Showing papers in "Fems Microbiology Letters in 1996"
TL;DR: Three enterotoxic components have been isolated from a strain of Bacillus cereus which was involved in a large food poisoning outbreak in Norway in 1995, and three proteins of 39, 45 and 105 kDa were found to be necessary for maximum cytotoxicity.
Abstract: Three enterotoxic components have been isolated from a strain of Bacillus cereus which was involved in a large food poisoning outbreak in Norway in 1995. The components were purified by chromatography on three different columns. Three proteins of 39, 45 and 105 kDa, respectively, were found to be necessary for maximum cytotoxicity. The amino acid N-terminal sequences of the 39 and 45 kDa proteins were determined. The 45 kDa component was the same protein as the main antigen detected in the Bacillus Diarrhoeal Enterotoxin Visual Immunoassay (Tecra). The 39 kDa protein showed some similarity to the l1 protein of haemolysin BL from B. cereus. Furthermore, the three toxic components were all recognised by a polyclonal antiserum reported to detect enterotoxin from B. cereus. The proteins were different from the B- and L2-components of haemolysin BL, previously suggested to be a primary virulence factor, and had no detectable haemolytic activity.
237 citations
TL;DR: Although most anaerobically expressed genes are subject to transcription control, none of them are totally switched off, so bacteria can respond equally rapidly whether times get better with the arrival of oxygen, or get worse when the nitrate is depleted.
Abstract: Anaerobic metabolism of the simplest, best understood enteric bacteria such as Escherichia coli is unexpectedly complex. Recent studies of the biochemistry and genetics of nitrate reduction via nitrite to ammonia by enteric bacteria have provided insights into the reasons for this complexity. An NADH-dependent nitrite reductase in the cytoplasm works in partnership with the respiratory nitrate reductase on the cytoplasmic side of the membrane when nitrate is abundant. There is also an electrogenic, formate-dependent nitrite reductase ready to work in partnership with a periplasmic nitrate reductase when nitrite is available but nitrate is scarce. A third E. coli nitrate reductase, NarZYWV, and the poorly expressed formate dehydrogenase O possibly facilitate rapid adaptation to oxygen starvation pending the synthesis of the major respiratory formate-nitrate oxidoreductase. Although most anaerobically expressed genes are subject to transcription control, none of them are totally switched off. This enables the bacteria to be ready for a change in fortune: when growing anaerobically with nitrate, they can respond equally rapidly whether times get better with the arrival of oxygen, or get worse when the nitrate is depleted. Far from being redundant, the complexity is essential for survival in a changing environment.
206 citations
TL;DR: A multiplex PCR assay was developed for characterization of B. anthracis isolates, and simultaneous confirmation of the species identity independent of plasmid content, and distinguished "anthrax-like' strains from other B. cereus group bacteria.
Abstract: Bacillus anthracis can be identified on the basis of the detection of virulence factor genes located on two plasmids, pXO1 and pXO2. Thus isolates lacking both pXO1 and pXO2 are indistinguishable from closely related B. cereus group bacteria. We developed a multiplex PCR assay for characterization of B. anthracis isolates, and simultaneous confirmation of the species identity independent of plasmid content. The assay amplifies lef, cya, pag (pXO1) and cap (pXO2) genes, and a B. anthracis specific chromosomal marker, giving an easy-to-read profile. This system unambiguously identified virulent (pXO1+/2+) and avirulent (pXO1+/2−, pXO1−/2+ and pXO1−/2−) strains of B. anthracis and distinguished ‘anthrax-like’ strains from other B. cereus group bacteria.
194 citations
TL;DR: It appears that CcpA acts globally to direct carbon flow in B. subtilis.
