Heavy metal effects on Proteus mirabilis Superoxide dismutase production

Abstract Levels of genomic DNA relatedness were determined using a SI nuclease procedure for reference bacteria identified as biotypes of Corynebacterium diphtheriae , biovars of Corynebacterium pseudotuberculosis , and ' Corynebacterium ulcerans '. These results showed that the three species are separate taxa at the genomospecies level whereas biotypes and biovars are closely related genomically within each species. Phylogenetic analyses of small-subunit rDNA sequences revealed that ' Corynebacterium ulcerans ' forms a tight cluster with Corynebacterium pseudotuberculosis within the robust branch that groups all Corynebacterium sequenced to date. Therefore, we propose that the species incertae sedis ' C. ulcerans ' should be conclusively recognized as a distinct species within the genus Corynebacterium with strain CCUG 2708 = NCTC 7910 as type strain. This species is characterized by urease production and fermentation of glycogen.     

In vivo insertional mutagenesis in Corynebacterium pseudotuberculosis: an efficient means to identify DNA sequences encoding exported proteins

The reporter transposon-based system TnFuZ was used to identify exported proteins of the animal pathogen Corynebacterium pseudotuberculosis. Thirty-four out of 1,500 mutants had detectable alkaline phosphatase (PhoZ) activity. This activity was from 21 C. pseudotuberculosis loci that code for fimbrial and transport subunits and for hypothetical and unknown-function proteins.     

Complete Genome Sequences of Corynebacterium pseudotuberculosis Strains 3/99-5 and 42/02-A, Isolated from Sheep in Scotland and Australia, Respectively

Here, we report the whole-genome sequences of two ovine-pathogenic Corynebacterium pseudotuberculosis isolates: strain 3/99-5, which represents the first C. pseudotuberculosis genome originating from the United Kingdom, and 42/02-A, the second from Australia. These genome sequences will contribute to the objective of determining the global pan-genome of this bacterium.     

Purification of 1-monoacylglycerols containing alpha-branched-beta-hydroxylated fatty acids from lipids of Corynebacterium pseudotuberculosis.

A complex mixture of monoacylglycerols, identified as (C30--C36) corynomycoloylglycerols was isolated from the diethyl ether/acetone-soluble lipids of Corynebacterium pseudotuberculosis.     

A description of genes of Corynebacterium pseudotuberculosis useful in diagnostics and vaccine applications

Corynebacterium pseudotuberculosis, a Gram-positive intracellular pathogen, is the etiological agent of caseous lymphadenitis or CLA. This bacterium infects goats and sheep and causes great economic losses worldwide annually, mainly for goat producers. Despite its importance, CLA is still poorly characterized. However, with advances in the genomic field, many C. pseudotuberculosis genes have already been characterized, mainly those related to virulence such as phospholipase D. Here, we examined the use of the several available genes of C. pseudotuberculosis and reviewed their applications in vaccine construction, more efficient diagnostics for CLA, and control of this disease, among other applications.     

Cloning and manipulation of the Corynebacterium pseudotuberculosis recA gene for live vaccine vector development

Abstract Corynebacterium pseudotuberculosis is an intracellular bacterial pathogen causing a chronic abscessing disease in sheep and goats called caseous lymphadenitis. We are developing this bacterial species as a live vector system to deliver vaccine antigens to the animal immune system. Foreign genes expressed in bacterial hosts can be unstable so we undertook to delete the C. pseudotuberculosis chromosomal recA gene to determine whether a recA ⁻ background would reduce the frequency of recombination in cloned DNA. Homologous DNA recombination within an isogenic recA ⁻ C. pseudotuberculosis was 10–12-fold lower than that in the recA ⁺ parental strain. Importantly, the recA mutation had no detectable affect upon the virulence of C. pseudotuberculosis in a mouse model. Taken together these results suggest that a recA ⁻ background may be useful in the further development of C. pseudotuberculosis as a vaccine vector.     

