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Three strains of a novel Bartonella species (Bartonella tamiae) were isolated from human patients from Thailand. Sequence analysis of six chromosomal regions (16S rRNA, gltA, groEL, ftsZ, rpoB, and the intergenic spacer region) and phenotypical analysis supported the similarity of the three strains and placed them within the genus Bartonella separately from previously described species.Several species of the genus Bartonella cause numerous disorders, and most are thought to be zoonoses (1, 3). These Bartonella-associated illnesses occur worldwide, including in Asia, and they encompass a broad clinical spectrum, including fever, skin lesions, lymphadenopathy, endocarditis, and abnormalities of the central nervous system, liver, eye, and bone tissues (5). There has been a report indicating that humans are being exposed to Bartonella species in Thailand, though the investigators did not isolate the agents (7).We report the characterization of three Bartonella strains isolated from blood samples of patients from Khon Kaen Province, Thailand, that belong to a novel Bartonella species. These strains were identified during the screening of blood clot specimens from a prospective study performed to determine the etiology of febrile illnesses in Thailand. It is the first report of culture-confirmed Bartonella infection in humans in Thailand.Blood clots were separated from sera, stored at Ϫ70°C, and shipped to the U.S. CDC (Fort Collins, CO) for testing. Two approaches were used for the isolation of Bartonella from human clots: (i) blood clots were cocultivated with Vero E6 cells at 35°C with 5% carbon dioxide for 7 days and then subcultured onto rabbit blood-enriched agar and (ii) blood clots were inoculated into a preenrichment liquid, the Bartonella-Alphaproteobacteria growth medium (BAPGM) developed by Maggi et al. (6), and after 7 days of incubation at 35°C with 5% carbon dioxide were plated onto rabbit blood agar. The agar plates were incubated at 35°C with an aerobic atmosphere of 5% carbon dioxide for up to 30 days. The cultured bacteria were visualized with Gram stain by using standard light microscopy with an oil immersion objective at a magnification of 1,000ϫ.For negative staining, a drop of suspension was placed on a copper grid coated with Formvar-carbon film and allowed to adhere for 10 min. The grids with adherent bacteria were stained by placing them on a drop of 2% potassium-phosphotungstic acid and air dried. The grids were examined with a Phillips 201 electron microscope. For ultrathin sectioning, the bacterial suspension was fixed in a mixture of 2.5% formaldehyde, 0.1% glutaraldehyde, 0.03% trinitrophenol, and 0.03% CaCl 2 in 0.05 M cacodylate buffer (pH 7.2). Ultrathin sections were cut with a Leica-Reichert Ultracut S ultramicrotome, stained with 2% aqueous uranyl acetate and lead citrate, and examined with a Phillips 201 electron microscope.The MicroScan rapid anaerobe identification panel (Dade Behring, Inc., West Sacramento, CA) was used to test the activities of preformed bacterial enzymes in ...

Ecological and bacteriologic observations of small mammals captured in Dhaka, Bangladesh, indicated that Bartonella infections occurred in high prevalence among lesser bandicoot rats (Bandicota bengalensis), black rats (Rattus rattus), and house shrews (Suncus murinus). Sequence analysis of the citrate synthase gene of Bartonella isolates showed that small mammals in Bangladesh harbored a diverse assemblage of strains. Some cultures were genetically related to Bartonella elizabethae, a species identified from a human patient in the United States. Sequences of some other cultures from Bandicota and Rattus rats were identical to sequences of cultures from domestic rats in France, Portugal, and the United States. The finding of Bartonella species in a high proportion of the mammalian samples from Dhaka suggests the need to study whether these agents might be responsible for human cases of febrile illness of unknown etiology in Bangladesh and elsewhere in south Asia.

Our longitudinal study of plague dynamics was conducted in north-central New Mexico to identify which species in the community were infected with plague, to determine the spatial and temporal patterns of the dynamics of plague epizootics, and to describe the dynamics of Yersinia pestis infection within individual hosts. A total of 3156 fleas collected from 535 small mammals of 8 species were tested for Y. pestis DNA. Nine fleas collected from six southern plains woodrats (Neotoma micropus) and from one rock squirrel (Otospermophilus variegatus) were positive for the pla gene of Y. pestis. None of 127 fleas collected from 17 woodrat nests was positive. Hemagglutinating antibodies to the Y. pestis-specific F1 antigen were detected in 11 rodents of 6 species. All parts of the investigated area were subjected to local disappearance of woodrats. Despite the active die-offs, some woodrats always were present within the relatively limited endemic territory and apparently were never exposed to plague. Our observations suggest that small-scale die-offs in woodrats can support maintenance of plague in the active U.S. Southwestern focus.

One of the biggest challenges in the effort to treat and contain influenza A virus infections is the emergence of resistance during treatment. It is well documented that resistance to amantadine arises rapidly during the course of treatment due to mutations in the gene coding for the M2 protein. To address this problem, it is critical to develop experimental systems that can accurately model the selection of resistance under drug pressure as seen in humans. We used the hollow-fiber infection model (HFIM) system to examine the effect of amantadine on the replication of influenza virus, A/Albany/1/98 (H3N2), grown in MDCK cells. At 24 and 48 h postinfection, virus replication was inhibited in a dose-dependent fashion. At 72 and 96 h postinfection, virus replication was no longer inhibited, suggesting the emergence of amantadine-resistant virus. Sequencing of the M2 gene revealed that mutations appeared at between 48 and 72 h of drug treatment and that the mutations were identical to those identified in the clinic for amantadine-resistant viruses (e.g., V27A, A30T, and S31N). Interestingly, we found that the type of mutation was strongly affected by the dose of the drug. The data suggest that the HFIM is a good model for influenza virus infection and resistance generation in humans. The HFIM has the advantage of being a highly controlled system where multiplicity parameters can be directly and accurately controlled and measured.

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Thirty bartonella strains were isolated from the blood of black-tailed prairie dogs (Cynomys ludovicianus) from Boulder County, Colorado, USA. The bacteria appeared as small, fastidious, aerobic, Gram-negative rods. The partial sequences of the citrate synthase gene (gltA) demonstrated five unique genetic variants. Phylogenetic analysis based on sequences of gltA, 16S rRNA, rpoB, ftsZ, and ribC showed that the black-tailed prairie dog-related Bartonella variants comprise a distinct monophyletic clade that is closely related to Bartonella washoensis, a species isolated from a human patient and subsequently from ground squirrels. These variants, however, are grouped together in 100% of the bootstrapped trees. These variants were not found in other small mammals trapped during the same study, showing some evidence of host specificity. We believe that the group being described here is typical of the black-tailed prairie dog. We propose to name the bacteria Candidatus Bartonella washoensis subsp. cynomysii. The type strain is CL8606co(T)(=ATCC BAA-1342(T) = CCUG 53213(T)), which is the representative isolate of the dominant variant of the characterized group.

Coccidioidomycosis is an infection caused by Coccidioides immitis or C. posadasii . We developed a TaqMan real-time PCR assay that rapidly and accurately differentiates the species. This assay can be used as a tool to improve disease surveillance, increase understanding of the natural history of the infection, and assist in clinical differentiation studies.