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Parental Strain (parental + strain)
Selected AbstractsGenotoxicity of nitrosulfonic acids, nitrobenzoic acids, and nitrobenzylalcohols, pollutants commonly found in ground water near ammunition facilitiesENVIRONMENTAL AND MOLECULAR MUTAGENESIS, Issue 2 2006Tamara Grummt Abstract 2-Amino-4,6-dinitrobenzoic acid (2-A-4,6-DNBA), 4-amino-2,6-dinitrobenzoic acid (4-A-2,6-DNBA), 2,4,6-trinitrobenzoic acid (2,4,6-TNBA), 2-amino-4, 6-dinitrobenzylalcohol (2-A-4,6-DNBAlc), 4-amino-2,6-dinitrobenzylalcohol (4-A-2,6-DNBAlc), 2,4-dinitrotoluol-5-sulfonic acid (2,4-DNT-5-SA), 2,4-dinitrotoluol-3-sulfonic acid (2,4-DNT-3-SA), and 2, 4-dinitrobenzoic acid (2,4-DNBA) are derivatives of nitro-explosives that have been detected in groundwater close to munitions facilities. In the present study, the genotoxicity of these compounds was evaluated in Salmonella/microsome assays (in strains TA100 and TA98, with and without S9 and in TA98NR without S9), in chromosomal aberration (CA) tests with Chinese hamster fibroblasts (V79), and in micronucleus (MN) assays with human hepatoma (HepG2) cells. All compounds except the sulfonic acids were positive in the bacterial mutagenicity tests, with 2,4,6-TNBA producing the strongest response (8023 revertants/,mol in TA98 without S9 activation). 2-A-4,6-DNBA was a direct acting mutagen in TA98, but negative in TA100. The other positive compounds were ,1,3 orders of magnitude less mutagenic than 2,4,6-TNBA in TA98 and in TA100; relatively strong effects (,50,400 revertants/,mol) were produced by the benzylacohols in the two indicator strains. With the exception of 2,4-DNBA, the mutagenic responses were lower in the nitroreductase-deficient strain TA98NR than in the parental strain. 2,4-DNBA produced a marginally positive response in the V79-cell CA assay; the other substances were devoid of activity. Only the benzoic acids were tested for MN induction in HepG2 cells, and all produced positive responses. As in the bacterial assays, the strongest effect was seen with 2,4,6-TNBA (significant induction at ,1.9 ,M). 4-A-2,6-DNBA was positive at 432 ,M; the weakest effect was observed with 2,4,-DNBA (positive at ,920 ,M). The differences in the sensitivity of the indicator cells to these agents can be explained by differences in the activities of enzymes involved in the activation of the compounds. The strong responses produced by some of the compounds in the human-derived cells suggest that environmental exposure to these breakdown products of nitro-explosives may pose a cancer risk in man. Environ. Mol. Mutagen., 2006. © 2005 Wiley-Liss, Inc. [source] d -Alanyl ester depletion of teichoic acids in Lactobacillus reuteri 100-23 results in impaired colonization of the mouse gastrointestinal tractENVIRONMENTAL MICROBIOLOGY, Issue 7 2007Jens Walter Summary The dlt operon of Gram-positive bacteria encodes proteins required for the incorporation of d -alanine esters into cell wall-associated teichoic acids (TA). d -Alanylation of TA has been shown to be important for acid tolerance, resistance to antimicrobial peptides, adhesion, biofilm formation, and virulence of a variety of pathogenic organisms. The aim of this study was to determine the importance of d -alanylation for colonization of the gastrointestinal tract by Lactobacillus reuteri 100-23. Insertional inactivation of the dltA gene resulted in complete depletion of d -alanine substitution of lipoteichoic acids. The dlt mutant had similar growth characteristics as the wild type under standard in vitro conditions, but formed lower population sizes in the gastrointestinal tract of ex- Lactobacillus -free mice, and was almost eliminated from the habitat in competition experiments with the parental strain. In contrast to the wild type, the dlt mutant was unable to form a biofilm on the forestomach epithelium during gut colonization. Transmission electron microscope observations showed evidence of cell wall damage of mutant bacteria present in the forestomach. The dlt mutant had impaired growth under acidic culture conditions and increased susceptibility to the cationic peptide nisin relative to the wild type. Ex vivo adherence of the dlt mutant to the forestomach epithelium was not impaired. This study showed that d -alanylation is an important cell function of L. reuteri that seems to protect this commensal organism against the hostile conditions prevailing in the murine forestomach. [source] The ttgGHI solvent efflux pump operon of Pseudomonas putida DOT-T1E is located on a large self-transmissible plasmidENVIRONMENTAL MICROBIOLOGY, Issue 6 2007José J. Rodríguez-Herva Summary Pseudomonas putida DOT-T1E is a solvent-tolerant strain able to grow in the presence of > 1% (v/v) toluene in the culture medium. A set of multidrug efflux pumps have been found to play a major role in the tolerance of this bacterium to organic solvents (Rojas et al., J Bacteriol 183: 3967,3973). In the course of studies of the mechanisms underlying solvent tolerance in DOT-T1E, we isolated a spontaneous solvent-sensitive mutant derivative which had lost the genes encoding the TtgGHI efflux pump, the most important extrusion element in quantitative terms. Genomic comparisons between the mutant and its parental strain by microarray analysis revealed that in addition to the ttgVW-ttgGHI gene cluster, another group of genes, highly similar to those found in the Tn4653A and ISPpu12 transposable elements of the TOL plasmid pWW0 from P. putida mt-2, were also absent from this strain. Further analysis demonstrated that strain DOT-T1E harboured a large plasmid (named pGRT1) that was lost from the solvent-sensitive mutant. Mapping analysis revealed that the ttgVW-ttgGHI genes and the Tn4653A -like transposon are borne by the pGRT1 plasmid. Plasmid pGRT1 is highly stable and its frequency of loss is below 10,8 per cell per generation under a variety of growth conditions, including nutritional and physical stresses. The pGRT1 plasmid is self-transmissible, and its acquisition by the toluene-sensitive P. putida