensitive to small departures from normality with large sample sizes.  The large
sample size gives one the power to detect very small statistical differences from normality that,
for analysis of the transformed data, are of low practical significance. Further details of the test
output are provided in Appendix D. The SAS programs are included in Appendix E.

     CHLOROPHYLLS CRITERIA ASSESSMENT PROTOCOL REFINEMENTS
                          USING LOG-TRANSFORMATIONS

Statistical treatment of chlorophyll a data from a review of non-Chesapeake Bay and Chesapeake
Bay  specific  peer  reviewed  scientific literature and  U.S. EPA  Chesapeake Bay criteria
documentation: 1) supports a common recognition of skewness with chlorophyll a data sets; and
2) shows a long  history with the  application  of log-transformations for analyses. Bland  and
Altman  (1996) recommend carrying out all calculations on the transform scale and transform
back once one has calculated the confidence intervals of the sample mean. Log transformation of
data  during  analyses  to better reflect a normal  distribution then  better support the inference
procedures  based on normal distributions. The chlorophyll  a criteria  assessment protocol
modifications described here (Table IV-2) constitute a more consistent and technically  sound
calculation than the currently published EPA methods (U.S. EPA 2003, 2007a, 2008). Analyses
conducted with data  approximating a  normal distribution throughout the  calculations supports
the use of a wide array of statistical inference  procedures based on normal  distributions. Tidal
James River chlorophyll a data was evaluated and showed fidelity to the normal distribution.
                                                                                      36

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Table IV-2. Previously published Chesapeake Bay chlorophyll a criteria assessment methods
and recommended modifications.
U.S. EPA 2008 Addendum
1 . Chlorophyll a data used for scenario assessments
comprise all chlorophyll a values in the CIMS
water quality database with layer flagged "S" for
surface.
2. Data are organized into individual "cruise" files foi
interpolation.
3. Individual cruise files are interpolated using the
Chesapeake Bay Interpolator (version 4.61), with
the "In-transform" and the "2-D Inverse-Distance
Squared" options selected. The Interpolator
automatically back-transforms chlorophyll a
values in its output files.
4. Interpolated chlorophyll a surfaces are averaged
for an entire season (on a cell-by-cell basis). The
current methodology calculates an arithmetic
mean on the back-transformed chlorophyll a
values
5. Seasonal arithmetic means are assessed (cell-by-
cell) against the criterion for the relevant river
segment-season.
U.S. EPA 2010 Addendum
No modification recommended.
No modification recommended.
No modification recommended.
4a. Interpolated chlorophyll a surfaces are In-transformed
4b. Seasonal means are calculated on In-transformed
chlorophyll a values.
5. Ln-transformed seasonal means are assessed (cell-by-
cell) against the In-transformed criterion for the
relevant river segment-season.
Source: U.S. EPA 2008.

           IMPLICATIONS OF THE REVISED ASSESSMENT PROTOCOL

Conducting the spatial and temporal analyses in log-space produces geometric means. Geometric
means will be less than the arithmetic means of the raw data, i.e. bias low for the estimator of the
arithmetic mean, for all data sets with at least one pair of nonequal values (Bland and Altman
1996). When all values in the data set are the same value and only then will the arithmetic mean
equal the geometric mean. However, while geometric means may be less than arithmetic means,
the values will always be above the minimum observed value and below the observed maximum
value in both approaches. For log-normally distributed data such as the chlorophyll a data, the
geometric mean is further a more efficient measure of central tendency, efficiently estimating the
median which might be considered more typical of observations from the sampled population (E.
Perry, 2010, Pers. Comm.).

Given the very small number of data points in the tidal James River data analyses that influence
the statistical measure of departure  from normality, then this departure  occurs  in  a small
percentile of the distribution. Overall, the data align very well with the expected up through the
10th percentile (see Appendix D). Because the CFD assessment method is defining the upper
bound chlorophyll a criteria somewhere around the 10th percentile, it is fair to conclude that the
log-normal is adequate for that purpose. While there may  be another distribution that matches the
data better  than  the normal distribution, one  would, however, have to weigh the  benefits  of
improved estimation against the costs of developing a suite of estimation procedures for this
other distribution.   One clear  advantage  of working  with the log-normal is  that  the log
transformation provides for a normal metric where one has many choices of well developed and
well tested statistical methods (E. Perry 2010, Pers. Comm.).
                                                                                    37

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The present Virginia water quality standards for tidal James River and the District of Columbia's
water quality standards for its tidal waters, stated as seasonal chlorophyll a means, reflect the
importance of the assessment in measuring central tendency compared to an acceptable upper
bound for acceptable  water quality conditions.  Chlorophyll a is a parameter whose measures
repeatedly show skewed distributions appropriate to log transformation to approximate a normal
distribution for  making inference  with well  developed,  well tested statistical methods. It is
therefore appropriate  to use  a  statistic that addresses  the central tendency respecting the
appropriate statistical properties of such data, i.e., the geometric mean.

The EPA Chesapeake Bay Program Office and its partners tested the recommended revised
assessment methodology for Chesapeake Bay data (e.g., tidal James River) and compared the
results with the application of the promulgated Virginia water quality standards' chlorophyll a
criteria. Results showed almost universally greater levels of chlorophyll a attainment using the
recommended revised  methodology compared with the previously EPA published criteria
assessment method (and adopted into Virginia's water quality standards). Acknowledging these
findings, the revisions to the published criteria assessment method are recommended for
ensuring consistency within the assessment procedures with acknowledged the statistical
properties of the chlorophyll a data.
                                 LITERATURE CITED

Barnes, H. 1952. The use of transformations in marine biological statistics. J. du Cornell 18:61-
71.

