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Plague is characterized by geographical foci from which it re-emerges after decades of silence, a fact currently explained by enzootic and epizootic cycles between plague-susceptible and plague-resistant rodents. To assess the potential role of soil in plague epidemiology, we experimentally investigated whether Yersinia pestis could persist alive and virulent in soil. Sterilized soil inoculated with virulent Y. pestis biotype Orientalis was regularly sampled for 40 weeks in duplicate. Each sample was observed by acridine orange staining and immunofluorescence using an anti-Y, pestis polyclonal antibody, and DNA was extracted for PCR amplification and sequencing of the Y. pestis ureD, caf1 and pla genes. All samples were inoculated onto selective agar, and samples from soil that had been incubated for 10, 60, 165, 210 and 280 days were also inoculated into each of two BALB/c female mice. The mouse experiment was performed in triplicate. Non-inoculated, sterilized soil samples were used as negative controls. Microorganisms fluorescing orange and detected by immunofluorescence were identified as Y. pestis in all samples. They were recovered in pure agar cultures for up to 30 weeks but thereafter were contaminated with Pseudomonas spp. Soil that had been inoculated with Y. pestis proved to be fully virulent in mice, which died with Y. pestis septicaemia and multiple organ involvement. Negative control mice showed no signs of disease. These data indicate that Y. pestis biotype Orientalis can remain viable and fully virulent after 40 weeks in soil. This study is a first step on which to base further investigations of a potential telluric reservoir for Y. pestis, which could represent an alternative mechanism for the maintenance of plague foci.

Abstract Background Bacteria of the genus Bartonella are responsible for a large variety of human and animal diseases. Serological typing of Bartonella is a method that can be used for differentiation and identification of Bartonella subspecies. Results We have developed a novel multiple antigenic microarray to serotype Bartonella strains and to select poly and monoclonal antibodies. It was validated using mouse polyclonal antibodies against 29 Bartonella strains. We then tested the microarray for serotyping of Bartonella strains and defining the profile of monoclonal antibodies. Bartonella strains gave a strong positive signal and all were correctly identified. Screening of monoclonal antibodies towards the Gro EL protein of B. clarridgeiae identified 3 groups of antibodies, which were observed with variable affinities against Bartonella strains. Conclusion We demonstrated that microarray of spotted bacteria can be a practical tool for serotyping of unidentified strains or species (and also for affinity determination) by polyclonal and monoclonal antibodies. This could be used in research and for identification of bacterial strains.

Ivermectin has emerged as very promising pediculicide, particularly in cases of resistance to commonly used pediculicides. Recently, however, the first field-evolved ivermectin-resistance in lice was reported. To gain insight into the mechanisms underlying ivermectin-resistance, we both looked for mutations in the ivermectin-target site (GluCl) and searched the entire proteome for potential new loci involved in resistance from laboratory susceptible and ivermectin-selected resistant body lice. Polymorphism analysis of cDNA GluCl showed no non-silent mutations. Proteomic analysis identified 22 differentially regulated proteins, of which 13 were upregulated and 9 were downregulated in the resistant strain. We evaluated the correlation between mRNA and protein levels by qRT-PCR and found that the trend in transcriptional variation was consistent with the proteomic changes. Among differentially expressed proteins, a complexin i.e. a neuronal protein which plays a key role in regulating neurotransmitter release, was shown to be the most significantly down-expressed in the ivermectin-resistant lice. Moreover, DNA-mutation analysis revealed that some complexin transcripts from resistant lice gained a premature stop codon, suggesting that this down-expression might be due, in part, to secondary effects of a nonsense mutation inside the gene. We further confirmed the association between complexin and ivermectin-resistance by RNA-interfering and found that knocking down the complexin expression induces resistance to ivermectin in susceptible lice. Our results provide evidence that complexin plays a significant role in regulating ivermectin resistance in body lice and represents the first evidence that links complexin to insecticide resistance., Competing Interests: The authors have declared that no competing interests exist.

Tropheryma whipplei, the agent of Whipple's disease, is a gram-positive rod-shaped bacterium that belongs to the group of actinobacteria. In order to produce monoclonal antibodies (MAbs) against this bacterium, we inoculated mice with two different strains, Slow2 and Endo5. We produced 13 and 10 MAbs against Slow2 and Endo5, respectively. Nine of the Slow2 MAbs and seven of the Endo5 MAbs recognized a 58-kDa epitope. In addition, three other Endo5 MAbs detected a unique 84-kDa epitope. These MAbs were species specific, as they did not react with a selection of 22 different bacterial species, but they were not strain specific, as they did react with six other strains of T. whipplei. Two-dimensional gel electrophoresis (2-DE) was combined with mass spectrometry (MS) to identify the 58-kDa and 84-kDa epitopes recognized by MAbs. After trypsin in-gel digestion of the spot, the 58-kDa protein was identified as an ATP synthase F1 complex beta chain, whereas the 84-kDa protein was identified as a polyribonucleotide nucleotidyltransferase by MS with matrix-assisted laser desorption ionization-time of flight. In an in vitro model, one of these MAbs allowed good detection of T. whipplei in stool samples, contrary to a rabbit polyclonal antibody, which led to high fluorescent background. In the prospective studies, the produced MAb will be tested for detection of T. whipplei in clinical samples, and the gene coding for identified 58-kDa and 84-kDa antigens will be tentatively cloned and then tested for its use in a diagnostic enzyme-linked immunosorbent assay for Whipple's disease.