KT2440 and Pseudomonas aeruginosa PAO1 increased the recipient's tolerance to toluene up to levels similar to those exhibited by P. putida DOT-T1E. We discuss the importance and potential benefits of this plasmid for the development of bacteria with enhanced solvent tolerance, and its potential impact for bioremediation and whole-cell biotransformations. [source] A NEGATIVE RELATIONSHIP BETWEEN MUTATION PLEIOTROPY AND FITNESS EFFECT IN YEASTEVOLUTION, Issue 6 2007Tim F. Cooper It is generally thought that random mutations will, on average, reduce an organism's fitness because resulting phenotypic changes are likely to be maladaptive. This relationship leads to the prediction that mutations that alter more phenotypic traits, that is, are more pleiotropic, will impose larger fitness costs than mutations that affect fewer traits. Here we present a systems approach to test this expectation. Previous studies have independently estimated fitness and morphological effects of deleting all nonessential genes in Saccharomyces cerevisiae. Using datasets generated by these studies, we examined the relationship between the pleiotropic effect of each deletion mutation, measured as the number of morphological traits differing from the parental strain, and its effect on fitness. Pleiotropy explained ,18% of variation in fitness among the mutants even once we controlled for correlations between morphological traits. This relationship was robust to consideration of other explanatory factors, including the number of protein,protein interactions and the network position of the deleted genes. These results are consistent with pleiotropy having a direct role in affecting fitness. [source] The Helicobacter pylori plasticity region locus jhp0947,jhp0949 is associated with duodenal ulcer disease and interleukin-12 production in monocyte cellsFEMS IMMUNOLOGY & MEDICAL MICROBIOLOGY, Issue 2 2004Ramon De Jonge Abstract Colonization with Helicobacter pylori always results in chronic gastritis, which is controlled by infiltration of mononuclear cells and the subsequent release of cytokines like interleukin (IL)-12. To identify H. pylori factors involved in inducing cytokine production in mononuclear cells, a random H. pylori mutant library was screened for the inability to induce IL-12 production in monocyte THP-1 cells. Of the 231 random mutants screened, one mutant (M1) showed a consistent twofold decrease in the amount of IL-12 induction compared to the parental strain 1061 (P<0.01). Further characterization of mutant M1 revealed that the kanamycin resistance cassette had integrated in the jhp0945 gene, which is situated in an H. pylori strain-specific plasticity region. Three reference strains possessing this plasticity region induced significantly higher amounts of IL-12 when compared to the H. pylori 26695 reference strain, which does not possess this plasticity region. The role in disease outcome of jhp0945 as well as the neighbouring plasticity region genes jhp0947 and jhp049 was assessed in a Dutch population cohort. Firstly, the presence of jhp0947 was completely linked with that of jhp0949 and was roughly associated with jhp0945 (P=0.072), but not with the cag pathogenicity island (PAI) (P=0.464). The presence of the jhp0947 and jhp0949 genes, but not of jhp0945, was significantly associated with duodenal ulcer disease when compared to gastritis (P=0.027). Therefore, the jhp0947,jhp0949 locus may be a novel putative H. pylori marker for disease outcome independent of the cag PAI. [source] Heterologous complementation of the exopolysaccharide synthesis and carbon utilization phenotypes of Sinorhizobium meliloti Rm1021 polyhydroxyalkanoate synthesis mutantsFEMS MICROBIOLOGY LETTERS, Issue 2 2004Punita Aneja Abstract A reduced exopolysaccharide phenotype is associated with inability to synthesize polyhydroxyalkanaote (PHA) stores in Sinorhizobium meliloti strain Rm1021. Loss of function mutations in phbB and phbC result in non-mucoid colony morphology on Yeast Mannitol Agar, compared to the mucoid phenotype exhibited by the parental strain. This phenotype is attributed to reduction in succinoglycan synthesis. We have used complementation of this phenotype and the previously described d -3-hydroxybutyrate/acetoacetate utilization phenotype to isolate a heterologous clone containing a Bradyrhizobium japonicum phbC gene. Sequence analysis confirmed that this clone contains one of the five predicted phbC genes in the B. japonicum genome. The described phenotypic complementation strategy should be useful for isolation of novel PHA synthesis genes of diverse origin. [source] Involvement of RNase G in in vivo mRNA metabolism in Escherichia coliGENES TO CELLS, Issue 5 2001Genryou Umitsuki Background Escherichia coli rng gene (previously called cafA) encodes a novel RNase, named RNase G, which is involved in the 5, end-processing of 16S rRNA. In rng mutant cells, a precursor form of 16S rRNA, 16.3S rRNA, is accumulated. Here we report a role of RNase G in the in vivo mRNA metabolism. Results We found that rng::cat mutant strains overproduced a protein of about 100 kDa. N-terminal amino acid sequencing of this protein showed that it was identical to the fermentative alcohol dehydrogenase, the product of the adhE gene located at 28 min on the E. coli genetic map. The level of adhE mRNA was significantly higher in the rng::cat mutant strain than that in its parental strain, while such differences were not seen in other genes we examined. A rifampicin-chase experiment revealed that the half-life of adhE mRNA was 2.5-fold longer in the rng::cat disruptant than in the wild-type. Conclusion These results indicate that, in addition to rRNA processing, RNase G is involved in in vivo mRNA degradation in E. coli. [source] Mouse chromosome 11 harbors genetic determinants of hippocampal strain-specific nicotinic receptor expressionHIPPOCAMPUS, Issue 8 2008Scott W. Rogers Abstract Differences between isogenic mouse strains in cellular expression of the neuronal nicotinic acetylcholine (ACh) receptor subunit alpha4 (nAChR,4) by the dorsal hippocampus are well known. To investigate further the genetic basis of these variations, expression of the nAChR,4 subunit was measured in congenic mouse lines derived from two strains exhibiting notable divergence in the expression of this subunit: C3H and C57BL/6. Congenic lines carrying reciprocally introgressed regions (quantitative trait loci; QTL) from chromosomes 4, 5, and 12 each retained the phenotype most closely associated with the parental strain. However, in congenic lines harboring the reciprocal transfer of a chromosome 11 QTL, a characteristic difference in the ratio of interneurons versus astrocytes expressing nAChR,4 in the CA1 region is reversed relative to the parental strain. These finding suggest that this chromosomal segment harbors genes that regulate strain distinct hippocampal morphology that is revealed by nAChR,4 expression. © 2008 Wiley-Liss, Inc. [source] Morphological features of Murray Valley encephalitis virus infection in the central nervous system of swiss miceINTERNATIONAL JOURNAL OF EXPERIMENTAL PATHOLOGY, Issue 1 2000Vance Matthews We have examined the histological and ultrastructural features of CNS infection with Murray Valley encephalitis (MVE) virus in mice inoculated with a virulent parental strain (BH3479). Light microscopic examination revealed neuronal necrosis in the olfactory bulb and hippocampus of MVE-infected brains by 5 days post-infection (pi). Electron microscopy of these regions showed endoplasmic reticulum membrane proliferation, and tubular and spherical structures in the cisternae of the endoplasmic reticulum, Golgi complex and nuclear envelope. At seven to eight days pi, infected neurones exhibited chromatin condensation and extrusion, nuclear fragmentation, loss of segments of the nuclear envelope, reduced surface contact with adjacent cells and loss of cytoplasmic organelles. This cell injury was particularly noticeable in the proximal CA3 and distal CA1 regions of the hippocampus. The inflammatory cell profile consisted of macrophages, lymphocytes and especially neutrophils, and many of these inflammatory cells were apoptotic. High mortality rates in the BH3479-infected population of mice correlated with the intense polymorphonuclear and mononuclear leucocyte inflammatory infiltrate in the CNS. [source] The role of GAP1 gene in the nitrogen metabolism of Saccharomyces cerevisiae during wine fermentationJOURNAL OF APPLIED MICROBIOLOGY, Issue 1 2009R. Chiva Abstract Aim:, The aim of this study was to analyse the relevance of the general amino acid permease gene (GAP1) of the wine yeast Saccharomyces cerevisiae on nitrogen metabolism and fermentation performance. Methods and Results:, We constructed a gap1 mutant in a wine strain. We compared fermentation rate, biomass production and nitrogen consumption between the gap1 mutant and its parental strain during fermentations with different nitrogen concentrations. The fermentation capacity of the gap1 mutant strain was impaired in the nitrogen-limited and -excessive conditions. The nitrogen consumption rate between the wild strain and the mutant was different for some amino acids, especially those affected by nitrogen catabolite repression (NCR). The deletion of GAP1 gene also modified the gene expression of other permeases. Conclusions:, The Gap1 permease seems to be important during wine fermentations with low and high nitrogen content, not only because of its amino acid transporter role but also because of its function as an amino acid sensor. Significance and Impact of the Study:, A possible biotechnological advantage of a gap1 mutant is its scarce consumption of arginine, whose metabolism has been related to the production of the carcinogenic ethyl carbamate. [source] Chromate tolerance caused by reduced hydroxyl radical production and decreased glutathione reductase activity in Schizosaccharomyces pombeJOURNAL OF BASIC MICROBIOLOGY, Issue 2 2003Zoltán Gazdag The stable Cr(VI)-tolerant chr1-66T mutant of Schizosaccharomyces pombe, which carries one simple gene mutation responsible for Cr(VI) tolerance, accumulated and reduced the chromate anion (CrO42,) significantly more slowly than did its parental strain 6chr+. The mutant chr1-66T proved to be sensitive to oxidative stressors such as H2O2, menadione, tert -butyl hydroperoxide and Cd2+. Both the Cr(VI) tolerance and the oxidative stress sensitivity were attributed to a decreased specific glutathione reductase activity. These effects were also enhanced with a decrease in the specific mitochondrial Mn-SOD activity. [source] PDR16 -mediated azole resistance in Candida albicansMOLECULAR MICROBIOLOGY, Issue 6 2006Saloua Saidane Summary Many Candida albicans azole-resistant (AR) clinical isolates overexpress the CDR1 and CDR2 genes encoding homologous multidrug transporters of the ATP-binding cassette family. We show here that these strains also overexpress the PDR16 gene, the orthologue of Saccharomyces cerevisiae PDR16 encoding a phosphatidylinositol transfer protein of the Sec14p family. It has been reported that S. cerevisiae pdr16, mutants are hypersusceptible to azoles, suggesting that C. albicans PDR16 may contribute to azole resistance in these isolates. To address this question, we deleted both alleles of PDR16 in an AR clinical strain overexpressing the three genes, using the mycophenolic acid resistance flipper strategy. Our results show that the homozygous pdr16,/pdr16, mutant is approximately twofold less resistant to azoles than the parental strain whereas reintroducing a copy of PDR16 in the mutant restored azole resistance, demonstrating that this gene contributes to the AR phenotype of the cells. In addition, overexpression of PDR16 in azole-susceptible (AS) C. albicans and S. cerevisiae strains increased azole resistance by about twofold, indicating that an increased dosage of Pdr16p can confer low levels of azole resistance in the absence of additional molecular alterations. Taken together, these results demonstrate that PDR16 plays a role in C. albicans azole resistance. [source] Characterization