Effect of temperature on viability of Campylobacter jejuni and Campylobacter coli on raw chicken or pork skin.

To determine growth and survival of Campylobacter jejuni and Campylobacter coli on chicken and pork, Campylobacter spp. (10(4) CFU/cm2) were inoculated on pieces of raw, irradiated chicken or pork skin and exposed to temperatures ranging from -20 to 42 degrees C under either microaerobic or aerobic conditions. Viable counts over 48 h declined 2 to 3 log CFU/cm2 at -20 degrees C and 1 to 2 log CFU/cm2 at 25 degrees C regardless of skin type, species of Campylobacter, or level of oxygen. At 4 degrees C, there was no significant change in the number of Campylobacter over 48 h. At both 37 and 42 degrees C, the number of viable Campylobacter increased significantly (2 to 3 log CFU/cm2, P < 0.0001) under microaerobic conditions but decreased 0.5 to 1.5 log CFU/cm2 in air. Preincubation of skins for 24 h at 42 degrees C under microaerobic conditions to establish Campylobacter on the surface prior to lowering the temperature to -20, 4, or 25 degrees C and incubating in air resulted in a decline in viability for the first 4 h (0.5 to 1 log CFU/cm2). However, after this initial drop in viability, no additional effect on viability was observed compared with incubation at -20, 4, or 25 degrees C in air without microaerobic preincubation at 42 degrees C. Preincubation of inoculated skins at -20, 4, or 25 degrees C in air for 24 h followed by a shift in temperature to 42 degrees C for 4, 8, 24, or 48 h and a shift to microaerobic conditions resulted in an overall decline in viability on raw pork skin but not on raw chicken skin. In contrast, preincubation of inoculated skins at -20, 4, or 25 degrees C for 24 h in air followed by a shift in temperature to 37 degrees C and microaerobic conditions did not result in a decrease in viable counts for either chicken or pork skins. Overall, viability of C. coli and C. jejuni on chicken and pork skins was similar. Therefore, a lower incidence of Campylobacter spp. in pork than in poultry postslaughter, despite a similar prevalence in live animals, is not due to differences in viability of C. coli versus C. jejuni on raw chicken or pork skin.

Complete nucleotide sequences of 84.5- and 3.2-kb plasmids in the multi-antibiotic resistant Salmonella enterica serovar Typhimurium U302 strain G8430.

The multi-antibiotic resistant (MR) Salmonella enterica serovar Typhimurium phage type U302 strain G8430 exhibits the penta-resistant ACSSuT-phenotype (ampicillin, chloramphenicol, streptomycin, sulfonamides and tetracycline), and is also resistant to carbenicillin, erythromycin, kanamycin, and gentamicin. Two plasmids, 3.2- and 84.5-kb in size, carrying antibiotic resistance genes were isolated from this strain, and the nucleotide sequences were determined and analyzed. The 3.2-kb plasmid, pU302S, belongs to the ColE1 family and carries the aph(3')-I gene (Kan(R)). The 84.5-kb plasmid, pU302L, is an F-like plasmid and contains 14 complete IS elements and multiple resistance genes including aac3, aph(3')-I, sulII, tetA/R, strA/B, bla(TEM-1), mph, and the mer operon. Sequence analyses of pU302L revealed extensive homology to various plasmids or transposons, including F, R100, pHCM1, pO157, and pCTX-M3 plasmids and TnSF1 transposon, in regions involved in plasmid replication/maintenance functions and/or in antibiotic resistance gene clusters. Though similar to the conjugative plasmids F and R100 in the plasmid replication regions, pU302L does not contain oriT and the tra genes necessary for conjugal transfer. This mosaic pattern of sequence similarities suggests that pU302L acquired the resistance genes from a variety of enteric bacteria and underscores the importance of a further understanding of horizontal gene transfer among the enteric bacteria.

Prevalence and characterization of shiga toxin-producing Escherichia coli in swine feces recovered in the National Animal Health Monitoring System's Swine 2000 study.