Bland, J.M and D.G. Altman. 1996. Transformations, means and confidence intervals. British
Medical Journal 312:1079.

Buchanan, C., R. V. Lacouture, H. G. Marshall, M.  Olson, J. Johnson. 2005. Phytoplankton
reference communities for Chesapeake Bay and its tidal tributaries. Estuaries 28(1): 138-159.

Colorado                                   lakes                                   study.
http://www.chatfieldwatershedauthoritv.org/pdf/Characterizing%20Chlorophyll%20Distribution
s%20in%20Colorado.pdf

Gilbert, R.O. 1987. Statistical methods for environmental pollution monitoring. Van Nostrand
Reinhold, New York, NY.320 pp.

Harding, L. Jr. 1994. Long term trends in the distribution of phytoplankton in Chesapeake Bay:
roles of light, nutrients and streamflow. Mar. Ecol. Prog. Ser.  104:267-291.

Harding,  L.W.,  and  E.S.  Perry.  1997. Long-term  increase of phytoplankton biomass  in
Chesapeake Bay, 1950-1994. Mar. Ecol. Prog.  Series. 157:39-52.
                                                                                      38

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Harris, G.P. 1986. Phytoplankton ecology:  Structure, function and fluctuation. Chapman  and
Hall, New York, NY. 384 pp.

Heiskary, Steven A and William W. Walker,Jr. 1995. Establishing a chlorophyll agoal for a run-
of-the-river reservoir. Lake andReserv. Manage. 11(1):67-76.

Jordan,  T.E., D.L. Correll, J. Miklas, and  D.E.  Weller. 1991. Long-term trends in  estuarine
nutrients and chlorophyll, and short term effects of variation in watershed discharge. Mar. Ecol.
Progr. Ser. 75:121-132.
Lacouture, R.V., Johnson, J.M., Buchanan, C., and Marshall, H.G. 2006. Phytoplankton index of
biotic integrity for Chesapeake Bay and its tidal tributaries. Estuaries Coasts 29: 598-616.

Llanso,  R.J., D.M. Dauer and J.H. Volstad. 2009. Assessing ecological integrity for impaired
waters decisions in Chesapeake Bay, USA. Marine Pollution Bulletin 59:48-53

Olson, M. 2009. Relative Status Indicator: Development and evolution of a relative measure of
condition for assessing the status of water quality and biological parameters tracked in the U.S.
EPA Chesapeake Bay Program long term monitoring programs. Final Report. September 2009.
Chesapeake Bay Program Office, Annapolis, MD.

Sokal, R.R. and F.J. Rohlf. 1981. Biometry, 2nd ed. W.H. Freeman, San Francisco, CA.

Tett, P.  and A. Wallis.  1978. The general annual cycle of chlorophyll  standing crop in Lock
Creran. J. Ecol. 66:227-239.

U.S. Environmental Protection Agency. 2003. Ambient Water Quality  Criteria for Dissolved
Oxygen,  Water Clarity and Chlorophyll a for the Chesapeake Bay and Its Tidal Tributaries
(Regional Criteria Guidance) April  2003.  EPA 903-R-03-002. Region III Chesapeake Bay
Program Office, Annapolis, MD.

U.S. EPA 2007a. Ambient Water Quality  Criteria for Dissolved Oxygen,  Water Clarity  and
Chlorophyll afar the Chesapeake Bay and its Tidal Tributaries - 2007 Addendum. July 2007.
EPA 903-R-07-003. Region III Chesapeake Bay Program Office, Annapolis, MD.

U.S. EPA 2007b. Ambient Water Quality  Criteria for Dissolved Oxygen,  Water Clarity  and
Chlorophyll a for the  Chesapeake  Bay and its Tidal Tributaries  - Chlorophyll Criteria
Addendum. November 2007. EPA 903-R-07-005. Region III Chesapeake Bay Program  Office,
Annapolis, MD.

U.S. EPA 2008. Ambient Water Quality Criteria for Dissolved Oxygen,  Water Clarity  and
Chlorophyll a for the Chesapeake Bay and  its Tidal Tributaries - 2008 Technical Support for
Criteria Protocols Addendum. September 2008. EPA 903-R-08-001. Region III Chesapeake Bay
Program Office, Annapolis, MD.
                                                                                    39

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Vollenweider, R.A.  and J.  Krekes.  1980. The loading concept as a  basis for controlling
eutrophication: philosophy and preliminary results of the OECD programme on eutrophication.
Prog. Wat.  Technol. 12:5-38.
                                                                                     40

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                                ACRONYMS

2-D          two-dimensional
B-IBI        benthic index of biotic integrity
CBP         Chesapeake Bay Program
CIMS        Chesapeake Information Management System
CFD         cumulative frequency distribution
CHLA       chlorophyll a
DC          deep channel
DO          dissolved oxygen
DW          deep water
EPA         U.S. Environmental Protection Agency
GLM        Generalized Linear Model
HRSD       Hampton Roads Sanitation District
m           meters
mg/L        milligrams per liter
OW          open water
P-IBI        phytoplankton index of biotic integrity
S            surface
SAS         Statistical Analysis Software
SD          Standard Deviation
STAC        Scientific and Technical Advisory Committee
VIMS        Virginia Institute of Marine Science
                                                                                   41

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                            Appendix A.

B-IBI Sample  Size and Standard Deviations on B-IBI

    Scoring when Screening Segments  for Reference

                 Community Characterization

The EPA Chesapeake Bay Program Office in cooperation with its  partners, examined the
effects  of relaxing the  data screening criteria to accept segment-period combinations as
"healthy" when defining reference communities with  sample size > 8  (instead  of the
recommended n > 10) and/or standard deviation < 1.2 (instead of the recommended  <  1.0).
Data were 1996-2006 from the CIMS database.

