Whole-Genome Sequencing in Microbial Forensic Analysis of Gamma-Irradiated Microbial Materials.

Effective microbial forensic analysis of materials used in a potential biological attack requires robust methods of morphological and genetic characterization of the attack materials in order to enable the attribution of the materials to potential sources and to exclude other potential sources. The genetic homogeneity and potential intersample variability of many of the category A to C bioterrorism agents offer a particular challenge to the generation of attributive signatures, potentially requiring whole-genome or proteomic approaches to be utilized. Currently, irradiation of mail is standard practice at several government facilities judged to be at particularly high risk. Thus, initial forensic signatures would need to be recovered from inactivated (nonviable) material. In the study described in this report, we determined the effects of high-dose gamma irradiation on forensic markers of bacterial biothreat agent surrogate organisms with a particular emphasis on the suitability of genomic DNA (gDNA) recovered from such sources as a template for whole-genome analysis. While irradiation of spores and vegetative cells affected the retention of Gram and spore stains and sheared gDNA into small fragments, we found that irradiated material could be utilized to generate accurate whole-genome sequence data on the Illumina and Roche 454 sequencing platforms.

Coxiella burnetii serology assays in goat abortion storm.

Many commercial antibody detection enzyme-linked immunosorbent assay (ELISA) kits for Q fever utilize the Nine Mile (Montana tick) strain of Coxiella burnetii as antigen. An ELISA kit manufactured in France employs ovine placenta-sourced antigen and has been used in Europe. Sera from goats experiencing a Q fever abortion storm in the United States were used to compare the sensitivity and specificity of these 2 ELISA formats and the Q fever complement fixation test (CFT). Latent class estimates of sensitivity ranged from 97% to 100% with a specificity of 95-100% for the 2 ELISA kits. Estimates for sensitivity and specificity of the CFT were 89% and 82%, respectively. There was not a significant increase in ELISA sensitivity observed with the ovine-sourced antigen kit in this study. Real-time polymerase chain reactions performed on a portion of the sera found that 15 out of 20 sera were congruent across 4 tests for positive and negative sera.