A study was conducted to determine the prevalence of Shiga toxin-producing Escherichia coli (STEC) in swine feces in the United States as part of the National Animal Health Monitoring System's Swine 2000 study. Fecal samples collected from swine operations from 13 of the top 17 swine-producing states were tested for the presence of STEC. After enrichment of swine fecal samples in tryptic soy broth, the samples were tested for the presence of stx1 and stx2 by use of the TaqMan E. coli STX1 and STX2 PCR assays. Enrichments of samples positive for stx1 and/or stx2 were plated, and colony hybridization was performed using digoxigenin-labeled probes complementary to the stx1 and stx2 genes. Positive colonies were picked and confirmed by PCR for the presence of the stx1, stx2, or stx2e genes, and the isolates were serotyped. Out of 687 fecal samples tested using the TaqMan assays, 70% (484 of 687) were positive for Shiga toxin genes, and 54% (370 of 687), 64% (436 of 687), and 38% (261 of 687) were positive for stx1, stx2, and both toxin genes, respectively. Out of 219 isolates that were characterized, 29 (13%) produced stx1, 14 (6%) produced stx2, and 176 (80%) produced stx2e. Twenty-three fecal samples contained at least two STEC strains that had different serotypes but that had the same toxin genes or included a strain that possessed stx1 in addition to a strain that possessed stx2 or stx2e. The STEC isolates belonged to various serogroups, including O2, O5, O7, O8, O9, OX10, O11, O15, OX18, O20, O57, O65, O68, O69, O78, O91, O96, O100, O101, O120, O121, O152, O159, O160, O163, and O untypeable. It is noteworthy that no isolates of serogroup O157 were recovered. Results of this study indicate that swine in the United States harbor STEC that can potentially cause human illness.

Quorum sensing and production of autoinducer-2 in Campylobacter spp., Escherichia coli O157:H7, and Salmonella enterica serovar Typhimurium in foods.

Autoinducer molecules are utilized by gram-negative and gram-positive bacteria to regulate density-dependent gene expression by a mechanism known as quorum sensing. PCR and DNA sequencing results showed that Campylobacter jejuni and Campylobacter coli possessed luxS, which is responsible for autoinducer-2 (AI-2) production. Using a Vibrio harveyi luminescence assay, the production of AI-2 was observed in milk, chicken broth, and brucella broth by C. coli, C. jejuni, Salmonella enterica serovar Typhimurium, and Escherichia coli O157:H7 under different conditions.

Phosphoprotein with phosphoglycerate mutase activity from the archaeon Sulfolobus solfataricus.

When soluble extracts of the extreme acidothermophilic archaeon Sulfolobus solfataricus were incubated with [gamma-(32)P]ATP, several proteins were radiolabeled. One of the more prominent of these, which migrated with a mass of approximately 46 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), was purified by column chromatography and SDS-PAGE and subjected to amino acid sequence analysis via both the Edman technique and mass spectroscopy. The best match to the partial sequence obtained was the potential polypeptide product of open reading frame sso0417, whose DNA-derived amino acid sequence displayed many features reminiscent of the 2,3-diphosphoglycerate-independent phosphoglycerate (PGA) mutases [iPGMs]. Open reading frame sso0417 was therefore cloned, and its protein product was expressed in Escherichia coli. Assays of its catalytic capabilities revealed that the protein was a moderately effective PGA mutase that also exhibited low levels of phosphohydrolase activity. PGA mutase activity was dependent upon the presence of divalent metal ions such as Co(2+) or Mn(2+). The recombinant protein underwent autophosphorylation when incubated with either [gamma-(32)P]ATP or [gamma-(32)P]GTP. The site of phosphorylation was identified as Ser(59), which corresponds to the catalytically essential serine residue in bacterial and eucaryal iPGMs. The phosphoenzyme intermediate behaved in a chemically and kinetically competent manner. Incubation of the (32)P-labeled phosphoenzyme with 3-PGA resulted in the disappearance of radioactive phosphate and the concomitant appearance of (32)P-labeled PGA at rates comparable to those measured in steady-state assays of PGA mutase activity.