For the "fixed station" samples both "total_score" and "grand_score" records were included.
"Total_score" records are replicate measurements of the same  sampling event; the average of
these is reported as the "grand_score."  Benthic experts (Llanso, Versar, Inc.) recommend
using the "grand_score" in these analyses. Four scenarios were explored (Table A-l). The
EPA accepted screening criteria is the default under Scenario  A.  Scenarios B, C  and D
relaxed the  standard  deviation,  sample size and both sample size and standard deviation,
respectively.

Table Al. Healthy deep-water segments as characterized with four scenarios of screening
criteria.  The accepted screening criteria is Scenario A.




Total number of
"healthy" deep water
segment-periods
Scenario A
(Default)
B-IBI > 3.0
n> 10
S.D. < 1.0

10
Scenario B
B-IBI > 3.0
n> 10
S.D. < 1.2

11
Scenario C
B-IBI > 3.0
w>8
S.D. < 1.0

13
Scenario D
B-IBI > 3.0
w>8
S.D. < 1.2

16
Relaxation of the criteria results in moderate increases (ranging from 1 to 6) in the number of
segment-periods classified  as "healthy."    Due  to  the  increased risk  of inaccurate
classification, it is important to examine not just the number of additional segment-periods,
but also the shape of these curves.  If a curve is classified as "healthy" but its location in CFD
space is consistent with DO violation CFDs  of segment-periods classified  as "degraded,"
then it is reasonable to question whether an inaccurate classification has occurred.

In the case of Scenario B (relaxing the standard deviation criterion from a maximum of 1.0 to
a maximum of  1.2), a single curve (CB5MH 1999-2001) is added to the group of "healthy"
segment-periods.  In  Figure A-l below, this curve is visible as a light blue line, while the
population of 10 curves identified  in Scenario A are presented by dark blue lines. Degraded
                                                                            42

-------
segment-periods are visible as red lines.  The biologically-based reference curve generated
from the 1001
line.
            ,th
percentile of "Scenario A" violations at each time step is visible as a yellow
      0.2

                                  OA
                                               0.6
                                                            0.8
                                       space
Figure A-l. Scenario B - illustrates the impact of maintaining the sample size criterion of n
>_10 while relaxing the standard deviation criterion from a maximum of 1.0 to a maximum of
1.2
The shape of the CB5MH 1999-2001 curve (light blue line in Figure A-l) raises the question
of  whether increasing the uncertainty  of the  screening  criteria  resulted  in  erroneous
classification of this segment-period as healthy.  In particular, the location of the top half of
this curve in CFD space that is dominated by degraded curves decreases confidence in the
accuracy of its classification.  The addition of this curve, particularly in combination with the
methodology of taking the 100th  percentile of each curve at each  point in time, would
increase the potential for the resulting biologically-based reference curve to allow  rates of
hypoxia that result in degradation of the benthic community.

In the case of Scenario C, the standard deviation is  kept consistent with the recommended
screening criteria but the sample size criterion is relaxed from 10 to 8.  This relaxation of the
recommended criteria results in the classification of 3 additional segments as "healthy."  The
CFD curves for these additional segments are shown as light blue curves in Figure A-2.

While two  of the additional curves  (CB6PH 1998-2000 and CB6PH 2000-2002) fall within
the cloud of violation rates deemed "acceptable," one curve (CB3MH 1996-1998) once again
extends into the cloud of data dominated by CFDs associated with degraded segment-periods
(Figure A-2).  As described earlier, this raises the concern that relaxation  of the criteria has
resulted in the inaccurate classification of a degraded segment-period as healthy.

-------
The  relaxation of both the sample  size and the standard deviation criteria (Scenario D)
increases the number of segment-periods classified as "healthy" from 10 to 16. However, 4
of these additional CFD curves extend into "degraded" CFD space to a degree that calls into
question the accuracy of their classification as healthy (Figure A-3).
Figure A-2. Scenario C - illustrates the impact from relaxing the sample size criterion from n
> 10 to n > 8 while maintaining the standard deviation criterion of S.D. < 1.0.

Relaxing the screening criteria for defining healthy segments based on the B-IBI with respect
to minimum sample size and maximum standard deviation increases the number of healthy
segments that can be used to generate the biologically-based reference curve.  However, the
increased uncertainty of accurate classification resulting from relaxation of the  criteria far
outweighs the potential benefit of increased sample size.  For the reference CEVIS dataset, the
EPA recommended methodology results in  a total sample  size of 24 segment-periods, of
which 10 are classified as healthy and 14 are classified as degraded. Accounting for the trade
offs  with segment  classification risks, this  present method is  supported as sufficient in
generating a low risk sample size for elucidating the boundary between acceptable (i.e. those
which allow a healthy benthic community to persist) and unacceptable violations of the deep-
water DO criteria.
                                                                                   44

-------
           *^^\\^\Xi
                   A\\
               ^^>,N»%j^i>ii
            iVl«w^x
Figure \-3. Scenario D - illustrates the impact from relaxing the sample size criterion from n >

10 to n > 8 and the standard deviation criterion of SD < 1.0 to SD < 1.2.
                                                                             45

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                                 Appendix B.

