B-Cell Epitope Mapping Using a Library of Overlapping Synthetic Peptides in an Enzyme-Linked Immunosorbent Assay.

This chapter describes a strategy for mapping linear B-cell epitopes of proteins using synthetic biotinylated peptides in an ELISA.A set of overlapping peptides were designed based upon a known amino acid sequence of the target protein, VapA (Virulence-associated Protein A) of the bacterium Rhodococcus equi, an important pulmonary pathogen in foals. The peptides synthesized as biotinylated peptides were coated directly onto micro titer plates which had been pre-coated with NeutrAvidin™ and used to screen sera from foals confirmed to have R.

Salmonella in the tropical household environment--Everyday, everywhere.

Objectives: To determine the prevalence of Salmonella in the environment of case and control houses, and compare serovars isolated from cases and their houses.

Methods: From 2005 to 2008, we tested samples from houses of 0-4 year old cases and community controls in Darwin and Palmerston for Salmonella. Case isolates were compared with environmental isolates.

The isolation and characterization of Campylobacter jejuni bacteriophages from free range and indoor poultry.

Six hundred and sixty one samples - primarily fresh chicken faeces - were processed to isolate wild type Campylobacter jejuni bacteriophages, via overlay agar methods using C. jejuni NCTC 12662. The aims of this study were to isolate and purify bacteriophages and then test for their ability to lyse field strains of C.

A new methodology for differentiation and typing of closely related Salmonella enterica serovar Heidelberg isolates.

This study describe the use of a combination of two recently proposed typing approaches, multiple amplification of prophage locus typing (MAPLT) and multiple-locus variable-number tandem-repeat analysis (MLVA) for subdividing within Salmonella enterica serovar Heidelberg (S. Heidelberg). The combined typing method was compared with pulsed-field gel electrophoresis (PFGE) by Simpson's index of diversity (DI).

Bacteriophage-mediated transduction of antibiotic resistance in enterococci.

Aims: Temperate bacteriophages are bacterial viruses that transfer genetic information between bacteria. This phenomenon is known as transduction, and it is important in acquisition of bacterial virulence genes and antimicrobial resistance determinants. The aim of this study was to demonstrate the role of bacteriophages in gene transfer (antibiotic resistance) in enterococci.

MLVA and phage typing as complementary tools in the epidemiological investigation of Salmonella enterica serovar Typhimurium clusters.

In South Australia serotyping and phage typing are employed for routine Salmonella surveillance. Molecular techniques such as Multiple-locus variable number tandem repeat analysis (MLVA) are increasingly utilized to aid outbreak investigations. During 2007 three Salmonella enterica serovar Typhimurium outbreaks involving phage types DT9, DT29, and DT44 were investigated.

Antimicrobial and heavy metal resistance in commensal enterococci isolated from pigs.

Antibiotic resistance in animal isolates of enterococci is of public health concern because of the risk of transfer of antibiotic resistance isolates or resistance determinants to consumers via the food chain. In this study, phenotypic and genotypic resistance in 192 pig isolates of enterococci to ampicillin, avilamycin, avoparcin, bacitracin, flavophospholipol, gentamicin, narasin, tetracycline, tiamulin, tylosin, vancomycin, virginiamycin, copper and zinc were investigated by susceptibility test and molecular methods. Resistance rates varied between the species but all isolates were susceptible to ampicillin, avilamycin, avoparcin, gentamicin and narasin but resistant to tetracycline and tylosin and intermediately resistant to copper.

Whole-genome sequencing and gene mapping of a newly isolated lytic enterococcal bacteriophage EFRM31.

Bacteriophages contribute greatly to bacterial evolution. There has been limited investigation of enterococcal bacteriophages, and only two enterococcal bacteriophages have been sequenced completely. In this study, a novel enterococcal bacteriophage, EFRM31, was isolated from a piggery effluent sample and then characterized.

Novel Bacteriophages in Enterococcus spp.

Most of the bacteriophages (phages) currently reported in Enterococcus spp. belong to tailed families of bacteriophages Podoviridae, Siphoviridae, and Myoviridae. There is a little information on non-tailed bacteriophages isolated from enterococci.

Linear B-cell epitope mapping using enzyme-linked immunosorbent assay for libraries of overlapping synthetic peptides.

The aim of this chapter is to provide a strategy for mapping linear antibody epitopes of protein antigens in order to discover candidates for vaccines or diagnostic tests. A set of overlapping peptides was designed and synthesised based upon a known amino acid sequence of the target protein, virulence-associated protein A (VapA) of the bacterium Rhodococcus equi, an important pulmonary pathogen in foals.The peptides were biotinylated and used in an ELISA to screen immune sera from foals.

A comparison of two PCR-based typing methods with pulsed-field gel electrophoresis in Salmonella enterica serovar Enteritidis.

Two novel molecular typing methods, multiple-locus variable-number tandem repeats (VNTR) analysis (MLVA) and multiple amplification of phage loci typing (MAPLT), were compared with pulsed-field gel electrophoresis (PFGE) for the discrimination of 128 Salmonella enterica serovar Enteritidis (S. Enteritidis) isolates. Selected epidemiologically unrelated isolates represented a cross-section of phage types routinely isolated in Australia and included 28 isolates that could not be assigned a phage type.

A comparison of three molecular typing methods for the discrimination of Salmonella enterica serovar Infantis.