Abstract: The CcpA protein has been identified as a key regulator of carbon metabolism in Bacillus subtilis. CcpA is a DNA binding protein in the LacI/GalR transcriptional repressor family, and genes which respond to CcpA contain icommon cis-acting target sequences (Ccp boxes). A number of pathways involved in carbon source utilization are repressed by CcpA, while at least one gene which is involved in excretion of excess carbon is activated by CcpA. Genes repressed by CcpA generally contain Ccp boxes within or downstream of the promoter, while ackA, which is activated by CcpA, contains Ccp boxes upstream of the promoter. It therefore appears that CcpA acts globally to direct carbon flow in B. subtilis.
193 citations
TL;DR: The B. cepacia genome is comprised of multiple chromosomes and is rich in insertion sequences, which may have played a key role in the evolution of novel degradative functions and the unusual adaptability of this bacterium.
Abstract: Burkholderia cepacia has attracted attention because of its extraordinary degradative abilities and its potential as a pathogen for plants and for humans. This bacterium was formerly considered to belong to the genus Pseudomonas in the γ-subclass of the Proteobacteria, but recently has been assigned to the β-subclass based on rrn gene sequence analyses and other key phenotypic characteristics. The B. cepacia genome is comprised of multiple chromosomes and is rich in insertion sequences. These two features may have played a key role in the evolution of novel degradative functions and the unusual adaptability of this bacterium.
189 citations
TL;DR: It is concluded, based on polar lipid composition, that square, gas vacuole-containing Archaea of the type first described by Walsby are unrelated to the genera Halobacterium and Haloarcula, and probably belong to a new genus.
Abstract: Square, gas vacuole-containing Archaea of the type first described by Walsby were found to dominate in a saltern crystallizer pond in Eilat, Israel. To obtain information on the taxonomic position of these yet uncultured bacteria, we analyzed the polar lipids present in the microbial community in the saltern brine. In addition to phosphatidylglycerol, phosphatidylglycerophosphate and phosphatidylglycerosulfate we found one glycolipid, chromatographically identical with the sulfated diglycosyl diether lipid found as the major glycolipid in Haloferax species. As the square bacteria contributed at least 85% of the total membrane surface in the biota of the sample examined, we concluded, based on polar lipid composition, that these organisms are unrelated to the genera Halobacterium and Haloarcula, and probably belong to a new genus.
187 citations
TL;DR: Polyvinylpolypyrrolidone spin columns were used to rapidly purify crude soil DNA extracts from humic materials for polymerase chain reaction (PCR) analysis, and could be applied to the amplification of biodegradative genes from indigenous microbial populations from a wide variety of soil types.
Abstract: Polyvinylpolypyrrolidone spin columns were used to rapidly purify crude soil DNA extracts from humic materials for polymerase chain reaction (PCR) analysis. The PCR detection limit for the tfdC gene, encoding chlorocatechol dioxygenase from the 2,4-dichlorophenoxyacetic acid degradation pathway, was 101−102 cells/g soil in inoculated soils. The procedure could be applied to the amplification of biodegradative genes from indigenous microbial populations from a wide variety of soil types, and the entire analysis could be performed within 8 h.
187 citations
TL;DR: This was the first paper in which diffuse reflectance absorbance FT-IR spectroscopy was used with a supervised learning method in the form of artificial neural networks, and showed that this combination could succeed in discriminating a series of closely related, clinically relevant, Gram-positive bacterial strains.
Abstract: Diffuse reflectance-absorbance Fourier transform infrared spectroscopy (FT-IR) was used to analyse 19 hospital isolates which had been identified by conventional means to one Enterococcus faecalis, E faecium, Streptococcus bovis, S mitis, S pneumoniae, or S pyogenes Principal components analysis of the FT-IR spectra showed that this 'unsupervised' learning method failed to form six separable clusters (one of each species) and thus could not be used to identify these bacteria base on their FT-IR spectra By contrast, artificial neural networks (ANNs) could be trained by 'supervised' learning (using the back-propagation algorithm) with the principal components scores of derivatised spectra to recognise the strains from their FT-IR spectra These results demonstrate that the combination of FT-IR and ANNs provides a rapid, novel and accurate bacterial identification technique
165 citations
TL;DR: Carbon flux and its regulation, and branches into anabolic pathways are discussed together with recent approaches to broaden the substrate range of the bacterium.
