Aims:  To characterize Francisella isolated from two natural warm springs in Utah and compare them to a strain isolated from a patient with probable exposure to one of the springs in 2001. Methods and Results:  A total of 39 presumptive Francisella isolates were obtained from two springs, Wasatch Hot Spring and Hobo Warm Spring, just north of Salt Lake City, Utah. All isolates were characterized by a combination of biochemical and molecular analyses, including novel PCR/electrospray ionization‐mass spectrometry (ESI‐MS) typing assays. Thirty‐one were identified as F. philomiragia, while the remaining eight were identified as F. tularensis ssp. novicida. Phylogenetic analysis of the 16S rRNA sequences revealed 27 isolates, which clustered with F. philomiragia, albeit into two distinct clades. The remaining isolates clustered along with other F. tularensis strains including the Utah clinical isolate. Testing with the PCR/ESI‐MS assays confirmed the identities of the isolates, but both yielded DNA signatures distinct from that of the clinical isolate. Conclusion:  We were successful in isolating several Francisella strains from natural warm springs; however, none appeared to genetically match the original 2001 clinical isolate. Significance and Impact of the Study:  This work highlights the presence of viable, potentially pathogenic Franscisella species living in the unique environmental niche of natural warm springs.

The Ibis T5000 is a novel diagnostic platform that couples PCR and mass spectrometry. In this study, we developed an assay that can identify all known pathogenic Vibrio species and field-tested it using natural water samples from both freshwater lakes and the Georgian coastal zone of the Black Sea. Of the 278 total water samples screened, 9 different Vibrio species were detected, 114 (41%) samples were positive for V. cholerae, and 5 (0.8%) samples were positive for the cholera toxin A gene (ctxA). All ctxA-positive samples were from two freshwater lakes, and no ctxA-positive samples from any of the Black Sea sites were detected.

Polymerase chain reaction/electrospray ionization-mass spectrometry (PCR/ESI-MS, previously known as “TIGER”) utilizes PCR with broad-range primers to amplify products from a wide array of organisms within a taxonomic group, followed by analysis of PCR amplicons using mass spectrometry. Computer analysis of precise masses allows for calculations of base compositions for the broad-range PCR products, which can then be compared to a database for identification. PCR/ESI-MS has the benefits of PCR in sensitivity and high-throughput capacity, but also has the distinct advantage of being able to detect and identify organisms with no prior characterization or sequence data. Existing broad range PCR primers, designed with an emphasis on human pathogens, were tested for their ability to amplify DNA of well characterized phytobacterial strains, as well as to populate the existing PCR/ESI-MS bacterial database with base counts. In a blinded panel study, PCR/ESI-MS successfully identified 93% of unknown bacterial DNAs to the genus level and 73% to the species/subspecies level. Additionally, PCR/ESI-MS was capable of detecting and identifying multiple bacteria within the same sample. The sensitivity of PCR/ESI-MS was consistent with other PCR based assays, and the specificity varied depending on the bacterial species. Preliminary tests with real life samples demonstrate a high potential for using PCR/ESI-MS systems for agricultural diagnostic applications.