Kathryn E. Stockbauer

Subspecific differentiation of Mycobacterium avium complex strains by automated sequencing of a region of the gene (hsp65) encoding a 65-kilodalton heat shock protein

To develop a strategy for rapid species assignment and strain differentiation of Mycobacterium avium complex (MAC) organisms, the sequence of a 360-bp region of the gene (hsp65) encoding a 65-kDa heat shock protein was determined for 56 isolates, including 21 patient isolates and 35 reference strains. Eleven hsp65 alleles were identified, and there was no sharing of alleles between strains classified as M. avium and Mycobacterium intracellulare based on serovar and species-specific DNA hybridization probes. Phylogenetic analysis showed that 30 strains had one of two hsp65 alleles which were found in known M. avium organisms, 23 strains had one of six alleles allied with known M. intracellulare organisms, and three MAC isolates had one of three hsp65 alleles that differed substantially from the consensus M. avium and M. intracellulare hsp65 sequences. Estimates of strain relationships based on the sequences of hsp65 and the 16S-23S ribosomal DNA internal transcribed spacer were similar. Automated DNA sequencing of a 360-bp region of the hsp65 gene from MAC organisms provides a rapid and unambiguous marker system for strain differentiation and permits specific assignment of these acid-fast organisms for diagnostic purposes.

Rapidly Progressive, Fatal, Inhalation Anthrax-like Infection in a Human: Case Report, Pathogen Genome Sequencing, Pathology, and Coordinated Response

CONTEXT Ten years ago a bioterrorism event involving Bacillus anthracis spores captured the nations interest, stimulated extensive new research on this pathogen, and heightened concern about illegitimate release of infectious agents. Sporadic reports have described rare, fulminant, and sometimes fatal cases of pneumonia in humans and nonhuman primates caused by strains of Bacillus cereus , a species closely related to Bacillus anthracis. OBJECTIVES To describe and investigate a case of rapidly progressive, fatal, anthrax-like pneumonia and the overwhelming infection caused by a Bacillus species of uncertain provenance in a patient residing in rural Texas. DESIGN We characterized the genome of the causative strain within days of its recovery from antemortem cultures using next-generation sequencing and performed immunohistochemistry on tissues obtained at autopsy with antibodies directed against virulence proteins of B anthracis and B cereus. RESULTS We discovered that the infection was caused by a previously unknown strain of B cereus that was closely related to, but genetically distinct from, B anthracis . The strain contains a plasmid similar to pXO1, a genetic element encoding anthrax toxin and other known virulence factors. Immunohistochemistry demonstrated that several homologs of B anthracis virulence proteins were made in infected tissues, likely contributing to the patients death. CONCLUSIONS Rapid genome sequence analysis permitted us to genetically define this strain, rule out the likelihood of bioterrorism, and contribute effectively to the institutional response to this event. Our experience strongly reinforced the critical value of deploying a well-integrated, anatomic, clinical, and genomic strategy to respond rapidly to a potential emerging, infectious threat to public health.