       Shape of the Biologically-based Reference Curve

The  shape of the  biologically-based reference  curve  is an  important factor  in  identifying
acceptable violations of the Chesapeake Bay water  quality criteria for dissolved oxygen. The
shape of biologically derived reference curve has  thus far reinforced the suitability  of the
hyperbolic  10% default  reference  curve  when  a biologically-based reference curve  is
unavailable. An alternative hypothesis, however,  is that comparing the total area under a CFD
assessment curve to the total area under the biologically-based reference curve  is a better
measure of the degree to which healthy biological  communities can tolerate violations of the DO
criteria than the existing "point" method. Arguments put forth to support this proposal include:
(1) a segment-period may  exceed the biologically-based reference curve in one area of CFD
space while the overall area of its exceedance is within than that represented by the biologically-
based reference curve; (2) there is high variability in the shape of CFD curves and the data do not
allow identification  of combinations of time  and volume that lead to poor B-IBI  scores in a
segment; and (3) the proposed "area" method has lower error rates than the published "point"
method, even with the modifications proposed by EPA to the latter method.

With regard to arguments  1 and 2,  application of the  method modifications outlined in this
addendum, Chesapeake Bay benthic communities  are now being accurately  classified as
"healthy" or "degraded" when there is sufficient data to do so.  As a result, the Chesapeake Bay
data  support a rather specific combination of time and volume that forms the boundary between
healthy and degraded benthic communities in the deep-water designated use (Figure B-l).
Figure B-l. Dissolved oxygen violation curves associated with healthy (blue) and degraded (red)
benthic communities in deep-water designated use habitats.  The deep-water biologically-based
reference curve (yellow) is also shown.
                                                                                   46

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Using a dataset with all duplicate records appropriately removed and all appropriate screening
criteria applied, the error rate for the "Point Method" is zero. In this case, all segment-periods
classified  as "healthy" using the recommended screening criteria (n >_10, SD <  1.0) pass the
EPA recommended  deep-water biologically-based reference  curve,  and all  segment-periods
classified  as "degraded" fail the recommended biologically-based reference  curve (Tables B-l
and B-2).

Table  B-l. Segment  classifications using  the recommended screening criteria:  deep-water
designated use.
Method

Published "Point"
Method
Proposed "Area"
Method
Correct
Healthy Segments
Passing
100%
100%
Degraded
Segments Failing
100%
100%
Incorrect
Healthy Segments
Failing
0%
0%
Degraded Segments
Passing
0%
0%
Table  B-2.  Segment-period  classifications under the
designated use.
recommended method:  deep-water
Method


Published "Point"
Method












Proposed "Area"
Method












Correct
Healthy Segments
Passing
CB6PH 1996 1998
CB7PH 1996 1998
CB6PH 1997 1999
CB7PH 1997 1999
CB7PH 1998 2000
CB6PH 1999 2001
CB7PH 1999 2001
CB7PH 2000 2002
CB6PH 2004 2006
CB7PH_2004_2006




CB6PH 1996-1998
CB6PH 1997-1999
CB6PH 1999-2001
CB6PH 2004-2006
CB7PH 1996-1998
CB7PH 1997-1999
CB7PH 1998-2000
CB7PH 1999-2001
CB7PH 2000-2002
CB7PH 2004-2006




Degraded Segments
Failing
PAXMH 1996 1998
POTMH 1996 1998
PAXMH 1997 1999
POTMH 1997 1999
POTMH 1998 2000
PAXMH 1999 2001
POTMH 1999 2001
PAXMH 2000 2002
RPPMH 2000 2002
PAXMH 2001 2003
PAXMH 2002 2004
PAXMH 2003 2005
PAXMH 2004 2006
RPPMH 2004 2006
POTMH 19992001
POTMH 19982000
RPPMH20022004
PAXMH 19992001
PAXMH20012003
PAXMH20042006
POTMH19971999
PAXMH20032005
PAXMH20002002
POTMH19961998
RPPMH20002002
PAXMH20022004
PAXMH 1996 1998
PAXMH 1997 1999
Incorrect
Healthy Segments
Failing




























Degraded
Segments Passing




























                                                                                      47

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Both methods result in the same  error rates when duplicate records are removed and  EPA's
criteria  are  applied  to  the classification of benthic communities.   However, in contrast to
Argument 2 as described above, it is EPA's position that this dataset does  provide convincing
biological information with regard to the degree and  distribution of deep-water DO criteria
violations that can be tolerated by the benthic  community.   Furthermore,  by using  the worst
violation rate allowed by any healthy community at  each point in time, EPA has allowed for
greater violation rates in regions of CFD-space  where CFD curves from healthy and degraded
communities overlap.  It is reasonable to postulate, based on the distribution in CFD-space of
curves associated with healthy and degraded benthic  communities, that violations occurring in
the CFD-space circled in black in Figure B-2 lead to degradation of the benthic community.
                  0.2
                              0.4
                                           0.6
                                                       0.8
                                    space
Figure B-2. Violations occurring in the CFD-space circled in black are postulated to lead to
degradation of the benthic community.

It  is suggested from the multiple lines of evidence that the shape of the  biologically-based
reference curve is an important factor in identifying acceptable violations of the DO criteria. The
shape of this biological reference curve also provides further support for the suitability of the
hyperbolic 10% reference curve, in that it illustrates the sensitivity of biological communities to
chronic violations of DO criteria.
                                                                                       48

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                                 Appendix C.
      Derivation of the Deep-Water Biologically-Based
                              Reference Curve
Step 1. We obtained a dataset of benthic scores for the Chesapeake Bay and tidal tributaries.
Data used comprised benthic communities sampled between  1996 and 2006,  from both the
"fixed station" and "random strata" sampling programs. Only "grand score" values - which are
an average of replicate samples - were included from  the fixed station program.   For the
purposes of deriving a deep-water biological reference curve, we restricted the  dataset to only
those samples taken in segments that contain a deep-water designated use.