Targeted allele replacement mutagenesis of Corynebacterium pseudotuberculosis

A two-step allele replacement mutagenesis procedure, using a conditionally replicating plasmid, was developed to allow the creation of targeted, marker-free mutations in Corynebacterium pseudotuberculosis. The relationship between homologous sequence length and recombination frequency was determined, and enhanced plasmid excision was observed due to the rolling-circle replication of the mutagenesis vector. Furthermore, an antibiotic enrichment procedure was applied to improve the recovery of mutants. Subsequently, as proof of concept, a marker-free, cp40-deficient mutant of C. pseudotuberculosis was constructed.     

Genotyping of Yersinia pestis strains on the basis of variation of ribosomal rRNA biosynthesis genes

Analysis of restriction fragment length polymorphism of rRNA genes of Yersinia pestis and Y. pseudotuberculosis strains, circulating in Russian Federation and abroad revealed the effectiveness of ribotyping for differentiation between these microorganisms, as well as for differentiation between different Y. pestis biovars and main and nonmain subspecies of this agent. Use of this method was shown to be promising as a component for the complex molecular typing system of Y. pestis. Variant ribotypes of main and non-main subspecies of Y. pestis strains are presented.     

Effect of olive oil mill wastewater on extracellular ligninolytic enzymes produced by Phanerochaete flavido-alba

Four genes, fagA, B, C and D, encoding products with 32-47% identity to proteins involved in bacterial iron uptake systems, were identified immediately downstream of the Corynebacterium pseudotuberculosis phospholipase D gene. beta-Galactosidase assays on a C. pseudotuberculosis strain carrying a fagA-lacZ fusion indicated that the putative fagABC operon was poorly expressed in iron-rich media. However, similar experiments in iron-limited media resulted in an approximately three-fold increase in beta-galactosidase activity, suggesting that this operon is regulated by iron in vitro. Although no defect in iron utilization could be determined for a C. pseudotuberculosis fagB(C) mutant in vitro, this mutant showed reduced virulence compared to wild-type in a goat model of caseous lymphadenitis. Thus, expression of the fag genes in the host appears to contribute to virulence.     

Identification of macrophage induced genes of Corynebacterium pseudotuberculosis by differential fluorescence induction

Corynebacterium pseudotuberculosis is the etiological agent of the sheep disease caseous lymphadenitis. We have developed a promoter reporter system for this organism based on expression of the green fluorescent protein (gfp) gene from Aequorea victoria. A promoterless vector, pSM20, containing the gfp gene was constructed, and promoters were inserted upstream of the gfp gene. Upon transformation into C. pseudotuberculosis, fluorescence could be visualised by fluorescence microscopy, and relative promoter strength measured by flow cytometry. The usefulness of this system for measuring changes in gene expression was demonstrated by measuring fluorescence levels of heat shocked C. pseudotuberculosis carrying a dnaK promoter construct. Replication of C. pseudotuberculosis within J774 macrophages could be monitored by fluorescence microscopy. The establishment of the system allowed the use of differential fluorescence to identify genes that showed up-regulation following macrophage infection. Genes coding for a non-ribosomal peptide synthetase and the beta chain of a propionyl CoA carboxylase were identified as possessing promoters that demonstrated enhanced activity following macrophage infection by C. pseudotuberculosis.     

Complete genome sequence of Corynebacterium pseudotuberculosis strain Cp267, isolated from a llama

In this work we report the genome of Corynebacterium pseudotuberculosis strain 267, isolated from a llama. This pathogen is of great veterinary and economic importance, as it is the cause of caseous lymphadenitis in several livestock species around the world and causes significant losses due to the high cost of treatment.     