Abstract: Sucrose, glucose and fructose are degraded in the Gram-negative bacterium Zymomonas mobilis via an anaerobic version of the Entner-Doudoroff pathway, to an equimolar mixture of ethanol and carbon dioxide. Sucrose is split extracellularly into glucose and fructose (or levan). The two sugars are transported into the cell via facilitated diffusion (uniport). A periplasmic enzyme, glucose-fructose oxidoreductase, provides the novel compatible solute, sorbitol, to counteract detrimental osmotic stress. Carbon flux and its regulation, and branches into anabolic pathways are discussed together with recent approaches to broaden the substrate range of the bacterium.
164 citations
TL;DR: Stress tolerance and cross-protection in Enterococcus faecalis ATCC19433 were examined after exposure to bile salts, acid or heat shock and whole-cell protein extract analysis revealed that each treatment induced a battery of stress proteins.
Abstract: Stress tolerance and cross-protection in Enterococcus faecalis ATCC19433 were examined after exposure to bile salts, acid or heat shock. Bile salts and heat adapted cells demonstrated induced homologous tolerance and cross-resistance. No cross-protection of heat adapted cells against acid stress is observed and pretreatment with bile salts even sensitized the cells to this challenge. Whole-cell protein extract analysis revealed that each treatment induced a battery of stress proteins. Some of these polypeptides are induced by more than one treatment. The greatest overlap is observed between bile salts and heat treatments. Eighteen stress proteins, including DnaK and GroEL, are common between these stresses.
154 citations
TL;DR: It is suggested that surfaces incorporating high densities of oligo(ethylene glycol) are good candidates for surfaces that interact minimally with bacteria.
Abstract: Bacterial cell attachment to the surfaces of self-assembled monolayers formed by the adsorption of omega-substituted alkanethiols on transparent gold films has been studied under defined bacterial culture and flow conditions. Phase contrast microscopy was used to quantify the attachment of two organisms, one of medical (Staphylococcus epidermidis) and one of marine (Deleya marina) importance. Self-assembled monolayers terminated with hexa(ethylene glycol), methyl, carboxylic acid and fluorocarbon groups were investigated. Over the range of experimental conditions, self-assembled monolayers formed from HS(CH2)11(OCH2CH2)6OH were found to be uniformly resistant to bacterial attachment, with a 99.7% reduction of attachment for both organisms when compared to the most fouled surface for each organism. On other surfaces, S. epidermidis and D. marina were shown to exhibit very different attachment responses to the wettability of the substratum. While the attachment of S. epidermidis correlated positively with surface hydrophilicity, D. marina showed a preference for hydrophobic surfaces. This study suggests that surfaces incorporating high densities of oligo(ethylene glycol) are good candidates for surfaces that interact minimally with bacteria.
TL;DR: A detailed analysis of groE and dnaK operons indicates that the organization of the groE operons is highly conserved, which means that genes have been added and deleted from it, and the gene order within the operon is variable.
Abstract: groEL and dnaK are the most highly conserved protein-coding genes known. Most groEL operons and several dnaK and dnaJ operons contain a highly conserved inverted repeat (IR) sequence in their regulatory region. So far, this IR has been found only as part of the groE, dnaK and dnaJ operons and genes. In most cases, the IR is part of the operon transcript, and is involved in the regulation of expression at both the DNA and the mRNA levels. A detailed analysis of groE and dnaK operons indicates that the organization of the groE operons is highly conserved. They contain only the groES and groEL genes and always in the same order. In contrast, the organization of the dnaK operons has changed during evolution: genes have been added and deleted from it, and the gene order within the operon is variable.