of the Staphylococcus aureus CidR regulon: elucidation of a novel role for acetoin metabolism in cell death and lysisMOLECULAR MICROBIOLOGY, Issue 2 2006Soo-Jin Yang Summary The Staphylococcus aureus cid and lrg operons encode a novel regulatory system that affects murein hydrolase activity, stationary-phase survival and antibiotic tolerance. Expression of the lrgAB operon is positively regulated by a two-component regulatory system encoded by the lytSR operon located immediately upstream to lrgAB. By comparison, the cidABC operon lies downstream from the cidR gene, encoding a protein homologous to the LysR-type family of transcriptional regulators. Transcription analysis of a cidR mutant revealed that CidR enhances cidABC expression in the presence of acetic acid generated by the metabolism of excess glucose. Microarray studies identified additional CidR-regulated operons including the IrgAB and alsSD encoding proteins involved in acetoin production. Surprisingly, Northern blot analyses revealed that cidABC and lrgAB transcription was uninducible in an alsSD mutant grown in the presence of excess glucose, suggesting that the CidR-mediated upregulation of cidABC and lrgAB transcription is dependent on the presence of intact alsSD genes. Zymographic and quantitative analyses of murein hydrolase activity also revealed that disruption of the alsSD genes results in significantly decreased extracellular murein hydrolase activity compared with that of the parental strain, UAMS-1. Furthermore, the alsSD mutant displayed decreased stationary-phase survival relative to UAMS-1, both in the presence and absence of glucose. The results of this study define the CidR regulon and demonstrate that the generation of acetoin is linked to the control of cell death and lysis in S. aureus. [source] The HOG MAP kinase pathway is required for the induction of methylglyoxal-responsive genes and determines methylglyoxal resistance in Saccharomyces cerevisiaeMOLECULAR MICROBIOLOGY, Issue 1 2005Jaime Aguilera Summary A sudden overaccumulation of methylglyoxal (MG) induces, in Saccharomyces cerevisiae, the expression of MG-protective genes, including GPD1, GLO1 and GRE3. The response is partially dependent on the transcriptional factors Msn2p/Msn4p, but unrelated with the general stress response mechanism. Here, we show that the high-osmolarity glycerol (HOG)-pathway controls the genetic response to MG and determines the yeast growth capacity upon MG exposure. Strains lacking the MAPK Hog1p, the upstream component Ssk1p or the HOG-dependent nuclear factor Msn1p, showed a reduction in the mRNA accumulation of MG-responsive genes after MG addition. Moreover, hyperactivation of Hog1p by deletion of protein phosphatase PTP2 enhanced the response, while blocking the pathway by deletion of the MAPKK PBS2 had a negative effect. In addition, the activity of Hog1p affected the basal level of GPD1 mRNA under non-inducing conditions. These effects had a great influence on MG resistance, as hog1, and other HOG-pathway mutants with impaired MG-specific expression displayed MG sensitivity, whereas those with enhanced expression exhibited MG resistance as compared with the wild-type. However, MG does not trigger the overphosphorylation of Hog1p or its nuclear import in the parental strain. Moreover, dual phosphorylation of Hog1p appears to be dispensable in the triggering of the transcriptional response, although a phosphorylable form of Hog1p is fundamental for the transcriptional activity. Overall, our results suggest that the basal activity of the HOG-pathway serves to amplify the expression of MG-responsive genes under non-inducing and inducing conditions, ensuring cell protection against this toxic glycolytic by-product. [source] Bacillus anthracis requires siderophore biosynthesis for growth in macrophages and mouse virulenceMOLECULAR MICROBIOLOGY, Issue 2 2004Stephen Cendrowski Summary Systemic anthrax infections can be characterized as proceeding in stages, beginning with an early intracellular establishment stage within phagocytes that is followed by extracelluar stages involving massive bacteraemia, sepsis and death. Because most bacteria require iron, and the host limits iron availability through homeostatic mechanisms, we hypothesized that B. anthracis requires a high-affinity mechanism of iron acquisition during its growth stages. Two putative types of siderophore synthesis operons, named Bacillus anthracis catechol, bac (anthrabactin), and anthrax siderophore biosynthesis, asb (anthrachelin), were identified. Directed gene deletions in both anthrabactin and anthrachelin pathways were generated in a B. anthracis (Sterne) 34F2 background resulting in mutations in asbA and bacCEBF. A decrease in siderophore production was observed during iron-depleted growth in both the ,asbA and ,bacCEBF strains, but only the ,asbA strain was attenuated for growth under these conditions. In addition, the ,asbA strain was severely attenuated both for growth in macrophages (M,) and for virulence in mice. In contrast, the ,bacCEBF strain did not differ phenotypically from the parental strain. These findings support a requirement for anthrachelin but not anthrabactin in iron assimilation during the intracellular stage of anthrax. [source] Haem utilization in Vibrio cholerae involves multiple TonB-dependent haem receptorsMOLECULAR MICROBIOLOGY, Issue 3 2001Alexandra R. Mey Vibrio cholerae has multiple iron transport systems, one of which involves haem uptake through the outer membrane receptor HutA. A hutA mutant had only a slight defect in growth using haemin as the iron source, and we show here that V. cholerae encodes two additional TonB-dependent haem receptors, HutR and HasR. HutR has significant homology to HutA as well as to other outer membrane haem receptors. Membrane fractionation confirmed that HutR is present in the outer membrane. The hutR gene was co-transcribed with the upstream gene ptrB, and expression from the ptrB promoter was negatively regulated by iron. A hutA, hutR mutant was significantly