Multiplex polymerase chain reaction to identify and determine the toxigenicity of Corynebacterium spp with zoonotic potential and an overview of human and animal infections

Corynebacterium diphtheriae, Corynebacterium ulcerans and Corynebacterium pseudotuberculosis constitute a group of potentially toxigenic microorganisms that are related to different infectious processes in animal and human hosts. Currently, there is a lack of information on the prevalence of disease caused by these pathogens, which is partially due to a reduction in the frequency of routine laboratory testing. In this study, a multiplex polymerase chain reaction (mPCR) assay that can simultaneously identify and determine the toxigenicity of these corynebacterial species with zoonotic potential was developed. This assay uses five primer pairs targeting the following genes: rpoB (Corynebacterium spp), 16S rRNA (C. ulcerans and C. pseudotuberculosis), pld (C. pseudotuberculosis), dtxR (C. diphtheriae) and tox [diphtheria toxin (DT) ]. In addition to describing this assay, we review the literature regarding the diseases caused by these pathogens. Of the 213 coryneform strains tested, the mPCR results for all toxigenic and non-toxigenic strains of C . diphtheriae, C. ulcerans and C. pseudotuberculosis were in 100% agreement with the results of standard biochemical tests and PCR-DT. As an alternative to conventional methods, due to its advantages of specificity and speed, the mPCR assay used in this study may successfully be applied for the diagnosis of human and/or animal diseases caused by potentially toxigenic corynebacterial species.     

Draft Genome Sequence of Corynebacterium diphtheriae Biovar Intermedius NCTC 5011

We report an annotated draft genome of the human pathogen Corynebacterium diphtheriae bv. intermedius NCTC 5011. This strain is the first C. diphtheriae bv. intermedius strain to be sequenced, and our results provide a useful comparison to the other primary disease-causing biovars, C. diphtheriae bv. gravis and C. diphtheriae bv. mitis. The sequence has been deposited at DDBJ/EMBL/GenBank with the accession number AJVH01000000.     

Molecular characterization of Corynebacterium pseudotuberculosis isolated from goats using ERIC-PCR

Corynebacterium pseudotuberculosis, the infectious agent of caseous lymphadenitis (CLA), is responsible for substantial economic losses in goat and sheep production. Molecular characterization of C. pseudotuberculosis isolates by enterobacterial repetitive intergenic consensus (ERIC)-PCR has shown promising results in genotyping strains isolated from sheep with CLA. We evaluated the genetic diversity of C. pseudotuberculosis isolates collected from the Sert?o region of the Pernambuco (PE) State, Brazil, and investigated the potential of ERIC-PCR as a tool for the molecular typing of strains of C. pseudotuberculosis isolated from goats. Thirty-two C. pseudotuberculosis strains isolated from goats in the municipalities of Floresta and Ibimirim, PE, C. pseudotuberculosis type strain ATCC 19410, the 1002 vaccine strain, and a field isolate of Rhodococcus equi were fingerprinted using the primers ERIC-1R and ERIC-2 and the primer pair ERIC- 1R+ERIC-2. Using 100% similarity as the cutoff, 8, 10, and 7 genotypes were obtained with ERIC-1-PCR, ERIC-2-PCR, and ERIC-1+2-PCR, respectively. The Hunter-Gaston discriminatory index calculated for the ERIC-1-PCR was 0.75. The index for the ERIC-2-PCR was 0.88, and the index for the ERIC-1+2-PCR was 0.79. Among goat isolates of C. pseudotuberculosis, three, two and four genotypes (found by ERIC-1-PCR, ERIC-2-PCR, and ERIC-1+2-PCR, respectively) had been previously described among sheep isolates from Minas Gerais State, Brazil. These results showed that ERIC-PCR has good discriminatory power and typeability, making it a useful tool for discrimination among C. pseudotuberculosis isolates from goats.     

Whole-genome sequence of Corynebacterium pseudotuberculosis PAT10 strain isolated from sheep in Patagonia, Argentina

In this work, we report the complete genome sequence of a Corynebacterium pseudotuberculosis PAT10 isolate, collected from a lung abscess in an Argentine sheep in Patagonia, whose pathogen also required an investigation of its pathogenesis. Thus, the analysis of the genome sequence offers a means to better understanding of the molecular and genetic basis of virulence of this bacterium.     