TL;DR: Results suggest that mutations in the rpoB gene are, mostly, but not necessarily, associated with rifampicin resistance of M. tuberculosis, and the sites of mutations on the rPOB gene will affect the level of resistance to rifampsicin.
Abstract: Using 39 clinical isolates of Mycobacterium strains with a broad range of susceptibility to rifampicin, we examined the relationship between the degree of resistance to rifampicin and mutational sites of the rpoB gene. All rifampicin-resistant strains had missense mutations. Twenty strains (95%) had a mutation in the cluster I region, which has also been reported in Escherichia coli [Jin and Gross (1988) J. Mol. Biol. 202, 45-58], and the remaining one strain had a mutation at codon 381 [Ala-->Val] in the N-terminal region, which has not been reported in E. coli. Among 18 rifampicin-susceptible strains, two had a mutation in the cluster I region and the other three strains had a mutation in the cluster III region. The mutations at codons 513 (5%), 526 (33%) or 531 (43%) in the cluster I region led to high level resistance to rifampicin (50 micrograms ml-1 < or = MIC). The mutations at the other sites, in the cluster III region (codons 679 or 687) and even in the cluster I region (codon 514, 521, or 533), showed low level (MIC = 12.5 micrograms ml-1) or no (MIC < 0.39 microgram ml-1) resistance to rifampicin. These results suggest that mutations in the rpoB gene are, mostly, but not necessarily, associated with rifampicin resistance of M. tuberculosis, and the sites of mutations on the rpoB gene will affect the level of resistance to rifampicin.
TL;DR: A set of vectors containing a mutated gfp gene was constructed for use with Gram-negative bacteria other than Escherichia coli, and two plasmids containing gfp expressed from a lac and an npt2 promoter were constructed.
Abstract: A set of vectors containing a mutated gfp gene was constructed for use with Gram-negative bacteria other than Escherichia coli. These constructs were: pTn3gfp for making random promoter probe gfp insertions into cloned DNA in E. coli for subsequent introduction into host strains; pUTmini-Tn5gfp for making random promoter probe gfp insertions directly into host strains; p519gfp and p519nfp, broad host range mob+ plasmids containing gfp expressed from a lac and an npt2 promoter, respectively.
TL;DR: This work has analysed the effect of oxidants on the activities of a number of enzymes with known antioxidant activity and found that C. albicans showed a greater level of resistance to oxidants, both H2O2 and redox-cycling agents, compared to that observed with S. cerevisiae.
Abstract: Treatment of Candida albicans with low concentrations of either hydrogen peroxide or menadione (a Superoxide generating agent) induces an adaptive response which protects cells from the lethal effects of a subsequent challenge with higher concentrations of these oxidants. Pre-treatment with either menadione or hydrogen peroxide is protective against cell killing by either oxidant. This suggests that the pathogenic yeast C. albicans (unlike the budding yeast Saccharomyces cerevisiae which has separate responses) possesses an adaptive response that responds to both these oxidants. In addition, we found that C. albicans showed a greater level of resistance to oxidants, both H2O2 and redox-cycling agents, compared to that observed with S. cerevisiae. In an attempt to characterise the oxidative stress response in more detail we have analysed the effect of oxidants on the activities of a number of enzymes with known antioxidant activity.
TL;DR: Analysis of phenazine gene ( phz ) expression demonstrates that phenazine biosynthesis in P. aureofaciens is regulated analogously to other bacterial genes involved in host-microbe interactions, and proposes a model to explain these interactions between the plant, the pathogen and the bacterium.