Step 2. We  removed from the dataset any samples obtained from  the  following segments:
PATMH, SBEMH,  and CB5MH. Benthic communities  in  PATMH and SBEMH are widely
understood to be impacted by chemical contaminants (pers. comm, Roberto Llanso, Versar Inc.);
a complication that confounds the relationship between hypoxia  and benthic community health in
these areas.

In the case of CB5MH, areas greater than 12 meters in depth - which account for 35 percent of
the bottom surface area of CB5MH - are excluded from the benthic sampling program because
they are assumed to be azoic or nearly azoic. For their analyses of benthic health, Llanso et al.
2009 assume that all areas greater than 12 m  in depth are  degraded, and perform a post-hoc
correction  to factor this assumption into their benthic assessment. For purposes of developing a
biological  reference curve, the exclusion from sampling of such a large portion of CB5MH calls
into  question our  ability to accurately characterize the health  of its deep-water benthic
communities.

Step 3. We  obtained water  quality  data from the Chesapeake Bay Program  Water Quality
database for the time period 1996-2006.  Using the standardized method for locating pycnocline
boundaries (see U.S. EPA 2008), we determined the depth of  the upper and lower pycnocline
boundaries for all sampling events in this time period.

Step 4. From this  dataset,  we  selected the sampling event that was  closest  in space (at a
minimum,  within the same segment) and time (at a minimum,  within the same month) to each
benthic sampling event.

Step 5. We then  classified each  benthic sample as an "open-water," "deep-water," or "deep-
channel"  benthic sample based on  its depth  relative to  the upper and  lower  pycnocline
boundaries of the paired water quality  sampling event.  Benthic samples that were taken at
depths between the upper and lower boundaries of the pycnocline were classified as "deep-
water" samples.  When no lower boundary was identified, benthic  samples from depths below
the  upper  boundary of the pycnocline were classified as "deep-water." Benthic samples that
could not be paired with a pycnocline boundary were discarded.
                                                                                 49

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Step 6.  For each 3-year time window from 1996-2006, we applied the following criteria to
classify  deep-water benthic communities as "healthy" for the purposes of generating a biological
reference curve for the dissolved oxygen criteria assessment: at least 10 benthic IBI scores; mean
score >  3.0; and standard deviation of the mean < 1.0. Segment-periods (e.g. "CB6PH 1996-
1998") that met the above criteria were classified as "healthy."

Step 7.  We obtained the "dissolved oxygen violation rates"  for each healthy segment-period.
These rates are an intermediate  product of the  dissolved oxygen criteria assessment procedure
(see U.S. EPA 2007 and 2008).   They represent the fraction of deep-water in a given segment
that violates water quality criteria for dissolved oxygen in a given time period.  Using these
violation rates, we generated a  CFD curve for  each healthy segment-period.  Most segment-
periods  contained 12 violation rates, but some contained only 11 rates.  To account for segment-
periods  with different  numbers  of violation rates, all violation rates  were  interpolated to a
common set of plotting  positions (y values).  To generate a biological reference curve that
represented the "100th percentile" of healthy violation rates, we then used the largest violation
rate (across healthy segment-periods) for each "y" value of the violation CFD.  The resulting  set
of violation rates represents the  largest of all healthy violation rates for each  plotting position.
See chapter 3 for more details on the selection of the 100th percentile curve.

The following segment-periods comprised the set of "healthy" segment-periods:

                                     CB6PH 1996-1998
                                     CB7PH 1996-1998
                                     CB6PH 1997-1999
                                     CB7PH 1997-1999
                                     PAXMH 1997-1999
                                     CB7PH 1998-2000
                                     CB6PH 1999-2001
                                     CB7PH 1999-2001
                                     CB7PH 2000-2002
                                     CB7PH 2003-2005
                                     CB6PH 2004-2006
                                     CB7PH 2004-2006
                                                                                     50

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The resulting deep-water biologically-based reference curve for dissolved oxygen assessment is
illustrated in Chapter 3, Figure III-4 of this addendum and defined as:
X
(violation
rate)
0
0
0
0
0.025641
0.029132
0.051185
0.200524
0.246642
0.271513
0.356639
0.402786
0.555376
1
Y
(plotting
position)
1
0.923077
0.846154
0.769231
0.692308
0.615385
0.538462
0.461538
0.384615
0.307692
0.230769
0.153846
0.076923
0
                                  LITERATURE CITED

Llanso, R.J., D.M. Dauer and J.H.  Volstad. 2009. Assessing ecological integrity for impaired
waters decisions in Chesapeake Bay, USA. Marine Pollution Bulletin 59:48-53

U.S. Environmental Protection Agency.  2007. Ambient Water Quality Criteria for Dissolved
Oxygen, Water Clarity and Chlorophyll a for the Chesapeake Bay and Its Tidal Tributaries -
2007 Addendum. July 2007. EPA 903-R-07-003.  Region III Chesapeake Bay Program Office,
Annapolis, MD.

U.S. Environmental Protection Agency.  2008. Ambient Water Quality Criteria for Dissolved
Oxygen, Water Clarity and Chlorophyll afar the Chesapeake Bay and Its Tidal Tributaries 2008
Technical Support for Criteria Assessment Protocols Addendum. September 2008. EPA 903-R-
08-001. Region III Chesapeake Bay Program Office, Annapolis, MD.
                                                                                    51

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                                 Appendix D.