impaired, but not completely defective, in the ability to use haemin as the sole iron source. HasR is most similar to the haemophore-utilizing haem receptors from Pseudomonas aeruginosa and Serratia marcescens. A mutant defective in all three haem receptors was unable to use haemin as an iron source. HutA and HutR functioned with either V. cholerae TonB1 or TonB2, but haemin transport through either receptor was more efficient in strains carrying the tonB1 system genes. In contrast, haemin uptake through HasR was TonB2 dependent. Efficient utilization of haemoglobin as an iron source required HutA and TonB1. The triple haem receptor mutant exhibited no defect in its ability to compete with its Vib, parental strain in an infant mouse model of infection, indicating that additional iron sources are present in vivo. V. cholerae used haem derived from marine invertebrate haemoglobins, suggesting that haem may be available to V. cholerae growing in the marine environment. [source] Effects of Bacillus thuringiensis Cry1C toxin on the metabolic rate of Cry1C resistant and susceptible Spodoptera exigua (Lepidoptera: Noctuidae)PHYSIOLOGICAL ENTOMOLOGY, Issue 5 2004Beatrice N. Dingha Abstract., The effects of Bacillus thuringiensis (Bt) Cry1C toxin on the metabolic rate of Cry1C resistant and susceptible Spodoptera exigua (Hübner) (Lepidoptera: Noctuidae) are investigated using closed-system respirometry. Mechanisms of resistance to the Bt toxin may be associated with an energetic cost that can be measured as an increase in metabolic rate compared with Bt-susceptible insects. This hypothesis is tested using third- and fifth-instar larvae and 1,7-day-old pupae. Metabolic rate is measured as the amount of O2 consumed and CO2 produced. V,O2 and V,CO2 (mL g,1 h,1) of third-instar Cry1C resistant larvae reared continuously on a diet containing 320 µg Cry1C toxin per g diet (CryonT) are significantly greater than third-instar Cry1C resistant larvae reared on toxin for 5 days and reared thereafter on untreated diet (Cry5dT), Cry1C resistant larvae reared on untreated diet (CryReg) and the susceptible parental strain (SeA) reared on untreated diet. There are no differences in V,O2 and V,CO2 (mL g,1 h,1) among treatment groups for fifth-instar larvae. CryonT larvae and pupae weigh significantly less than larvae and pupae receiving other treatments. Smaller body mass may be an important biological cost to individuals exposed continuously to Bt toxin. One-day-old pupae of all treatment groups exhibit a high V,O2 (mean approximately 0.174 mL g,1 h,1) with CryonT having a significantly greater value than all other treatments; there are no differences among the other treatments. Pupal metabolic rates of all treatment groups decline to a minimum between days 2 and 4 then increase linearly between days 4 and 7 until adult emergence. These results demonstrate no difference in metabolic rates, and possibly fitness costs, between resistant (CryReg and Cry5dT) and susceptible (SeA) S. exigua except when larvae were reared continuously on toxin (CryonT). [source] A comparative proteomic approach to understand the adaptations of an H+ -ATPase-defective mutant of Corynebacterium glutamicum ATCC14067 to energy deficienciesPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 18 2007Liyuan Li Abstract F172-8, an H+ -ATPase-defective mutant of the glutamic acid-producing bacterium Corynebacterium glutamicum ATCC 14067, exhibits enhanced rates of glucose consumption and respiration compared to the parental strain when cultured in a biotin-rich medium with glucose as the carbon source. We conducted a comparative proteomic analysis to clarify the mechanism by which the enhanced glucose metabolism in this mutant is established using a proteome reference map for strain ATCC 14067. A comparison of the proteomes of the two strains revealed the up-regulated expression of the several important enzymes such as pyruvate kinase (Pyk), malate:quinone oxidoreductase (Mqo), and malate dehydrogenase (Mdh) in the mutant. Because Pyk activates glycolysis in response to cellular energy shortages in this bacterium, its increased expression may contribute to the enhanced glucose metabolism of the mutant. A unique reoxidation system has been suggested for NADH in C. glutamicum consisting of coupled reactions between Mqo and Mdh, together with the respiratory chain; therefore, the enhanced expression of both enzymes might contribute to the reoxidation of NADH during increased respiration. The proteomic analysis allowed the identification of unique physiological changes associated with the H+ -ATPase defect in F172-8 and contributed to the understanding of the adaptations of C. glutamicum to energy deficiencies. [source] Isoprene Formation in Bacillus subtilis: A Barometer of Central Carbon Assimilation in a Bioreactor?BIOTECHNOLOGY PROGRESS, Issue 5 2002Megan C. Shirk Isoprene (2-methyl-1,3-butadiene) is a volatile hydrocarbon of uncertain function in Bacillus subtilis, and we hypothesized that it is an overflow metabolite produced during excess carbon utilization. Here we tested this idea for phase 2 of isoprene release, a phase that occurs during extracellular acetoin accumulation and its reassimilation. Phase 2 isoprene formation could be disrupted in three different ways, all related to acetoin metabolism. Disruption of a gene essential for acetoin biosynthesis (acetolactic acid synthase, alsS) blocked acetoin formation and led to cessation of phase 2 isoprene formation as well as a variety of pleiotropic effects related to loss of pH control. Growth of the alsS mutant with external pH control reversed most of these effects. Disruption of acetoin catabolism (acetoin dehydrogenase, acoA), also eliminated phase 2 isoprene formation and caused cells to transition directly from phase 1 to phase 3; the latter is attributed to amino acid catabolism. A third alteration of acetoin metabolism was detected in the widely used strain 168 ( trpC2) but not in strain MS175, a trpC mutant constructed in the Marburg strain genetic background. Strain 