Abstract: Pseudomonas aureofaciens are soil-borne root-colonizing bacteria that produce phenazine antibiotics. Populations of P. aureofaciens strain 30–84 increase over time on wheat roots in response to the presence of a specific fungal pathogen. Phenazine production is the primary mechanism responsible for the competitive fitness of P. aureofaciens in the rhizosphere. Analysis of phenazine gene ( phz ) expression demonstrates that phenazine biosynthesis in P. aureofaciens is regulated analogously to other bacterial genes involved in host-microbe interactions. Two genes, phz R and phz I , the products of which belong to the LuxR/LuxI family of cell density-dependent regulatory proteins, are required for phz expression. PhzR encodes a transcriptional activator that induces phz expression in response to the accumulation of a diffusible signal produced by PhzI. The requirement for phzI can be bypassed by exogenous diffusible signals produced by several other rhizosphere bacteria. We discuss phz regulation in the context of the ecology of P. aureofaciens on the plant root. We integrate both molecular and field observations to propose a model to explain these interactions between the plant, the pathogen and the bacterium. In addition, we discuss signaling among different bacterial populations in the rhizosphere and hypothesize that this signaling may influence phz expression and thus biological control by P. aureofaciens .
TL;DR: Electroporation at 37 degrees C has been chosen as a standard condition for slow-growing species as it usually resulted in a transformation efficiency several orders of magnitude higher than that obtained at 0 degree C.
Abstract: The effect of electroporation temperature, biochemical pretreatment of cells and stage of culture on electroporation efficiency for slow-growing mycobacteria were investigated. The efficiency of transformation into Mycobacterium tuberculosis, Mycobacterium bovis and Mycobacterium intracellulare increased markedly with temperature. In contrast, the efficiency of transformation into Mycobacterium smegmatis, a fast-growing species, was higher at 0°C and decreased with temperature. While stage of culture had little effect, a further increase in efficiency of 2–4-fold was obtained following glycine or ethionamide pretreatment. Electroporation at 37°C has been chosen as a standard condition for slow-growing species as it usually resulted in a transformation efficiency several orders of magnitude higher than that obtained at 0°C.
TL;DR: Evidence is provided that sigB mutants have an obvious phenotype compared to the wild type and indicates that delta B-dependent general stress proteins may function in providing starving cells with resistance against oxidative stress.
Abstract: Two catalases of B. subtilis have been studied which are subject to two different regulatory mechanisms. Whereas KatA belongs to the group of proteins specifically induced by oxidative stress, KatE is a general σB-dependent stress protein, not induced by oxidative stress. There are two mechanisms of oxidative stress resistance, the adaptive resistance induced by low H2O2 concentrations and an unspecific resistance acquired in glucose-starved cells. Mutants lacking KatA are defective in the adaptive resistance and both exponentially growing and glucose-starved cells are 100-fold more sensitive against lethal concentrations of H2O2. Under both conditions, however, a katE mutant was just as resistant as the wild type. Therefore, the role of KatE in oxidative stress tolerance remains obscure. sigB mutants which are non longer able to induce σB-dependent general stress proteins in glucose-starved cells are characterized by a strong impairment in the unspecific oxidative stress resistance but not in the H2O2-induced oxidative stress resistance. This is the first evidence that sigB mutants have an obvious phenotype compared to the wild type and indicates that σB-dependent general stress proteins may function in providing starving cells with resistance against oxidative stress.
TL;DR: The purpose of this review is to survey molecular mechanisms of multidrug resistance in enteric and other Gram-negative bacteria, and to speculate on the origins and natural physiological functions of the genes involved.
Abstract: In Gram-negative bacteria, multidrug resistance is a term that is used to describe mechanisms of resistance by chromosomal genes that are activated by induction or mutation caused by the stress of exposure to antibiotics in natural and clinical environments. Unlike plasmid-borne resistance genes, there is no alteration or degradation of drugs or need for genetic transfer. Exposure to a single drug leads to cross-resistance to many other structurally and functionally unrelated drugs. The only mechanism identified for multidrug resistance in bacteria is drug efflux by membrane transporters, even though many of these transporters remain to be identified. The enteric bacteria exhibit mostly complex multidrug resistance systems which are often regulated by operons or regulons. The purpose of this review is to survey molecular mechanisms of multidrug resistance in enteric and other Gram-negative bacteria, and to speculate on the origins and natural physiological functions of the genes involved.
