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-1B1 score > 3.0. However, no sample size 
restriction was introduced. As a result, a large segment could contain a single B-1BI score, and if 
that single score exceeded 3, then the segment was classified as healthy. The likelihood of a 
degraded segment containing 10 B-1BI 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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