  History of EPA Guidance Regarding the Deep-Channel

                              Reference Curve

In April 2003, the EPA published the guidance document, Ambient Water Quality Criteria for
Dissolved Oxygen,  Water  Clarity and Chlorophyll a for the  Chesapeake Bay and Its Tidal
Tributaries (U.S. EPA 2003).   In  this publication, EPA documented the derivation of the
dissolved oxygen criterion protective of the seasonal deep channel designated use.  For seasonal
deep-channel  designated use, an instantaneous minimum criterion of 1 mg/L was determined to
protect benthic organisms residing in the:

       "deep  water-column and adjacent bottom  surficial sediment habitats located
       principally in the river channel at the lower reaches of the  major rivers and
       along  the spine of the middle mainstem  Chesapeake Bay  at depths  below
       which seasonal anoxic (< 0.2 mg/L dissolved oxygen) to severe hypoxic
       conditions  (<  1 mg/L dissolved oxygen) routinely set in and persist  for
       extended periods of time under current conditions" (p. 60 in U.S. EPA 2003).

In support of the instantaneous minimum criterion of 1  mg/L, U.S. EPA  (2003) summarized
findings published in peer-reviewed literature sources indicating that several  keystone benthic
species  "are  resistant to  dissolved oxygen concentrations as low as 0.6  mg/L," and that
"extensive mortality is likely only  under  persistent exposure to  very low dissolved oxygen
concentrations at high summer temperatures" (p. 61).

U.S. EPA (2003) also reported that in the mesohaline Chesapeake Bay (the primary location of
the seasonal deep-channel  designated use), "dissolved oxygen concentrations of less than 1 mg/L
lead to  mortality for even tolerant species (p.  61) and that "when dissolved  oxygen  drops
significantly below 1 mg/L for even short  periods of time (on the order  of hours) mortality
increases, even for tolerant species" (p. 65).  Furthermore, it was  stated that  "States and other
users must  recognize that  the deep-channel dissolved oxygen criterion  is  stated as  an
instantaneous minimum,  thus any exceedance is  assumed to have direct consequences to the
survival of the bottom-dwelling community" (p. 151).

Regarding the definition of a water quality  standard, it is explained in U.S. EPA 2003 and in
Chapter 3 of the U.S. EPA's Water Quality Standards Handbook, 2nd Edition  (U.S. EPA 1994)
that water quality criteria definition and assessment comprises not just the magnitude of a water
quality criterion (i.e.  "the  quantifiable  condition," in  this case the concentration of dissolved
oxygen), but also the duration and frequency of that condition.

In this context, duration  is  addressed by  restricting  the applicability of the criterion to the
summer period (June - September) when stratification and severe hypoxia occur in deep-channel
regions of the Chesapeake Bay, and by defining the assessment period as "the  most recent three
consecutive years for which relevant monitoring data are available" (U.S. EPA 2003; p. 150-1).
                                                                                   52

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The frequency component  of the criterion "is directly addressed through comparison of the
generated cumulative frequency distribution (CFD) with the applicable criterion reference curve"
(U.S. EPA 2003; p. 151).

In summary, statements were made in U.S. EPA 2003 suggesting that the benthic community can
tolerate small  violations of the deep-channel instantaneous minimum criterion,  but  statements
were also made suggesting that any violation of this criterion has negative effects on the survival
of  deep-channel  benthic species. However, as  also described in U.S.  EPA  2003,  national
guidelines define a water quality standard as  comprising not only the magnitude of a given
condition, but also the  duration over which that condition is assessed and the frequency of
violation allowed within the given assessment duration. For the case of the Chesapeake Bay, the
frequency of allowable violation is defined by the location of a reference CFD, more  commonly
called a "reference curve"  (both the rationale for use of a biological reference curve and the
development of the 10 percent reference curve are also well documented in U.S. EPA 2003).
                                  LITERATURE CITED

U.S. Environmental Protection Agency. 1994. Water Quality Standards Handbook, 2nd Edition.
August 1994. EPA 823-8-94-005a. Washington D.C.

U.S. Environmental Protection Agency. 2003. Ambient Water Quality Criteria for Dissolved
Oxygen,  Water Clarity and Chlorophyll a for the Chesapeake Bay and Its Tidal Tributaries
(Regional Criteria Guidance).  April 2003. EPA  903-R-03-002.  Region III Chesapeake Bay
Program Office, Annapolis, MD.
                                                                                     53

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                               Appendix E.
    James River Chlorophyll a Data normality Analysis
  Checking Normality  of Log-transformed Chlorophyll a
                                    Data
Summary Notes regarding results of test log-normal assumption for James River chlorophyll. E.
Perry 2/24/2010.


SUMMER:
Even without standardizing for heterogeneous variance, the In(chl) residuals from the Year X
Segment model seem to be fairly close to a normal  distribution.  The normality test show
significant departure from normality

Tests for Normality for un-standardized residuals
Test
Shapi ro-Wilk
Kolmogorov-Smi rnov
         --Statistic	
         W     0.988201
         D     0.059704
                     Pr
                     Pr
          -p Value	
           W     <0.0001
           D     <0.0100
but this is not surprising with a sample size of 828. The large sample size gives you the power to
detect very small differences from normality. The Shapiro-Wilk statistic of 0.988 shows that the
residuals are very highly correlated with the expected residuals from a normal distribution. The
normal  probability plot shows very high concordance between the expected residuals and the
observed residuals except for two outlier points in the extreme tails of the sample. These outliers
probably reflect a failure of our simple model to capture some extreme event rather than a failure
of log normality.
Levene's test shows that the data do exhibit heterogeneous variances even in the log-metric. This
heterogeniety seems to be associated with changing variance over segments.

