A Novel Loop-Mediated Isothermal Amplification Assay for Rapid Detection of Yersinia pestis.

Rapid detection of Yersinia pestis, the causative agent of plague, is essential during field investigations to enable prompt control measures for prevention of the spread of the disease. Affordable, efficient, reliable, and simple detection assays are extremely useful, particularly in plague-endemic regions with limited resources. We developed a loop-mediated isothermal amplification (LAMP) assay that detects Y. pestis within 30 min by simply incubating at 65°C on a dry bath heater. The assay targeted the caf1A gene that is situated on the pMT1 plasmid using six specific primers. Y. pestis presence is visually detected based on the color change in the reactions. For comparison of the assay performance, a real-time LAMP with fluorescent dye detection was conducted on a real-time PCR instrument using the same six primers. Sensitivity assessment showed that the limit of detection (LOD) was 0.2 and 0.03 pg when performed on the dry bath heater and on the real-time PCR instrument, respectively. The assay was 100% specific, having no cross-reactivity with closely related Yersinia spp. and other bacterial species. We tested the LAMP assay on field-collected fleas and showed that it successfully detected Y. pestis with identical results to that of a previously published pentaplex real-time PCR assay. These findings suggest that the relatively inexpensive and simpler LAMP assay could be used to support field investigations, yielding comparable results to more expensive and complex PCR assays.

Human Exposure to Novel Bartonella Species from Contact with Fruit Bats.

Twice a year in southwestern Nigeria, during a traditional bat festival, community participants enter designated caves to capture bats, which are then consumed for food or traded. We investigated the presence of Bartonella species in Egyptian fruit bats (Rousettus aegyptiacus) and bat flies (Eucampsipoda africana) from these caves and assessed whether Bartonella infections had occurred in persons from the surrounding communities. Our results indicate that these bats and flies harbor Bartonella strains, which multilocus sequence typing indicated probably represent a novel Bartonella species, proposed as Bartonella rousetti. In serum from 8 of 204 persons, we detected antibodies to B. rousetti without cross-reactivity to other Bartonella species. This work suggests that bat-associated Bartonella strains might be capable of infecting humans.

Bartonella melophagi in blood of domestic sheep (Ovis aries) and sheep keds (Melophagus ovinus) from the southwestern US: Cultures, genetic characterization, and ecological connections.

Bartonella melophagi sp. nov. was isolated from domestic sheep blood and from sheep keds (Melophagus ovinus) from the southwestern United States. The sequence analyses of the reference strain performed by six molecular markers consistently demonstrated that B. melophagi relates to but differ from other Bartonella species isolated from domestic and wild ruminants. Presence of 183 genes specific for B. melophagi, being absent in genomes of other Bartonella species associated with ruminants also supports the separation of this bacterial species from species of other ruminants. Bartonella DNA was detected in all investigated sheep keds; however, culturing of these bacteria from sheep blood rejects a speculation that B. melophagi is an obligatory endosymbiont. Instead, the results support the hypothesis that the domestic sheep is a natural host reservoir for B. melophagi and the sheep ked its main vector. This bacterium was not isolated from the blood of bighorn sheep and domestic goats belonging to the same subfamily Caprinae. B. melophagi has also been shown to be zoonotic and needs to be investigated further.