TL;DR: In vitro excystation, vital dyes, DAPI and propidium iodide, and infectivity in neonatal CD-1 mice were used to assess the viability of Cryptosporidium parvum oocysts after chemical disinfection.
Abstract: In vitro excystation, vital dyes (4′, 6-diamidino-2-phenylindole (DAPI) and propidium iodide (PI)), and infeictivity in neonatal CD-1 mice were used to assess the viability of Cryptosporidium parvum oocysts after chemical disinfection. In vitro excystation and DAPI/PI staining provided similar estimates of viability in bench-scale experiments, but both of these methods significantly overestimated the viability when compared with infectivity (Pr ≤ 0.01). Infectivity was the most reliable measure of the viability of C. parvum oocysts following chemical disinfection.
TL;DR: In this paper, the role of glutathione in the protection of the yeast Saccharomyces cerevisiae against oxidative stress was investigated and it was shown that it is an important antioxidant molecule in yeast, with gamma-glutamylcysteine synthetase (gsh1) mutants, deficient in glutathion synthesis, being hypersensitive to H2O2 and superoxide anions.
Abstract: The tripeptide gamma-L-glutamyl-L-cystinylglycine (glutathione) is one of the major antioxidant molecules of cells and is thought to play a vital role in buffering the cell against reactive oxygen species and toxic electrophiles. We wished to determine the role of glutathione in the protection of the yeast Saccharomyces cerevisiae against oxidative stress. This study shows that glutathione is an important antioxidant molecule in yeast, with gamma-glutamylcysteine synthetase (gsh1) mutants, deficient in glutathione synthesis, being hypersensitive to H2O2 and superoxide anions in both exponential- and stationary-phase cultures. Despite this, these mutants are still able to induce adaptive stress responses to oxidants.
TL;DR: The PCR-RFLP assay described here is a sensitive tool to distinguish C. parvum isolates and Typing of 23 isolates showed that 10/10 calf isolates had the same profile, while 2 patterns were observed among human isolates: 7/13 displayed the calf profile, and 6/13 presented another pattern.
Abstract: In order to define transmission routes of cryptosporidiosis and develop markers that distinguish Cryptosporidium parvum isolates, we have identified 2 polymorphic restriction enzyme sites in a C. parvum repetitive DNA sequence. The target sequence was amplified by polymerase chain reaction from 100 to 500 oocysts and the amplified product was subjected to restriction enzyme digestion. Typing of 23 isolates showed that 10 / 10 calf isolates had the same profile. In contrast, 2 patterns were observed among human isolates: 7 / 13 displayed the calf profile, and 6 / 13 presented another pattern. The PCR-RFLP assay described here is a sensitive tool to distinguish C. parvum isolates.
TL;DR: It is shown that the collagen binding protein from Lactobacillus reuteri NCIB 11951 has sequence similarities to the solute binding component of bacterial ABC transporters.
Abstract: The gene coding for a collagen binding protein from Lactobacillus reuteri NCIB 11951 was cloned and sequenced. A genomic lambda library was constructed and recombinant plaques were screened using antisera raised against purified collagen binding proteins from the same L. reuteri strain. The positive plaques were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis, which revealed the expression of a 29 kDa protein, which reacted with the antisera and bound 125I-labelled type I collagen. The sequence of the corresponding gene, cnb showed that the collagen binding protein has sequence similarities to the solute binding component of bacterial ABC transporters.
TL;DR: It is indicated that both toxic and non-toxic strains of M. aeruginosa possess genes coding for peptide synthetases and that hepatotoxin-producing andNon-Toxic strains differ in their content of genes for specific peptide Synthetases.