Seventy-six epidemiologically unrelated Salmonella enterica serovar Infantis (S. Infantis) isolates were typed by pulsed-field gel electrophoresis (PFGE), multiple amplification of phage loci typing (MAPLT) and multiple-locus variable-number tandem-repeat analysis (MLVA). PFGE, using the restriction endonuclease XbaI, generated 23 different profiles for the 76 isolates (DI=0.

The use of MAPLT and MLVA analyses of phenotypically closely related isolates of Salmonella enterica serovar Typhimurium.

The resolving power of multiple-locus variable-number tandem repeat analysis (MLVA) was undertaken on 78 phenotypically closely related isolates of Salmonella enterica serovar Typhimurium that were previously analysed with multiple amplification of phage locus typing (MAPLT) and pulsed-field gel electrophoresis (PFGE). Isolates were tested with 10 primer sets targeting tandem repeat loci in the S. Typhimurium genome.

Chimeric vapA/groEL2 DNA vaccines enhance clearance of Rhodococcus equi in aerosol challenged C3H/He mice.

Rhodococcus equi remains a significant bacterial pathogen, causing severe pyogranulomatous pneumonia in foals aged 1-3 months. There is no effective vaccine currently available for the prevention of R. equi pneumonia.

A study of the transfer of tetracycline resistance genes between Escherichia coli in the intestinal tract of a mouse and a chicken model.

Experiments to demonstrate the transfer of genes within a natural environment are technically difficult because of the unknown numbers and strains of bacteria present, as well as difficulties designing adequate control experiments. The results of such studies should be viewed within the limits of the experimental design. Most experiments to date have been based on artificial models, which only give approximations of the real-life situation.

Clinical evaluation of a peptide-ELISA based upon N-terminal B-cell epitope of the VapA protein for diagnosis of Rhodococcus equi pneumonia in foals.

A total of 227 field samples from naturally exposed foals aged between 3 weeks and 6 months were used in an evaluation of a peptide-based enzyme-linked immunosorbent assay (ELISA) for diagnosis of Rhodococcus equi infection. A biotinylated peptide derived from the virulence-associated protein A (VapA) of R. equi, a horse pathogen, was synthesized and designated as PN11-14.

Recognition of a B-cell epitope of the VapA protein of Rhodococcus equi in newborn and experimentally infected foals.

The aim of this study was to evaluate the usefulness of the previously identified B-cell epitope TSLNLQKDEPNGRASDTAGQ of the VapA protein of Rhodococcus equi and its association with R. equi pneumonia. A modified peptide designated PN11-14 corresponding to the epitope was recognized by all sera from experimentally infected foals with virulent R.

Use of AFLP and PFGE to discriminate between Salmonella enterica serovar Typhimurium DT126 isolates from separate food-related outbreaks in Australia.

In 2001 Salmonella enterica serovar Typhimurium definitive phage-type (DT) 126 was isolated at higher frequency in Australia compared to other S. Typhimurium phage types and in comparison to previous years. Associated with this increase was the implication of this phage type in a number of food-related outbreaks.

Discrimination within phenotypically closely related definitive types of Salmonella enterica serovar typhimurium by the multiple amplification of phage locus typing technique.

Multilocus sequence typing (MLST) is a relatively new high-resolution typing system employed for epidemiological studies of bacteria, including Salmonella. Discrimination based on MLST of housekeeping genes may be problematical, due to the high identity of gene sequences of closely related Salmonella species. The presence of genomic sequences derived from stable temperate phages in Salmonella offers an alternative for MLST of Salmonella.

Immune response to vaccines based upon the VapA protein of the horse pathogen, Rhodococcus equi, in a murine model.

Rhodococcus equi is a significant pathogen in foals predominantly causing a pyogranulomatous bronchopneumonia. Many vaccine candidates have been tested for the prevention of R. equi disease in foals.

Antimicrobial resistance in Campylobacter spp., Escherichia coli and enterococci associated with pigs in Australia.

The major influences on the amplification and spread of antibiotic-resistant bacteria are the therapeutic use of antibiotics in human medicine and their use in livestock for therapy, prophylaxis and growth promotion. The use of veterinary antibiotics has many benefits to the livestock industries ensuring animal health and welfare, but use at subtherapeutic levels also exerts great selective pressure on emergence of resistant bacteria. The possible effect on human health is a problem of current debate.

ST64B is a defective bacteriophage in Salmonella enterica serovar Typhimurium DT64 that encodes a functional immunity region capable of mediating phage-type conversion.

The Salmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium) defective bacteriophage ST64B has a putative immunity (immC) region consisting of cI, cro and cII-like genes.

The immunogenicity of Rhodococcus equi GroEL2-based vaccines in a murine model.

Rhodococcus equi is a significant intracellular bacterial pathogen in foals. However, at present there is no commercially available vaccine for the prevention of R. equi-induced disease in these animals.

Fresh garlic: a possible vehicle for Salmonella Virchow.

A sustained increase in Salmonella enterica serovar Virchow notifications in South Eastern Australia between September 1997 and May 1998 instigated a case-control study and environmental investigations. Cases were defined as having locally acquired culture-confirmed S. Virchow phage-type 8 infection and diarrhoeal disease.

A fluorescent amplified fragment length polymorphism study of Salmonella enterica serovar Sofia, the major Salmonella serovar isolated from chickens in Australia.

Fluorescent amplified fragment length polymorphism (FAFLP) analysis was performed on 68 isolates of Salmonella enterica subsp. salamae serovar Sofia (S. Sofia).