Diagnostic Accuracy of a Loop-Mediated Isothermal PCR Assay for Detection of Orientia tsutsugamushi during Acute Scrub Typhus Infection

There is an urgent need for alternative diagnostic methods for scrub typhus, but evaluation of these is hampered because the current serological gold standard (IFA) is imperfect. In a study from Thailand, 3 of 20 (15%) patients with fever had a positive Orientia tsutsugamushi PCR result despite negative serology. These findings could reflect potential benefits of the PCR assay in detecting rickettsaemia before antibody responses set in and/or a diagnostic advantage in endemic areas with high background levels of antibody in the population. Serology is complicated by the heterogeneity of strains present in Southeast Asia, but high resource costs and training make realtime PCR assays impractical for many areas where scrub typhus is endemic. This is where the new LAMP methodology has potential: it is inexpensive, simple to perform and requires only a waterbath or simple heating block instead of a thermocycler. In the context of a prospective fever study in a scrub typhus-endemic area in Thailand, the results support the validity of LAMP methodology for the diagnosis of scrub typhus, highlight the difficulties in comparing antibody- with DNA-based methods and also contribute towards understanding the dynamics of bacteraemia in this under recognised and under studied disease

Genome Sequence of Kitasatospora setae NBRC 14216T: An Evolutionary Snapshot of the Family Streptomycetaceae

Kitasatospora setae NBRC 14216T (=KM-6054T) is known to produce setamycin (bafilomycin B1) possessing antitrichomonal activity. The genus Kitasatospora is morphologically similar to the genus Streptomyces, although they are distinguishable from each other on the basis of cell wall composition and the 16S rDNA sequence. We have determined the complete genome sequence of K. setae NBRC 14216T as the first Streptomycetaceae genome other than Streptomyces. The genome is a single linear chromosome of 8 783 278 bp with terminal inverted repeats of 127 148 bp, predicted to encode 7569 protein-coding genes, 9 rRNA operons, 1 tmRNA and 74 tRNA genes. Although these features resemble those of Streptomyces, genome-wide comparison of orthologous genes between K. setae and Streptomyces revealed smaller extent of synteny. Multilocus phylogenetic analysis based on amino acid sequences unequivocally placed K. setae outside the Streptomyces genus. Although many of the genes related to morphological differentiation identified in Streptomyces were highly conserved in K. setae, there were some differences such as the apparent absence of the AmfS (SapB) class of surfactant protein and differences in the copy number and variation of paralogous components involved in cell wall synthesis

Expression of divIB of Bacillus subtilis during vegetative growth

Expression of the division initiation gene, divIB, of Bacillus subtilis vegetative growth was examined. lacZ fusion studies and transcription start point mapping have established that a sigma A promoter proximal to divIB is utilized in vivo. The -10 region of this promoter, which is located 93 bp upstream of the start codon, has been defined precisely by site-directed mutagenesis that destroys the promoter. Examination of transcripts by Northern (RNA) blotting has shown that there are at least two transcripts for divIB. The established proximal promoter was found to give rise to a very minor transcript which could not be convincingly demonstrated in wild-type cells but which became apparent upon insertion of a plasmid into the chromosome just upstream of this promoter. The major transcript for divIB originated from a site several kb upstream of the gene and is probably the same as the long polycistronic message also traversing the murD-spoVE-murG genes that was identified previously by others (A.D. Henriques, H. de Lencastre, and P.J. Piggot, Biochimie 74:735-748, 1992). Transcription from the proximal promoter alone, in an upstream-deletion mutant strain, provided sufficient DivIB for normal growth and division as well as sporulation

Primary myoepithelial carcinoma of the lung: a rare entity treated with parenchymal sparing resection

Primary lung myoepithelial carcinomas are rare neoplasms arising from the salivary glands of the respiratory epithelium. Given the rare occurrences and reports of these tumors, appropriate recommendations for resection are difficult to formulate. Although classified as low-grade neoplasms, these tumors have a significant rate of recurrence and distant metastasis

Orientia tsutsugamushi in Human Scrub Typhus Eschars Shows Tropism for Dendritic Cells and Monocytes Rather than Endothelium

Scrub typhus is a common and underdiagnosed cause of febrile illness in Southeast Asia, caused by infection with Orientia tsutsugamushi. Inoculation of the organism at a cutaneous mite bite site commonly results in formation of a localized pathological skin reaction termed an eschar. The site of development of the obligate intracellular bacteria within the eschar and the mechanisms of dissemination to cause systemic infection are unclear. Previous postmortem and in vitro reports demonstrated infection of endothelial cells, but recent pathophysiological investigations of typhus patients using surrogate markers of endothelial cell and leucocyte activation indicated a more prevalent host leucocyte than endothelial cell response in vivo. We therefore examined eschar skin biopsies from patients with scrub typhus to determine and characterize the phenotypes of host cells in vivo with intracellular infection by O. tsutsugamushi, using histology, immunohistochemistry, double immunofluorescence confocal laser scanning microscopy and electron microscopy. Immunophenotyping of host leucocytes infected with O. tsutsugamushi showed a tropism for host monocytes and dendritic cells, which were spatially related to different histological zones of the eschar. Infected leucocyte subsets were characterized by expression of HLADR+, with an “inflammatory” monocyte phenotype of CD14/LSP-1/CD68 positive or dendritic cell phenotype of CD1a/DCSIGN/S100/FXIIIa and CD163 positive staining, or occasional CD3 positive T-cells. Endothelial cell infection was rare, and histology did not indicate a widespread inflammatory vasculitis as the cause of the eschar. Infection of dendritic cells and activated inflammatory monocytes offers a potential route for dissemination of O. tsutsugamushi from the initial eschar site. This newly described cellular tropism for O. tsutsugamushi may influence its interaction with local host immune responses