168 exhibited slow acetoin assimilation compared to that of MS175 or the parental strain, with little or no isoprene formation during this growth phase. These findings support the idea that isoprene release occurs primarily when the rate of carbon catabolism exceeds anabolism and that this volatile hydrocarbon is a product of overflow metabolism when precursors are not required for higher isoprenoid biosynthesis. It is suggested that isoprene release might serve as a useful barometer of the rise and fall of central carbon fluxes during the growth of Bacillus strains in industrial bioreactors. [source] Congenic Rats For Hypertension: How Useful Are They For The Hunting Of Hypertension Genes?CLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 4 2000Toru Nabika SUMMARY 1. Linkage studies have revealed quantitative trait loci (QTL) for blood pressure in the rat genome using genetic hypertensive rat models. To identify the genes responsible for hypertension, the construction of congenic rats is essential. 2. To date, several congenic strains have been obtained from spontaneously hypertensive or Dahl salt-sensitive rats. The results of these studies should be interpreted according to whether the rats carry the whole QTL region or not. 3. After establishing congenic strains, three strategies are possible: (i) an orthodox positional cloning in which, using subcongenic strains, the QTL region is cut down to smaller fragments suitable for physical mapping; (ii) a positional candidate strategy in which candidate genes in the QTL regions are studied; or (iii) physiological studies in which intermediate phenotypes directly associated with the hypertension gene are explored. Several other experimental strategies are also available using congenic strains as new animal models for hypertension. 4. To make the most of advances in DNA technology, the precise evaluation of the phenotypic difference between congenic strains carrying different QTL or between a congenic and parental strain is critical. [source] Identification of a Chr 11 quantitative trait locus that modulates proliferation in the rostral migratory stream of the adult mouse brainEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 4 2010Anna Poon Abstract Neuron production takes place continuously in the rostral migratory stream (RMS) of the adult mammalian brain. The molecular mechanisms that regulate progenitor cell division and differentiation in the RMS remain largely unknown. Here, we surveyed the mouse genome in an unbiased manner to identify candidate gene loci that regulate proliferation in the adult RMS. We quantified neurogenesis in adult C57BL/6J and A/J mice, and 27 recombinant inbred lines derived from those parental strains. We showed that the A/J RMS had greater numbers of bromodeoxyuridine-labeled cells than that of C57BL/6J mice with similar cell cycle parameters, indicating that the differences in the number of bromodeoxyuridine-positive cells reflected the number of proliferating cells between the strains. AXB and BXA recombinant inbred strains demonstrated even greater variation in the numbers of proliferating cells. Genome-wide mapping of this trait revealed that chromosome 11 harbors a significant quantitative trait locus at 116.75 ± 0.75 Mb that affects cell proliferation in the adult RMS. The genomic regions that influence RMS proliferation did not overlap with genomic regions regulating proliferation in the adult subgranular zone of the hippocampal dentate gyrus. On the contrary, a different, suggestive locus that modulates cell proliferation in the subgranular zone was mapped to chromosome 3 at 102 ± 7 Mb. A subset of genes in the chromosome 11 quantitative trait locus region is associated with neurogenesis and cell proliferation. Our findings provide new insights into the genetic control of neural proliferation and an excellent starting point to identify genes critical to this process. [source] Genetic determinants of adult hippocampal neurogenesis correlate with acquisition, but not probe trial performance, in the water maze taskEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 1 2002G. Kempermann Abstract A number of reports have indicated that adult neurogenesis might be involved in hippocampal function. While increases in adult neurogenesis are paralleled by improvements on learning tasks and learning itself can promote the survival of newly generated neurons in the hippocampus, a causal link between learning processes and adult hippocampal neurogenesis is difficult to prove. Here, we addressed the related question of whether the baseline level of adult neurogenesis is predictive of performance on the water maze task as a test of hippocampal function. We used ten strains of recombinant inbred mice, based on C57BL/6, which are good learners and show high baseline levels of neurogenesis, and DBA/2, which are known to be poor learners and which exhibit low levels of adult neurogenesis. Two of these strains, BXD-2 and BXD-8, showed a 26-fold difference in the number of newly generated neurons per hippocampus. Over all strains, including the parental strains, there was a significant correlation between the number of new neurons generated in the dentate gyrus and parameters describing the acquisition of the water maze task (slope of the learning curves). Similar results were seen when the parental strains were not included in the analysis. There was no correlation between adult hippocampal neurogenesis and probe trial performance, performance on the rotarod, overall locomotor activity, and baseline serum corticosterone levels. This result supports the hypothesis that adult neurogenesis is involved in specific aspects of hippocampal function, particularly the acquisition of new information. [source] Evidence that the keratinocyte colony number is genetically controlledEXPERIMENTAL DERMATOLOGY, Issue 6 2002Natalia V. Popova Abstract: We tested five inbred strains and two outbred stocks of female mice in a quantitative assay for clonogenic keratinocytes from the cutaneous epithelium. We found three