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Salmonella Gene rma (ramA) and Multiple-Drug-Resistant Salmonella enterica Serovar Typhimurium. Tahar van der Straaten, 2004.MarA and its homologue, RamA, have been implicated in multidrug resistance (MDR) . RamA overexpression in Salmonella enterica serovar Typhimurium and Escherichia coli conferred MDR independently of marA . Inactivation of ramA did not affect the antibiotic susceptibilities of wild-type S . enterica serovar Typhimurium or 15 unrelated clinical MDR isolates . Thus, ramA overexpression is not a common MDR mechanism in Salmonella . Growth of Polychlorinated-Biphenyl-Degrading Bacteria in the Presence of Biphenyl and Chlorobiphenyls Generates Oxidative Stress and Massive Accumulation of Inorganic Polyphosphate. Francisco P. Chávez, 2004.Inorganic polyphosphate (polyP) plays a significant role in increasing bacterial cell resistance to unfavorable environmental conditions and in regulating different biochemical processes . Using transmission electron microscopy of the polychlorinated biphenyl (PCB)-degrading bacterium Pseudomonas sp . strain B4 grown in defined medium with biphenyl as the sole carbon source, we observed large and abundant electron-dense granules at all stages of growth and following a shift from glucose to biphenyl or chlorobiphenyls . Using energy dispersive X-ray analysis and electron energy loss spectroscopy with an integrated energy-filtered transmission electron microscope, we demonstrated that these granules were mainly composed of phosphate . Using sensitive enzymatic methods to quantify cellular polyP, we confirmed that this polymer accumulates in PCB-degrading bacteria when they grow in the presence of biphenyl and chlorobiphenyls . Concomitant increases in the levels of the general stress protein GroEl and reactive oxygen species were also observed in chlorobiphenyl-grown cells, indicating that these bacteria adjust their physiology with a stress response when they are confronted with compounds that serve as carbon and energy sources and at the same time are chemical stressors . Membrane Topology of the Streptococcus pneumoniae FtsW Division Protein. Philippe Gérard, 2002.The topology of FtsW from Streptococcus pneumoniae, an essential membrane protein involved in bacterial cell division, was predicted by computational methods and probed by the alkaline phosphatase fusion and cysteine accessibility techniques . Consistent results were obtained for the seven N-terminal membrane-spanning segments . However, the results from alkaline phosphatase fusions did not confirm the hydropathy analysis of the C-terminal part of FtsW, whereas the accessibility of introduced cysteine residues was in agreement with the theoretical prediction . Based on the combined results, we propose the first topological model of FtsW, featuring 10 membrane-spanning segments, a large extracytoplasmic loop, and both N and C termini located in the cytoplasm . Targeted Disruption of the  -Amylase Gene in the Hyperthermophilic Archaeon Sulfolobus solfataricus.Penny Worthington, 2003.Sulfolobus solfataricus secretes an acid-resistant -amylase (amyA) during growth on starch as the sole carbon and energy source . Synthesis of this activity is subject to catabolite repression . To better understand
-amylase function and regulation, the structural gene was identified and disrupted and the resulting mutant was characterized . Internal
-amylase peptide sequences obtained by tandem mass spectroscopy were used to identify the amyA coding sequence . Anti--amylase antibodies raised against the purified protein immunoprecipitated secreted
-amylase activity and verified the enzymatic identity of the sequenced protein . A new gene replacement method was used to disrupt the amyA coding sequence by insertion of a modified allele of the S . solfataricus lacS gene . PCR and DNA sequence analysis were used to characterize the altered amyA locus in the recombinant strain . The amyA::lacS mutant lost the ability to grow on starch, glycogen, or pullulan as sole carbon and energy sources . During growth on a non-catabolite-repressing carbon source with added starch, the mutant produced no detectable secreted amylase activity as determined by enzyme assay, plate assay, or Western blot analysis . These results clarify the biological role of the
-amylase and provide additional methods for the directed genetic manipulation of the S . solfataricus genome .
Microbial Quality and Direct PCR Identification of Lactic Acid Bacteria and Nonpathogenic Staphylococci from Artisanal Low-Acid Sausages. T. Aymerich, 2003.Detection of six species of lactic acid bacteria and six species of gram-positive catalase-positive cocci from low-acid fermented sausages (fuets and chorizos) was assessed by species-specific PCR . Without enrichment, Lactobacillus sakei and Lactobacillus curvatus were detected in 11.8% of the samples, and Lactobacillus plantarum and Staphylococcus xylosus were detected in 17.6% . Enriched samples allowed the detection of L . sakei and S . xylosus in all of the samples (100%) and of Enterococcus faecium in 11.8% of the sausages . The percentages of L . curvatus, L . plantarum, Staphylococcus carnosus, and Staphylococcus epidermidis varied depending on the sausage type . L . curvatus was detected in 80% of fuets and in 57% of chorizos . L . plantarum was found in 50% of fuets and 100% of chorizos . S . epidermidis was detected in only 11.8% of fuets, and S . carnosus was detected in only 5.9% of chorizos . Lactococcus lactis, Staphylococcus warneri, and Staphylococcus simulans were not detected in any sausage type . From a microbiological point of view, 70.6% of the samples could be considered of high quality, as they had low counts of Enterobacteriaceae and did not contain any of the food-borne pathogens assayed .
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