The virulence plasmids of the equine virulent strains Rhodococcus equi ATCC 33701 and 103 were sequenced, and their genetic structure was analyzed. p33701 was 80,610 bp in length, and p103 was 1 bp shorter; their sequences were virtually identical. The plasmids contained 64 open reading frames (ORFs), 22 of which were homologous with genes of known function and 3 of which were homologous with putative genes of unknown function in other species. Putative functions were assigned to five ORFs based on protein family characteristics. The most striking feature of the virulence plasmids was the presence of a 27,536-bp pathogenicity island containing seven virulence-associated protein (vap) genes, including vapA. These vap genes have extensive homology to vapA, which encodes a thermoregulated and surface-expressed protein. The pathogenicity island contained a LysR family transcriptional regulator and a two-component response regulator upstream of six of the vap genes. The vap genes were present as a cluster of three (vapA, vapC, and vapD), as a pair (vapE and vapF), or individually (vapG; vapH). A region of extensive direct repeats of unknown function, possibly associated with thermoregulation, was present immediately upstream of the clustered and the paired genes but not the individual vap genes. There was extensive homology among the C-terminal halves of all vap genes but not generally among the N-terminal halves. The remainder of the plasmid consisted of a large region which appears to be associated with conjugation functions and a large region which appears to be associated with replication and partitioning functions.Rhodococcus equi is an important pulmonary pathogen of foals and is increasingly isolated from pneumonic infections and other infections in human immunodeficiency virus (HIV)-infected patients (19,33). Isolates from foals possess a large virulence plasmid, varying in size from 80 to 90 kb (45,47,49). Isolates lacking the plasmid are avirulent to foals (16,51). Little is known about the function of the plasmid apart from its encoding a virulence-associated surface protein (VapA) (45,49), the presence of a family of four vap genes (5), and the origin of replication (53). Infection with R. equi bacteria carrying the virulence plasmid may lead to immunomodulation in foals by causing failure to mount an effective Th1-based cellular immune response, but the basis of this effect is undefined (17). The expression of VapA is thermoregulated (Ն34°C) and pH regulated (41, 42), so that in this respect the plasmid has similarities to the virulence plasmids of pathogenic Yersinia species, such as Yersinia pestis, and of Shigella species (11,22,30). The plasmid is of significant interest, since it is associated with survival of the bacterium inside macrophages (16,21,33). Understanding its structure and function may therefore yield insights not only into the basis of virulence of this organism but also into the mechanisms of macrophage survival of other facultative intracellular pathogens, including Mycobacterium tub...

We investigated the prevalence of virulent Rhodococcus equi in clinical isolates from 69 sporadic cases (60 men, 8 women, and 1 patient of unknown sex) in Chiang Mai, Thailand, between 1993 and 2001. Fifty were human immunodeficiency virus (HIV) positive, 3 were HIV negative, and HIV status was unknown for 16. Fifty-two (75%) of 69 isolates were strains of intermediate virulence that contained the virulence-associated 20-kDa antigen, and 17 isolates (25%) were avirulent. No virulent strains with the virulence-associated 15-17-kDa antigens were identified. R. equi was isolated from HIV-positive patients' houses and those of their neighbors: avirulent strains were widespread, but only 1 strain of intermediate virulence was isolated. R. equi strains of intermediate virulence were isolated from 4 (0.8%) of 500 submaxillary lymph nodes from apparently healthy pigs in Chiang Mai. The routes of R. equi acquisition should be investigated from the viewpoint of zoonosis and public health.

Campylobacter jejuni infection is a leading cause of bacterial gastroenteritis in the United States and is acquired primarily through the ingestion of contaminated poultry products. Here, we describe the C. jejuni orthologue of ZnuA in other gram-negative bacteria. ZnuA (Cj0143c) is the periplasmic component of a putative zinc ABC transport system and is encoded on a zinc-dependent operon with Cj0142c and Cj0141c, which encode the other two likely components of the transport system of C. jejuni. Transcription of these genes is zinc dependent. A mutant lacking Cj0143c is growth deficient in zinc-limiting media, as well as in the chick gastrointestinal tract. The protein is glycosylated at asparagine 28, but this modification is dispensable for zinc-limited growth and chick colonization. Affinity-purified FLAG-tagged Cj0143c binds zinc in vitro. Based on our findings and on its homology to E. coli ZnuA, we conclude that Cj0143c encodes the C. jejuni orthologue of ZnuA.

The systematics of benign and moderately pathogenic Theileria isolates from cattle and deer originating from different geographic regions was undertaken by small-subunit ribosomal RNA (SSU rRNA) gene nucleotide-sequence analysis. A maximum-likelihood phylogenetic tree constructed from these sequences resulted in two major divisions, each with a common ancestor. One major division branches into four relatively divergent groups, including (1) bovine Theileria sp. Type D (USA and Korea), (2) T. mutans Intona and Theileria sp. MSD (Africa), (3) T. cervi (USA), and (4) well-characterized pathogenic Theileria spp. (Africa). The other major division branches into two groups: (1) T. buffeli Warwick and T. buffeli Marula and (2) a second branch of closely related isolates with SSU rRNA gene Types B, B1, C, E, and H. Putative geographically associated diversity was noted only in the Korean bovine Theileria spp. with SSU rRNA gene types C and H and in African T. mutans Intona and Theileria sp. MSD. The current results show that the United States bovine Theileria isolates are not T. mutans because they have T. buffeli Marula (Type A) and/or Type D (species undesignated) SSU rRNA gene sequences. The taxonomic separation of T. buffeli Warwick from African T. mutans is confirmed in this study.