significantly different subsets of colony counts such that: C57BL/6 , C3H = DBA/2 = SENCAR = BALB/c > FVB = CD,1 in culture conditions optimized for CD,1 0. C57BL/6 and BALB/c, two inbred parental strains, were chosen for further analysis. The F1 generation of these two parental strains had an intermediate number of colonies. The keratinocyte colony number from the two backcross generations was significantly different, while the colony number in the F2 generation was intermediate between the two backcrosses. We conclude that the number of keratinocyte colonies represents a new genetically definable quantitative trait. Analysis suggests that this trait is multigenic where the genes have an additive but not necessarily equal effect. We have therefore laid the foundation for identifying these stem cell regulatory genes, which may provide a new perspective on the mechanism of carcinogenesis and a new target for gene therapy. [source] Use of a breeding approach for improving biocontrol efficacy of Phlebiopsis gigantea strains against Heterobasidion infection of Norway spruce stumpsFEMS MICROBIOLOGY ECOLOGY, Issue 2 2009Hui Sun Abstract Sixty-four wild heterokaryotic isolates of Phlebiopsis gigantea were analysed for asexual spore production, growth rate and competitive ability against Heterobasidion in vitro, as well as growth rate in Norway spruce wood. These P. gigantea traits were considered important for controlling infection of Norway spruce stumps by spores of Heterobasidion spp. Ten most promising P. gigantea isolates were crossed with each other and 172 F1 progeny heterokaryons were analysed for the above-mentioned traits. Thirteen most promising progeny heterokaryons were selected and their biocontrol ability against infection by Heterobasidion was compared with the parental isolates in stem pieces of Norway spruce. The results indicated that the progeny strains had generally better traits and control efficacy than the parental strains. The genetic effects accounted for a part of the variations between progeny and parental strains. This further suggests that there is a potential to improve the biocontrol properties of P. gigantea through breeding. [source] Magnetic resonance imaging as a tool for in vivo and ex vivo anatomical phenotyping in experimental genetic modelsHUMAN BRAIN MAPPING, Issue 6 2007Alain Pitiot Abstract This article describes a suite of computational approaches suitable for deriving various quantitative phenotypes from structural magnetic resonance (MR) images obtained in rodents and used subsequently in genetic studies of complex traits. We begin by introducing the basic principles of genetic studies of complex traits in experimental models. We then illustrate the use of MR-based computational anatomy in vivo and ex vivo, and in combination with histology. This work was carried out in two inbred strains of rats, namely spontaneously hypertensive rats and Brown Norway rats; these are parental strains of the only existing panel of recombinant inbred strains of rats. The rats were scanned in vivo at two time points (at 8 and 12 weeks of age) and ex vivo (at 12 weeks of age). We describe between-strain differences and across-time changes in brain and kidney volumes, as well as regional variations in brain structure using surface- and deformation-based approaches. We conclude by discussing the power of the population-based computational analysis of MR images, and their fusion with histology, in studies of complex traits. Hum Brain Mapp, 2007. © 2007 Wiley-Liss, Inc. [source] Non-random reassortment in human influenza A virusesINFLUENZA AND OTHER RESPIRATORY VIRUSES, Issue 1 2008Raul Rabadan Background, The influenza A virus has two basic modes of evolution. Because of a high error rate in the process of replication by RNA polymerase, the viral genome drifts via accumulated mutations. The second mode of evolution is termed a shift, which results from the reassortment of the eight segments of this virus. When two different influenza viruses co-infect the same host cell, new virions can be released that contain segments from both parental strains. This type of shift has been the source of at least two of the influenza pandemics in the 20th century (H2N2 in 1957 and H3N2 in 1968). Objectives, The methods to measure these genetic shifts have not yet provided a quantitative answer to questions such as: what is the rate of genetic reassortment during a local epidemic? Are all possible reassortments equally likely or are there preferred patterns? Methods, To answer these questions and provide a quantitative way to measure genetic shifts, a new method for detecting reassortments from nucleotide sequence data was created that does not rely upon phylogenetic analysis. Two different sequence databases were used: human H3N2 viruses isolated in New York State between 1995 and 2006, and human H3N2 viruses isolated in New Zealand between 2000 and 2005. Results, Using this new method, we were able to reproduce all the reassortments found in earlier works, as well as detect, with very high confidence, many reassortments that were not detected by previous authors. We obtain a lower bound on the reassortment rate of 2,3 events per year, and find a clear preference for reassortments involving only one segment, most often hemagglutinin or neuraminidase. At a lower frequency several segments appear to reassort in vivo in defined groups as has been suggested previously in vitro. Conclusions, Our results strongly suggest that the patterns of reassortment in the viral population are not random. Deciphering these patterns can be a useful tool in attempting to understand and predict possible influenza pandemics. [source] Functional enhancement of Sake yeast strains to minimize the production of ethyl carbamate in Sake wineJOURNAL OF APPLIED MICROBIOLOGY, Issue 3 2010M.S. Dahabieh Abstract Aims:, In fermented alcoholic beverages and particularly in Japanese Sake wine, the ubiquitous presence of the probable human carcinogen ethyl carbamate (EC) is a topic of significant concern. This study