Complement receptor type 3 plays an important role in development of protective immunity to primary and secondary Corynebacterium pseudotuberculosis infection in mice

The present study was performed to investigate the role of CR3, the type 3 complement receptor, in host defense against primary and secondary Corynebacterium (C.) pseudotuberculosis infection in mice. Treatment of mice with 5C6, an anti-CR3 monoclonal antibody (mAb), resulted in unrestricted multiplication of bacteria in the organs and dramatically increased mortalities of the infected mice. Histological examinations showed the inflammation, degeneration and necrosis of organs and revealed that the infection-enhancing effect of 5C6 mAb was associated with the failure of mice to focus mononuclear phagocytes at sites of bacterial multiplication. These results suggest that CR3 plays an important role in host defense against primary as well as secondary C. pseudotuberculosis infection in mice.     

Complete genome sequence of Corynebacterium pseudotuberculosis strain CIP 52.97, isolated from a horse in Kenya

In this work, we report the whole-genome sequence of Corynebacterium pseudotuberculosis bv. equi strain CIP 52.97 (Collection Institut Pasteur), isolated in 1952 from a case of ulcerative lymphangitis in a Kenyan horse, which has evidently caused significant losses to agribusiness. Therefore, obtaining this genome will allow the detection of important targets for postgenomic studies, with the aim of minimizing problems caused by this microorganism.     

Etiologic role of Corynebacterium non diphtheriae in patients with different pathology

Bacteriologic examination of 1589 patients showed that, aside from C. diphtheriae, 11% of acute upper respiratory tract infections were caused by other Corynebacterium species. Such bacteria can cause infections of various localizations (bronchitis, pyelonephritis, urethritis, colpitis, dermatitis, arthritis, etc.). C. pseudodiphtheriticum and C. xerosis were isolated from clinical specimens most frequently. Corynebacterium spp. have adhesive, hemolytic, hemagglutinating, and neuraminidase activity; some of them are highly pathogenic. The most virulent, were following species: C. diphtheriae, C. pseudotuberculosis, C. urealyticum, and C. ulcerans. Corynebacterium non diphtheriae were frequently isolated from clinical specimens in association with staphylococci and streptococci. In such cases, factors of pathogenicity and resistance to antibiotics were more pronounced. Strains isolated with association with other bacteria have lost susceptibility to tetracycline, oleandomycin, penicillin, and erythromycin. It is important to be vigilant about bacteria from Corynebacterium genus in clinical settings, and thoroughly study their biologic characteristics, especially in immunocompromised patients.     

Evaluation of three methods for DNA fingerprinting of Corynebacterium pseudotuberculosis strains isolated from goats in Poland

Phenotypic approaches based on metabolic and biological characteristics of Corynebacterium pseudotuberculosis have been limited due to insufficient discrimination between closely related isolates. In this paper we present performance and convenience of three molecular typing methods: BOX-PCR, random amplification of polymorphic DNA (RAPD) and amplification of DNA fragments surrounding rare restriction site (ADSRRS-fingerprinting) in genome analysis of these bacteria. Among examined 61 strains there were distinguished four, eight and 10 different genotypes by BOX-PCR, RAPD and ADSRRS-fingerprinting, respectively. The value of discrimination index was the lowest for BOX-PCR (D = 0.265), much bigger for RAPD (D = 0.539) and the highest for ADSRRS-fingerprinting (D = 0.604). The good discriminatory ability and reproducibility of RAPD and ADSRRS-fingerprinting indicates that those techniques may be particularly applied for epidemiological studies of C. pseudotuberculosis isolates. We found that ADSRRS-fingerprinting is a rapid method offering good discrimination power, excellent reproducibility and may be applied for epidemiological studies of intraspecific genetic relatedness of C. pseudotuberculosis strains.     

A Seroepidemiological Investigation of Corynebacterium Pseudotuberculosis Infection in Sheep Flocks in Southern Norway

Corynebacterium pseudotuberculosis causes caseous lymphadenitis, a disease primarily affecting sheep and goats which occurs as a chronic infection characterized by suppurative lymphadenitis (Lovell 1959, Marsh 1965, Jubb & Kennedy 1970, Ayers 1977, Addo & Eid 1978). The disease has an insidious nature and a high prevalence, resulting in large economic losses due to condemnation of carcasses and organs at slaughter (Turner 1973).