Levene's test for un-standardized  residuals
                                      12:18 Wednesday, February  24, 2010
                           The GLM Procedure
Dependent Variable: aRChl
 Source
 Model
 Error
 Corrected Total
 Source
 year
 CBSEG_2003
DF
 9
 6

DF
15
812
827
Sum of
Squares
96.2326827
218.9548618
315.1875445

Mean Square
6.4155122
0.2696488

Type III SS
 3.87412641
91.23339617
Mean Square
 0.43045849
15.20556603
Value
 1.60
56.39
                                            Value
                                            23.79
                                             p-value
                                             <.0001
p-value
0.1120
<.0001
                                                                            54

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Standardizing the residuals by estimates of standard deviation by segment-group and year leads
to improvement in both normality and homogeneous variance.   However, both non-normality
and heterogeneous variance remain statistically significant.
Tests for Normality for standardized residuals
Test
Shapi ro-Wilk
Kolmogorov-Smi rnov
          --Statistic	
          W      0.99529
          D     0.032238
                       Pr
                       Pr
           -p Value	
            W       0.0121
            D       0.0361
Levene's test for standardized residuals
Dependent Variable:  aStdResChl
 Source
 Model
 Error
 Corrected Total

DF
15
812
827


12
309
322
Sum of
Squares
.9100323
.7902578
.7002901

Mean
0.
0.


Square
8606688
3815151


F Val
2.



ue
26



p-val ue
0.0041


 Source
 year
 CBSEG_2003
DF
 9
 6
Type  III SS
 3.48204541
 9.43256796
Mean  Square
 0.38689393
 1.57209466
F Value
   1.01
   4.12
p-value
0.4267
0.0004
Again the heterogeniety seems to be associated with segments which suggests that the grouping
algorithm could be improved.
SPRING:

Similar  to  summer results,  without  standardizing for heterogeneous  variance, the In(chl)
residuals from the Year X Segment model seem to be fairly close to a normal distribution.  The
normality test show significant departure from normality but  the p-value is larger than for
Summer.
Tests for Normality for un-standardized residuals
Test
Shapi ro-Wi'l k
Kolmogorov-Smirnov
          --Statistic	
          W     0.994437
          D     0.034024
                       	p  Value	
                       Pr < W       0.0070
                       Pr > D       0.0323
The Shapiro-Wilk statistic of 0.994 shows that the residuals are very highly correlated with the
expected residuals from a  normal distribution. The normal probability plot shows very high
concordance between the expected residuals and  the observed residuals and like the result for
summer, the departure from normality  appears  as outlier points in the  extreme  tails of the
sample.


Levene's test shows that the data do exhibit heterogeneous variances even in the log-metric. This
heterogeniety seems to be associated with changing variance over both segments and years.
                                                                                   55

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Levene's test for  un-standardized residuals
                                       12:18 Wednesday, February  24,  2010

Dependent Variable:  aRChl
 Source
 Model
 Error
 Corrected Total

DF
15
742
757

28,
243,
272,
Sum of
Squares
.9931483
.8524755
.8456238

Mean
1.
0.


Square
9328766
3286422


F Val
5.



ue
88



Pr > F
<.0001


 Source
 year
 CBSEG_2003
DF
 9
 6
Type III SS
12.18939188
16.81566976
Mean Square
 1.35437688
 2.80261163
Value  Pr > F
 4.12  <.0001
 8.53  <.0001
Standardizing the residuals by estimates of standard deviation by  segment-group and year
appears to resolve the heterogeneous variance issue but yields little improvement on normality.

Tests for Normality  for  standardized residuals
Test
Shapi ro-Wilk
Kolmogorov-Smi rnov
         --Statistic	
         W     0.994055
         D     0.038676
                      	p Value	
                      Pr  <  W      0.0044
                      Pr  >  D     <0.0100
Levene's test for standardized residuals

Dependent Variable:  aStdResChl
 Source             DF
 Model              15
 Error             742
 Corrected Total   757
                 Sum of
                Squares
              3.7421584
            251.8744219
            255.6165803
                      Mean  Square
                        0.2494772
                        0.3394534
                     F Value
                        0.73
              p-value
              0.7499
                                                                             56

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                             Appendix F.
    SAS Computer Code for James  River Chlorophyll a
        Normality Tests,  Spring and Summer Season
* PROGRAM:  JAMES_RIVER. sas
* This program will TEST CHLOROPHYLL DATA FOR NORMALITY
* Base code from Elgin Perry  02/16/2010
* additional code written by  Jackie Johnson 02/17/2010
***********************************.
                                  i
libname ALGAE "G: \LR\OTHER_LR_DATA\Criteria_work\chlorophyll\2010";
*libname ALGAE "C:\Projects\CBP\CHLCRIT\LogNormal\";
options ls=72;
^OPTIONS LS=120 PS=55 REPLACE NOCENTER;
OPTIONS formchar = '  ---- | + | --- +=|-/\<>*';
*PROC IMPORT OUT= ALGAE . JAMES_SPRING_CHL
            DATATABLE= " JAMES_SPRING_CHL"
            DBMS=ACCESS2000 REPLACE;
*
DATABASE="G: \LR\OTHER_LR_DATA\Criteria_work\chlorophyll\2010\james_river.mdb"
f
*RUN;
*PROC CONTENTS DATA=ALGAE . JAMES_SPRING_CHL;RUN;
data one;
set  ALGAE. JAMES_SPRING_CHL;
logE Chl=log ( reported value);
label logE_Chl="LOG_E~UG/LITER" ;
  if cbseg_2003 = "APPTF"
  or cbseg_2003 = "CHKOH"
  or cbseg_2003 = "EBEMH"
  or cbseg_2003 = "LAFMH"
  or cbseg_2003 = "WBEMH" then  delete;

  if cbseg_2003 = "JMSPH" then  SegGrp = 1;
  if cbseg_2003 = "JMSMH"
  or cbseg_2003 = "SBEMH" then  SegGrp = 2;
  if cbseg_2003 = "JMSOH"
  or cbseg  2003 = "CB8PH"
  or cbseg_2003 = "ELIPH" then  SegGrp = 3;
  if cbseg_2003 = "JMSTF" then  SegGrp = 4;
  sdate = DatePart (SAMPLE_DATE) ;
  year = year(SDATE);
RUN;
*Proc Contents;
run;
title "Spring James River Data  1991-2000";
proc glm data=one;
  class year cbseg 2003;
  model logE_Chl=year cbseg_2003;
  output out= reschl r=rchl;
run;
proc Univariate normal plot data=work . reschl ;
  title2 "Normality test on raw residuals";
                                                                        57