aims to develop novel methods for the reduction of EC in Sake wine. Methods and Results:, To reduce the high levels of EC in Sake wine, urea-degrading and urea-importing yeast strains were created by integrating linear cassettes containing either the respective DUR1,2 or DUR3 genes, under the control of the constitutively active Saccharomyces cerevisiae PGK1 promoter, into the Sake yeast strains K7 and K9. The self-cloned, urea-degrading Sake strains K7DUR1,2 and K9DUR1,2 produced Sake wine with 87 and 68% less EC, respectively, while the urea-importing Sake yeast strain K7DUR3 reduced EC by 15%. All functionally enhanced yeast strains were shown to be substantially equivalent to their parental strains in terms of fermentation rate, ethanol production, phenotype and transcriptome. Conclusions:, Under the conditions tested, urea-degrading yeast (constitutive DUR1,2 expression) are superior to urea-importing yeast (constitutive DUR3 expression) for EC reduction in Sake wine, and constitutive co-expression of DUR1,2 and DUR3 does not yield synergistic EC reduction. Significance and Impact of the Study:, The self-cloned, substantially equivalent, urea-degrading Sake yeast strains K7DUR1,2 and K9DUR1,2, which contain the integrated DUR1,2 cassette, are capable of highly efficacious EC reduction during Sake brewing trials, are suitable for commercialization and are important tools for modern Sake makers in their efforts to reduce high EC levels in Sake wine. [source] Nisin-resistant (Nisr) Listeria monocytogenes and Nisr Clostridium botulinum Are Not Resistant to Common Food PreservativesJOURNAL OF FOOD SCIENCE, Issue 5 2000A.S. MAZZOTTA ABSTRACT Nisin-resistant (Nisr) strains of Clostridium botulinum and Listeria monocytogenes may arise as nisin becomes more widely used as an additional safety barrier in minimally-processed foods. The sensitivity of Nisr L. monocytogenes ATCC 700301 and ATCC 700302 and toxigenic Nisr C. botulinum 169B to low pH, salt, sodium nitrite, and potassium sorbate was assayed using discontinuous gradients in broth and compared to the parental wild-type strains. The nisin-resistant strains did not have intrinsic resistance to low pH, sodium chloride, potassium sorbate, or sodium nitrite. In no case were the Nisr L. monocytogenes and C. botulinum strains examined more resistant to inhibitors than the parental strains. [source] Dopamine D2 Receptor Binding, Drd2 Expression and the Number of Dopamine Neurons in the BXD Recombinant Inbred Series: Genetic Relationships to Alcohol and Other Drug Associated PhenotypesALCOHOLISM, Issue 1 2003Robert Hitzemann Background: It has not been established to what extent the natural variation in dopamine systems contribute to the variation in ethanol response. The current study addresses this issue by measuring D2 dopamine (DA) receptor binding, the expression of Drd2, the number of midbrain DA neurons in the BXD recombinant inbred (RI) series and then compares these strain means with those previously reported for a variety of ethanol and other drug-related phenotypes. Methods: Data were collected for 21 to 23 of the BXD RI strains and the parental strains. D2 DA receptor autoradiography was performed using 125I-epidepride as the ligand [Kanes S, Dains K, Cipp L, Gatley J, Hitzemann B, Rasmussen E, Sanderson S, Silverman S, Hitzemann R (1996) Mapping the genes for haloperidol-induced catalepsy. J Pharmacol Exp Ther 277:1016,1025]. Drd2 expression was measured using the Affymetrix oligoarray system. Immunocytochemical techniques were used to determine the number of midbrain DA neurons [Hitzemann B, Dains K, Hitzemann R (1994) Further studies on the relationship between dopamine cell density and haloperidol response. J Pharmacol Exp Ther 271:969,976]. Results and Conclusions: The range of difference in receptor binding for the RI strains was approximately 2-fold in all regions examined, the core, the shell of the nucleus accumbens (NAc) and the dorsomedial caudate-putamen (CPu); heritability in all regions was moderate,(h 2,0.35). Drd2 expression in forebrain samples from the RI and parental strains ranged 1.5- to 2-fold and h2 was moderate,0.47. Variation in the number of tyrosine hydroxylase (TH) positive neurons was moderate, 41% and 26% and h2 was low,0.19 and 0.15 for the ventral tegmental area (VTA) and substantia nigra compacta (SNc), respectively. Significant correlations were found between D2 DA receptor binding and the low dose (1.33 g/kg) ethanol stimulant response. (p < 0.002) and between Drd2 expression and conditioned place preference (CPP) (p < 0.0005). No significant correlations were detected between ethanol preference and either receptor binding or Drd2 expression; however, a significant correlation was found between preference and Ncam expression. Ncam is approximately 0.2 Mb from Drd2. Overall, the data suggest ethanol preference and CPP are associated with the expression of Drd2 or closely linked genetic loci. [source] Single nucleotide polymorphism genotyping of the barley waxy gene by polymerase chain reaction with confronting two-pair primersPLANT BREEDING, Issue 3 2004E. Domon Abstract A high-throughput single nucleotide polymorphism (SNP) genotyping procedure was developed to select amylose-free barley mutants whose waxy genes had a C- to T-base substitution in exon 5, which converted Gln-89 of the wild-type gene into a termination codon. An F2 population carrying an amylose-free waxy gene was checked for segregation. Polymerase chain reaction with confronting two-pair primers (PCR-CTPP) produced allele-specific PCR products that have different sizes and are inherited in a co-dominant manner. Two alleles of the barley waxy gene with SNP were correctly identified in parental strains using the PCR-CTPP procedure. Segregation of the SNP as detected by PCR-CTPP in an F2 population fitted the expected 1:2:1 ratio. The PCR-CTPP procedure can provide a time saving and cost-effective alternative to derived cleaved amplified polymorphic sequence in marker-assisted selection. [source] | ||||||||||||||||||||||||||||