An ultrastructural comparison of spores from various strains of Clostridium perfringens and correlations with heat resistance parameters.

It has been shown that Clostridium perfringens isolates associated with food poisoning carry a chromosomal cpe gene, whereas nonfood-borne human gastrointestinal disease isolates carry a plasmid cpe gene. In addition, the chromosomal cpe gene isolates exhibit greater heat resistance as compared with the plasmid cpe strains. Therefore, the current study conducted ultrastructural measurements of spores from several plasmid and chromosomal cpe-positive C. perfringens isolates. In support of the dehydration mechanism of spore heat resistance, the C. perfringens spore core average size was found to show a negative correlation with D-values for spores obtained at 100 degrees C. Dipicolinic acid (DPA) concentrations assayed for the spores did not correlate well with C. perfringens spore core averages nor with D(10)-values at 100 degrees C. Spore core thickness might be a distinguishing phenotypic characteristic used to identify heat resistance and survival potential of C. perfringens in improperly cooked foods.

Detection of heat injury in Listeria monocytogenes Scott A.

Methods of detecting live pathogens in foods that may be growth inhibited following heat treatment are essential to food safety. Among the techniques available, reverse transcription polymerase chain reaction (RT-PCR) amplification of messenger RNA from heat-injured Listeria monocytogenes Scott A is preferable to direct PCR in an attempt to avoid false positives from dead cells. The RT-PCR has a detection limit of 3 x 10(6) CFU/g, compared to 3 CFU/g for untreated controls, but may not be suitable for the identification of all viable cells. Physically apparent changes in cellular structures from heat injury in L. monocytogenes are expected to result. Ultrastructural analyses did depict notable heat damage as cytoplasmic clearing after 5 min at 60 degrees C. The heat-injured survivors can be readily distinguished from total viable cells using selective media. As a result, combinations of molecular and visual methods including selective media improve detectability of heat-injured, viable L. monocytogenes Scott A.

Heat treatment adaptations in Clostridium perfringens vegetative cells.

Vegetative cells of Clostridium perfringens enterotoxigenic strains NCTC 8679, NCTC 8238. and H6 were grown at 37 degrees C followed by a 60-min exposure to 28 degrees C or 46 degrees C. D10-values, as a measure of thermal resistance at 60 degrees C, were significantly lower for 28 degrees C exposures as compared with cultures given 37 and 46 degrees C exposures. Following refrigeration at 4 degrees C for 24 h, D10-values for the 37 and 46 degrees C samples could not be differentiated from 28 degrees C samples. Western immunoblot analyses of lysates from heat-adapted cells also detected the increased expression of proteins reacting with antiserum directed against the molecular chaperonins from Escherichia coli; GroEL, DnaJ, and the small acid soluble protein from Bacillus subtilis, SspC. Differential scanning calorimetry (DSC) identified thermal transitions corresponding to ribosomal protein denaturations at 72.1 +/- 0.5 degrees C. Any cellular heat adaptations in the DSC profiles were lost following refrigeration for several days to simulate minimally processed food storage conditions. Further analyses of high-speed pellets from crude cell extract fractions using two-dimensional gel electrophoresis detected the differential gene expression of at least four major proteins in heat-adapted vegetative cells of C. perfringens. N-terminal amino acid analyses identified two of the proteins as glyceraldehyde 3-phosphate dehydrogenase and rubrerythrin. Both appear to have roles in this anaerobe under stressful conditions.

Molecular approaches to probe differential NADH activation of phosphoribulokinase isozymes from Rhodobacter sphaeroides.

