Epizootic bovine abortion (EBA), first identified in the 1950s, is a major contributor of economic loss to western U.S. beef producers. The causative agent proved elusive for over fifty years until a novel Deltaproteobacteria was identified as the etiologic agent in 2005. The microbe, which has yet to be successfully cultured in vitro, has proven difficult to purify from necropsy tissues. Thus, phylogenetic characterization has been limited to analysis of the 16S ribosomal RNA (rRNA) gene (AF503916), which placed this bacterium in the order Myxococcales, suborder Sorangiineae, family Polyangiaceae and most closely related to Sorangium cellulosum. The focus of the current study was to further expand the morphologic characterization and taxonomic placement of this bacteria, named here as Pajaroellobacter abortibovis. Modified Gram staining, combined with transmission electron microscopy, provide strong evidence that the bacterium is gram negative. Flow cytometric analysis identified the presence of P. abortibovis in murine leukocytes. While attempts to sequence ten universally conserved protein-coding genes using previously published degenerative primers failed, redesigned primers based solely upon Deltaproteobacteria facilitated the partial sequencing of two genes; fusA (JQ173112) and pyrG (JQ173111). Primers designed in a similar fashion generated a partial sequence of the 23S rRNA gene (JQ173113) These sequences, combined with a revised 16S rRNA phylogenic analysis, support the placement of this bacteria as a unique genus separate from Sorangium.

Since the advent of highly pathogenic variants of avian influenza virus (HPAIV), the main focus of avian influenza research has been the characterization and detection of HPAIV hemagglutinin (HA) from H5 and H7 subtypes. However, due to the high mutation and reassortation rate of influenza viruses, in theory any influenza strain may acquire increased pathogenicity irrespective of its subtype. A comprehensive antigenic characterization of influenza viruses encompassing all 16 HA and 9 neuraminidase subtypes will provide information useful for the design of differential diagnostic tools, and possibly, vaccines. We have expressed recombinant HA proteins from 3 different influenza virus HA subtypes in the baculovirus system. These proteins were used to generate polyclonal rabbit antisera, which were subsequently employed in epitope scanning analysis using peptide libraries spanning the entire HA. Here, we report the identification and characterization of linear, HA subtype-specific as well as inter subtype-conserved epitopes along the HA proteins. Selected subtype-specific epitopes were shown to be suitable for the differentiation of anti-HA antibodies in an ELISA.

Abstract. Epizootic bovine abortion (EBA), also commonly known as "foothill abortion," is a late-term abortion primarily in beef cattle with significant economic impacts in California, Nevada, and Oregon. The causative agent is a novel deltaproteobacterium (aoEBA) closely related to the order Myxococcales and vectored by the soft-shelled tick Ornithodoros coriaceus. Historically, diagnosis has relied upon the pathologic examination of the fetus and the presence of elevated fetal serum immunoglobulins. Identification of the etiologic agent, a unique deltaproteobacterium, permitted the development of a quantitative duplex real-time polymerase chain reaction (qPCR) using a unique 90-bp sequence of aoEBA 16S ribosomal RNA gene in conjunction with an 88-bp sequence of the bovine β-actin gene. Reaction efficiencies were 100.9% for the 16S aoEBA gene and 93.1% for the bovine β-actin gene. Application of the duplex TaqMan to a set of aoEBA-infected fetal bovine necropsy tissues demonstrated the assay to be robust in quantitatively identifying the aoEBA bacteria and establishing hosttissue pathogen load. Consistent with previously reported immunohistochemical data, organized lymphoid tissue generally carried the heaviest bacterial load as compared to non-lymphoid tissue. The newly developed duplex TaqMan assay will facilitate diagnosis in difficult cases and provide an invaluable tool for delineating the pathogenesis of EBA.