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  var rchl;
run;
*( step to get rid of heterogeneous variances;
Proc Sort data=reschl;
  by SegGrp year;
  run;
Proc Means data=reschl noprint;
  by SegGrp year;
  var rchl;
  output out=sdchl StdDev = sdchl n=n;
run;
*(proc print data=sdchl;
*(  title "standard deviation results";
*(  var SegGrp year sdchl n;
data ResChi;
  merge ResChl sdchl;
  by SegGrp year;
  StdResChl = rchl/sdchl;
run;

proc Univariate normal plot data=work.reschl;
  title2 "Normality test on standardized residuals";
  var StdResChl;
run;
*(proc contents data=work.reschl;
*(run;
data four;
  set work.reschl;
  aRChl= abs(RChl);
  aStdResChl= abs(StdResChl);
run;
proc glm data=work.four;
  title2 "Levene's test for both types of residuals";
  class year cbseg_2003;
  model aRChl aStdResChl=year cbseg_2003;
  *(means cbseg_2003/snk;
  Mlsmeans cbseg_2003;
run;
* PROGRAM: JAMES_RIVER.sas
* This program will TEST CHLOROPHYLL DATA FOR NORMALITY
* Base code from Elgin Perry 02/16/2010
* additional code written by Jackie Johnson 02/17/2010
************************************
                                    r
libname ALGAE "G:\LR\OTHER_LR_DATA\Criteria_work\chlorophyll\2010";
*libname ALGAE "C:\Projects\CBP\CHLCRIT\LogNormal\";
options ls=72;
^OPTIONS LS=120 PS=55 REPLACE NOCENTER;
OPTIONS  formchar = '	| + |	+=|-/\<>*';
*PROC IMPORT OUT= ALGAE.JAMES_SPRING_CHL
            DATATABLE= "JAMES_SPRING_CHL"
            DBMS=ACCESS2000 REPLACE;
                                                                             58

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DATABASE="G:\LR\OTHER_LR_DATA\Criteria_work\chlorophyll\2010\james_river.mdb"
f
*RUN;
*PROC CONTENTS DATA=ALGAE.JAMES_SUMMER_CHL;RUN;
data one;
set  ALGAE.JAMES_SUMMER_CHL;
logE Chl=log(reported value);
label logE_Chl="LOG_E~UG/LITER";
  if cbseg_2003 = "APPTF"
  or cbseg_2003 = "CHKOH"
  or cbseg_2003 = "EBEMH"
  or cbseg_2003 = "LAFMH"
  or cbseg_2003 = "WBEMH" then delete;

  if cbseg_2003 = "JMSPH" then SegGrp = 1;
  if cbseg 2003 = "JMSMH"
  or cbseg_2003 = "SBEMH" then SegGrp = 2;
  if cbseg_2003 = "JMSOH"
  or cbseg_2003 = "CB8PH"
  or cbseg_2003 = "ELIPH" then SegGrp = 3;
  if cbseg_2003 = "JMSTF" then SegGrp = 4;
  sdate = DatePart(SAMPLE_DATE);
  year = year(SDATE);
RUN;
*Proc Contents;
run;
title "Summer James River Data 1991-2000";
proc glm data=one;
  class year cbseg 2003;
  model logE_Chl=year cbseg_2003;
  output out= reschl r=rchl;
run;
proc Univariate normal plot data=work.reschl;
  title2 "Normality test on raw residuals";
  var rchl;
run;
*( step to get rid of heterogeneous variances;
Proc Sort data=reschl;
  by SegGrp year;
  run;
Proc Means data=reschl noprint;
  by SegGrp year;
  var rchl;
  output out=sdchl StdDev = sdchl n=n;
run;
*(proc print data=sdchl;
*(  title "standard deviation results";
*(  var SegGrp year sdchl n;
data ResChi;
  merge ResChl sdchl;
  by SegGrp year;
  StdResChl = rchl/sdchl;
run;

proc Univariate normal plot data=work.reschl;
  title2 "Normality test on standardized residuals";
                                                                            59

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  var StdResChl;
run;
*(proc contents data=work.reschl;
*(run;
data four;
  set work.reschl;
  aRChl= abs(RChl);
  aStdResChl= abs(StdResChl);
run;
proc glm data=work.four;
  title2 "Levene's test for both types of residuals";
  class year cbseg_2003;
  model aRChl aStdResChl=year cbseg_2003;
  *(means cbseg_2003/snk;
  *(lsmeans cbseg 2003;
run;
                                                                             60

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               May 2010
  U.S. Environmental Protection Agency
               Region III
     Chesapeake Bay Program Office
          Annapolis, Maryland

                  and

               Region III
        Water Protection Division
       Philadelphia, Pennsylvania

          in coordination with

            Office of Water
    Office of Science and Technology
           Washington, D.C.

                  and

             The states of
     Delaware, Maryland, New York
       Pennsylvania, Virginia and
West Virginia and the District of Columbia

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