No abstract

Campylobacter is a normal inhabitant of the chicken gut. Pathogenic infection with this organism in humans is accompanied by severe inflammation of the intestinal mucosal surface. The aim of this study was to evaluate the ability of Lactobacillus gasseri SBT2055 (LG2055) to inhibit the adhesion and invasion of Campylobacter jejuni in vitro and to suppress C. jejuni colonization of chicks in vivo. Pretreatment with LG2055 significantly reduced adhesion to and invasion of a human epithelial cell line, Intestine 407, by C. jejuni 81–176. Methanol (MeOH)-fixed LG2055 also reduced infection by C. jejuni 81–176. However, proteinase K (ProK)-treated LG2055 eliminated the inhibitory effects. Moreover, LG2055 co-aggregated with C. jejuni 81–176. ProK treatment prevented this co-aggregation, indicating that the co-aggregation phenotype mediated by the proteinaceous cell-surface components of LG2055 is important for reducing C. jejuni 81–176 adhesion and invasion. In an in vivo assay, oral doses of LG2055 were administered to chicks daily for 14 days after oral inoculation with C. jejuni 81–176. At 14 days post-inoculation, chicks treated with LG2055 had significantly reduced cecum colonization by C. jejuni. Reduction in the number of C. jejuni 81–176 cells adhering to and internalized by human epithelial cells demonstrated that LG2055 is an organism that effectively and competitively excludes C. jejuni 81–176. In addition, the results of the chick colonization assay suggest that treatment with LG2055 could be useful in suppressing C. jejuni colonization of the chicks at early growth stages.

Abstract. Rhodococcus equi isolates (462) obtained from 64 soil samples collected on 5 R. equi-endemic horse-breeding farms and isolates from 100 infected foals in Texas were examined to determine the prevalence and genotypic diversity of virulence-associated plasmids. Isolates were tested for the presence of 15-17-kDa virulence-associated protein antigens (VapA) by immunoblotting and virulence-associated plasmids by PCR. Plasmid DNAs were isolated and analyzed by digestion with restriction endonucleases for estimation of size and comparison of polymorphisims. Rhodococcus equi were isolated from soil of all 5 farms; however, virulent R. equi were only isolated from 3 of the 5 farms and represented 18.8% (87 of 462) of total isolates. Of the 87 virulent soil isolates, 56 (64.5%) contained an 85-kb type I plasmid, 23 (26.4%) an 87-kb type I plasmid, 7 (8%) a newly defined 85-kb type III plasmid (Tx 43), and 1 (1.1%) a newly defined 85-kb type IV plasmid (Tx 47). Of the 100 isolates from infected foals, 96 were virulent. Of the 96 virulent isolates, 51 (53.1%) contained an 85-kb type I plasmid, 39 (40.6%) an 87-kb type I plasmid, 4 (4.2%) an 85-kb type III plasmid (Tx 43), and 2 (2.1%) an 85-kb type IV plasmid (Tx 47). There are at least 4 different R. equi virulenceassociated plasmids in Texas, 2 of which have not previously been described. Based upon virulence plasmid typing, there is geographic diversity among isolates of R. equi from clinical and environmental samples on horse-breeding farms in Texas. There is not a strong correlation between the presence of virulent R. equi in farm soils and the R. equi disease status of those farms.Rhodococcus equi is one of the most important bacterial pathogens of young foals. Infections caused by this organism are characterized by chronic, suppurative bronchopneumonia and enteritis associated with a high mortality rate. 1,7,18 It was previously reported that the 15-17-kDa virulence-associated protein antigens (VapA) of R. equi are associated with virulence in mice 15,19 and that the presence of an 85-or a 90-kb plasmid is essential for virulence and the expression of VapA. 10,14,[22][23][24] These virulence-associated antigens and virulence plasmids have been used as epidemiologic markers to identify virulent R. equi isolates from horses and their environment by Western blot (immunoblot) assay using a monoclonal antibody and plasmid profiles. 4,11,12,16,17,21 More recently, restriction enzyme digestion patterns of virulence plasmids in human and foal isolates from several countries were examined. 6,20 The digestion patterns divided the plasmids of virulent isolates into 5 closely related types. Three of the 5 types had already been reported in Canadian and Japanese isolates, 6 and 2 new types were identified in French and Japanese isolates. 9,20 These 5 types of plasmids were designated as 85-kb type I (p-REAT701), 85-kb type II (a new type), 87-kb type I (EcoRI and BamHI type 2), 6 87-kb type II (a new type), and 90-kb (pREL1). The 85-kb type I plasmid was found in isolat...

This study documented a region in the N-terminal portion of SeM that varies in a nonsynonymous manner. This information should be useful in molecular epidemiologic studies of S equi.