Abstract: Toxic strains of Microcystis aeruginosa produce cyclic heptatoxins (microcystins) that are believed to be synthesized non-ribosomally by peptide synthetases. We analysed toxin-producing and non-toxic strains of M. aeruginosa with respect to the presence of DNA sequences potentially encoding peptide synthetases. Hybridizations of genomic DNA of various M. aeruginosa strains with PCR-amplificated fragments possessing homologies to adenylate-forming domains of peptide synthetase genes provided first evidence for the existence of corresponding genes in cyanobacteria. Furthermore we isolated and sequenced from genomic libraries overlapping fragments of M. aeruginosa DNA with a total length of 2982 bp showing significant homology to genes encoding peptide synthetases and hybridizing exclusively with DNA from toxic strains. Our results indicate that both toxic and non-toxic strains of M. aeruginosa possess genes coding for peptide synthetases and that hepatotoxin-producing and non-toxic strains differ in their content of genes for specific peptide synthetases.
TL;DR: Random amplified polymorphic DNA and restriction fragment end-labeling analysis of the isolates showed close genetic relatedness between pre- and post-treatment isolates, indicating that resistance was the result of selection of variants of the infecting strain rather then infection with an exogenous resistant strain.
Abstract: Seventy-three Helicobacter pylori-positive patients were treated with a combination of clarithromycin and ranitidine in order to eradicate the bacterium. Eradication was successful in 79.5%. In 15 patients eradication failed, and in 11 cases this was due to clarithromycin resistance. In one patient the infecting strain was resistant at the onset of treatment, while in the remaining 10 patients resistance developed during therapy. These isolates had also become resistant to various other antibiotics. Random amplified polymorphic DNA and restriction fragment end-labeling analysis of the isolates showed close genetic relatedness between pre- and post-treatment isolates, indicating that resistance was the result of selection of variants of the infecting strain rather then infection with an exogenous resistant strain. Nucleotide sequence comparisons revealed that all resistant isolates had a single base pair mutation in the 23S rRNA. Since this single point mutation results in co-resistance to various antibiotics at high frequencies, caution should be taken when using clarithromycin as a single antibiotic.
TL;DR: The local and transient sensitivity observed in vivo, and the in vitro adaptations suggest that several detergent-resistance mechanisms operate in S. typhimurium.
Abstract: Salmonella serotype typhimurium transpositional mutants altered in resistance to biliary salts and detergents were isolated previously. We have characterized further the LX1054 mutant strain, the most sensitive of them. The chromosomal DNA segment flanking transposon insertion was cloned and sequenced. The highest level of identity was found for the acrB (formerly acrE) gene of Escherichia coli, a gene encoding a drug efflux pump of the Acr family. LX1054 exhibited a reduced capacity to colonize the intestinal tract. After passages in mice, the mutant strain lost the sensitive phenotype. In vitro, a resumption of growth appeared after 17 h of culture in medium with cholate or other tested biological or chemical detergents. Then, the acquired resistant phenotype seemed stable. The data suggested a role of S. typhimurium acrB-like gene in resistance to biliary salts and detergents and in mice intestinal colonization. However, the local and transient sensitivity observed in vivo, and the in vitro adaptations suggest that several detergent-resistance mechanisms operate in S. typhimurium.
TL;DR: Analysis of the production kinetics for the autoinducer during the generation of conditioned media indicates that it differs from other more well characterized autoinducers, and it is proposed that the neurohumoral environment of the host may contribute to the production of bacterial growth factors.
Abstract: Escherichia coli O157:H7 were cultured in the presence or absence of norepinephrine to generate conditioned media. The presence of a growth-inducing factoKs) in the conditioned media was examined by measurement of the ability of conditioned media to support the growth of fresh cultures of E. coli O157:H7. Supplementation of fresh cultures with as little as 0.024% (v/v) norepinephrine conditioned medium resulted in increased growth as compared to controls, thereby indicating the presence of an autoinducer of growth. Analysis of the production kinetics for the autoinducer during the generation of conditioned media indicates that it differs from other more well characterized autoinducers. It is proposed that the neurohumoral environment of the host may contribute to the production of bacterial growth factors.
