SPADA Draft Documents

Additional considerations with the status quo testing of assays against inclusivity/exclusivity 101 panels 102 The SPADA inclusivity/exclusivity panels for the biodefense relevant bacterial pathogens, 103 such as Bacillus anthracis , Yersinia pestis , Brucella suis , Burkholderia mallei, Burkholderia 104 pseudomallei, and Francisella tularensis , comprise a total of approximately 100 strains. These 105 strains are used to validate the inclusivity/exclusivity criteria for the respective detection assays. 106 Most of the inclusivity strains, and some exclusivity strains, are considered Biosafety Level 3 107 (BSL3) agents and, as a result, are limited to labs that are registered and certified for such work. 108 Moreover, extensive testing adds cost and time to the assay development effort. Many whole 109 genome sequences of these bacterial strains now are available (2-6), which allows the in silico 110 evaluation of existing assays that are currently widely used in biosurveillance and a 111 representative analyses is illustrated in Figure 2. The majority of the evaluated assay signatures 112 had perfect sequence matches to the target inclusivity genome sequences, while they had less 113 (0% to 40%) sequence identity to the exclusivity panel genome sequences. For all the assays 114 evaluated, there was no “perfect” assay (i.e. no false positives and no false negatives). Some 115 assays are computationally predicted to have both false negatives (e.g., Bacillus anthracis assay 116 1 against strain 10 in inclusivity panel) and false positives (e.g., Bacillus anthracis assay 1 117 against strain 8 in exclusivity panel). Many of these predicted assay failures correspond to the 118 expected deviations based on the genotypes of these strains. There are some assays that simply 119 fail the inclusivity and/or exclusivity criteria (e.g., Bacillus anthracis assay 7 or Yersinia pestis 120 assay 15) and are therefore not reliable diagnostics due to low specificity. However, given the 121 high conservation of the assay signatures to the target strains in the inclusivity panel and their 122 low conservation in the exclusivity panel, the “brute force” testing of all available strains is not 123

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