The cbbPI and cbbPII genes from Rhodobacter sphaeroides, encoding highly similar phosphoribulokinase (PRK) isozymes, PRK I and PRK II, respectively, exhibited differential allosteric activation by NADH. The two cbbP genes were cloned into expression vectors and homogeneous recombinant protein prepared. PRK II was found to be inherently less stable than PRK I; however, the addition of substrate ATP resulted in the complete protection of both isozymes to a 15-min incubation at 50 degrees C. The relative molecular masses for both octameric isozymes were determined to be approximately 230,000; however, the protective effect of ATP was in accordance with aggregation of monomers to a molecular mass of approximately 750,000. While PRK I exhibited a nearly absolute dependence upon NADH for activity, PRK II retained substantial activity in the absence of NADH. PRK chimeras were thus constructed to facilitate elucidation of the basis for the differential effect of NADH, with advantage taken of the relative sequence identity of about 90% between the two isozymes. Chimeras were constructed either by in vivo homologous recombination, using the sacB gene from Bacillus subtilis as a conditionally lethal marker, or by using convenient restriction sites to combine different parts of the two cbbP genes. The PRK chimeras generated contained either the amino-terminal domain of PRK II and the carboxy-terminal domain of PRK I or the opposite configuration. Subsequent analyses of the chimeras pointed to particular regions and residue(s) as likely being important for NADH activation.

Nonsurgical management of acute jejunal diverticulitis: a review.

BACKGROUND: Diverticular disease of the colon and its complications are well known and readily considered when patients present with the proper clinical scenario. Conversely, complications of small bowel diverticula are very uncommon entities and are not often thought of as a cause of bleeding, obstruction, or an acute abdomen. OBJECTIVE: To report two patients presenting with an acute abdomen caused by acute jejunal diverticulitis who were treated nonsurgically as opposed to surgically as the literature dictates. METHODS: Two patients presented with sudden onset of acute periumbilical pain that had increased progressively over 1-2 days before admission. An emergent CT scan performed in each patient with localized peritonitis revealed "collections" consistent with abscess cavities. One patient was treated with antibiotics alone and the other with a combination of antibiotics and percutaneous CT-guided aspiration. CT-guided needle aspiration was performed and the injection of contrast clearly revealed communication with a jejunal diverticulum. Both patients did well and were subsequently discharged without incident or surgical intervention. CONCLUSIONS: Acute jejunal diverticulitis must be considered in the differential diagnosis of an acute abdominal process and may be successfully treated nonsurgically despite the recommendations of previous reports.

The protein tyrosine phosphatase LAR has a major impact on insulin receptor dephosphorylation.

Transmembrane protein tyrosine phosphatases (PTPases) may act as regulators of the insulin receptor. Supporting this hypothesis, antisense suppression of the PTPase LAR in McA-RH7777 hepatoma cells increased insulin receptor signaling (Kulas et. al., J. Biol. Chem. (1996) 271, 748-754). The effects of decreased LAR expression may be mediated by decreased dephosphorylation of the insulin receptor. The rate of insulin receptor dephosphorylation was examined in situ, following elution of surface bound insulin at pH 4.0. In LAR antisense cells, dephosphorylation was prolonged by 2.6-fold with a t(1/2) of 87+/-11 sec compared to a t(1/2) of 34+/-6 sec in control cells. EGF receptor dephosphorylation was also prolonged in LAR antisense cells. These results are further evidence that LAR is a physiological regulator of the insulin receptor and is consistent with its direct interaction with the tyrosine phosphorylated insulin receptor.

Heteropolysaccharide Formation by Arthrobacter viscosus Grown on Xylose and Xylose Oligosaccharides.

Arthrobacter viscosus NRRL B-1973 produces its viscous extracellular polysaccharides (EPS) when grown on media containing xylose or enzymatic xylan hydrolysates. Crude EPS formation from xylose averaged 12 g/liter when initial culture pH was adjusted to 8.0 and total nitrogen was limited to 0.03%. Purified EPS from pentose and hexose substrates were analyzed for their monosaccharide, acetyl, and uronic acid components, intrinsic viscosities, and average molecular masses. Differences were apparent in degrees of acetylation, molecular masses, and intrinsic viscosities of the heteropolysaccharides produced on different carbon sources.