TL;DR: The bradyzoite and tachyzoite forms of Toxoplasma gondii, purified from infected animals, were analysed for their activities of phosphofructokinase, pyruvate kinase, lactate dehydrogenase, NAD(+)- and NADH-linked isocitrate dehydrogenases, and succinic dehydrogensase, suggesting that energy metabolism in both forms may centre around a high glycolytic flux linked to lactate production.
Abstract: The bradyzoite and tachyzoite forms of Toxoplasma gondii, purified from infected animals, were analysed for their activities of phosphofructokinase, pyruvate kinase, lactate dehydrogenase, NAD(+)- and NADH-linked isocitrate dehydrogenases, and succinic dehydrogenase. Both developmental stages contained high activities of phosphofructokinase (specific for pyrophosphate rather than ATP), pyruvate kinase and lactate dehydrogenase, suggesting that energy metabolism in both forms may centre around a high glycolytic flux linked to lactate production. The markedly higher activity of the latter two enzymes in bradyzoites suggests that lactate production is particularly important in this developmental form. NAD(+)-specific isocitrate dehydrogenase was not detectable in either stage of the parasite (and proved useful as a measure of the purity of the bradyzoite preparation), whereas both parasite forms contained low activities of NADP(+)-linked isocitrate dehydrogenase. The results are consistent with the bradyzoites lacking a functional TCA cycle and respiratory chain and are suggestive of a lack of susceptibility of this developmental stage to atovaquone.
Metz1
TL;DR: A rapid screening procedure was developed for detection of Escherichia coli mutants with altered adhesion abilities using polystyrene 96-well microtiter plates as attachment surfaces and 34 mutants displayed altered motility.
Abstract: A rapid screening procedure was developed for detection of Escherichia coli mutants with altered adhesion abilities using polystyrene 96-well microtiter plates as attachment surfaces. During this assay, bacterial strains grew and adhered simultaneously, and attached cells were measured after crystal violet staining. Starting with a total of 7000 W3110::Tn10 insertion mutants of E. coli K-12 W3110, 50 adhesion-deficient mutants were isolated which showed less than 40% attachment, and 22 mutants were found with an attachment of 40–75%. Motility assays were performed on these 72 mutants, and 34 displayed altered motility.
TL;DR: The major OMP genes display diversity among Brucella species, biovars and strains allowing their differentiation, and the polymorphic markers identified have brought new insights into the evolutionary development of the genus brucella, antigenic variability of brucellae, and future prospects in the field of vaccine development.
Abstract: The major outer membrane proteins (OMPs) of Brucella spp. were initially identified in the early 1980s by selective extraction techniques and classified according to their apparent molecular mass as 36–38 kDa OMPs or group 2 porin proteins and 31–34 kDa and 25–27 kDa OMPs which belong to the group 3 proteins. Variation in apparent molecular mass is essentially due to association with peptidoglycan subunits of different sizes. Two genes, omp2a and omp2b, which are closely linked in the Brucella genome, and which share a great degree of homology (>85%), encode the 36 kDa porin proteins, now named Omp2a and Omp2b proteins respectively. Two genes code for the group 3 OMPs and are names omp25 and omp31. The predicted amino acid sequences of omp25 and omp31 share 34% identity. Furthermore, all Brucella major OMPs share amino acid sequence homology with the major OMPs RopA or RopB of Rhizobium leguminosarum, which supports the close genetic relationship of brucellae with members of the α-2 subdivision of the class Proteobacteria. Another characteristic common to the major OMPs of R. leguminosarum and Brucella is that they are tightly, probably covalently, associated with the peptidoglycan. The major OMP genes display diversity among Brucella species, biovars and strains allowing their differentiation, and the polymorphic markers identified have brought new insights into the evolutionary development of the genus Brucella, antigenic variability of brucellae, and future prospects in the field of vaccine development.