Familial aggregation of VO(2max) response to exercise training: results from the HERITAGE Family Study.

The aim of this study was to test the hypothesis that individual differences in the response of maximal O(2) uptake (VO(2max)) to a standardized training program are characterized by familial aggregation. A total of 481 sedentary adult Caucasians from 98 two-generation families was exercise trained for 20 wk and was tested for VO(2max) on a cycle ergometer twice before and twice after the training program. The mean increase in VO(2max) reached approximately 400 ml/min, but there was considerable heterogeneity in responsiveness, with some individuals experiencing little or no gain, whereas others gained >1.0 l/min. An ANOVA revealed that there was 2.5 times more variance between families than within families in the VO(2max) response variance. With the use of a model-fitting procedure, the most parsimonious models yielded a maximal heritability estimate of 47% for the VO(2max) response, which was adjusted for age and sex with a maternal transmission of 28% in one of the models. We conclude that the trainability of VO(2max) is highly familial and includes a significant genetic component.     

Genotypic Diversity among Brevibacillus laterosporus Strains

In comparison with other entomopathogenic Bacillus species, the genome of Brevibacillus laterosporus is poorly characterized. The aim of this study was to examine genetic variability in B. laterosporus by using a range of typing methodologies. Strains of B. laterosporus were examined for variation in 13 chromosomal genes encoding enzymes by multilocus enzyme electrophoresis. Optimal conditions of pulsed-field gel electrophoresis and randomly amplified polymorphic DNA were established that allowed analysis of the genome of B. laterosporus. None of these techniques allowed the identification of a convenient molecular marker for entomopathogenic strains, although one specific primer amplified only DNA from almost all mosquitocidal strains.     

Biotransformation of 2,4,6-Trinitrotoluene with Phanerochaete chrysosporium in Agitated Cultures at pH 4.5

The biotransformation of 2,4,6-trinitrotoluene (TNT) (175 μM) by Phanerochaete chrysosporium with molasses and citric acid at pH 4.5 was studied. In less than 2 weeks, TNT disappeared completely, but mineralization (liberated ¹⁴CO₂) did not exceed 1%. A time study revealed the presence of several intermediates, marked by the initial formation of two monohydroxylaminodinitrotoluenes (2- and 4-HADNT) followed by their successive transformation to several other products, including monoaminodinitrotoluenes (ADNT). A group of nine acylated intermediates were also detected. They included 2-N-acetylamido-4,6-dinitrotoluene and its p isomer, 2-formylamido-4,6-dinitrotoluene and its p isomer (as acylated ADNT), 4-N-acetylamino-2-amino-6-nitrotoluene and 4-N-formylamido-2-amino-6-nitrotoluene (as acetylated DANT), 4-N-acetylhydroxy-2,6-dinitrotoluene and 4-N-acetoxy-2,6-dinitrotoluene (as acetylated HADNT), and finally 4-N-acetylamido-2-hydroxylamino-6-nitrotoluene. Furthermore, a fraction of HADNTs were found to rearrange to their corresponding phenolamines (Bamberger rearrangement), while another group dimerized to azoxytoluenes which in turn transformed to azo compounds and eventually to the corresponding hydrazo derivatives. After 30 days, all of these metabolites, except traces of 4-ADNT and the hydrazo derivatives, disappeared, but mineralization did not exceed 10% even after the incubation period was increased to 120 days. The biotransformation of TNT was accompanied by the appearance of manganese peroxidase (MnP) and lignin-dependent peroxidase (LiP) activities. MnP activity was observed almost immediately after TNT disappearance, which was the period marked by the appearance of the initial metabolites (HADNT and ADNT), whereas the LiP activity was observed after 8 days of incubation, corresponding to the appearance of the acyl derivatives. Both MnP and LiP activities reached their maximum levels (100 and 10 U/liter, respectively) within 10 to 15 days after inoculation.     

Viability and Virulence of Experimentally Stressed Nonculturable Salmonella typhimurium

Maintenance of pathogenicity of viable but nonculturable Salmonella typhimurium cells experimentally stressed with UV-C and seawater, was investigated relative to the viability level of the cellular population. Pathogenicity, tested in a mouse model, was lost concomitantly with culturability, whereas cell viability remained undamaged, as determined by respiratory activity and cytoplasmic membrane and genomic integrities.     

Cloning and Characterization of the Genes Encoding a Cytochrome P450 (PipA) Involved in Piperidine and Pyrrolidine Utilization and Its Regulatory Protein (PipR) in Mycobacterium smegmatis mc2155

Transposon mutagenesis of Mycobacterium smegmatis mc2155 enabled the isolation of a mutant strain (called LGM1) altered in the regulation of piperidine and pyrrolidine utilization. The complete nucleotide sequence of the gene inactivated in mutant LGM1 was determined from the wild-type strain. This gene (pipR) encoded a member of the GntR family of bacterial regulatory proteins. An insertion element (IS1096), previously described for M. smegmatis, was detected downstream of the gene pipR. Three additional open reading frames were found downstream of IS1096. The first open reading frame (pipA) appeared to encode a protein identified as a cytochrome P450 enzyme. This gene is the first member of a new family, CYP151. By a gene replacement experiment, it was demonstrated that the cytochrome P450 pipA gene is required for piperidine and pyrrolidine utilization in M. smegmatis mc2155. Genes homologous to pipA were detected by hybridization in several, previously isolated, morpholine-degrading mycobacterial strains. A gene encoding a putative [3Fe-4S] ferredoxin (orf1) and a truncated gene encoding a putative glutamine synthetase (orf2') were found downstream of pipA.     

The impact of mating and sugar feeding on blood-feeding physiology and behavior in the arbovirus vector mosquito Aedes aegypti

BackgroundAedes aegypti mosquitoes are globally distributed vectors of viruses that impact the health of hundreds of millions of people annually. Mating and blood feeding represent fundamental aspects of mosquito life history that carry important implications for vectorial capacity and for control strategies. Females transmit pathogens to vertebrate hosts and obtain essential nutrients for eggs during blood feeding. Further, because host-seeking Ae. aegypti females mate with males swarming near hosts, biological crosstalk between these behaviors could be important. Although mating influences nutritional intake in other insects, prior studies examining mating effects on mosquito blood feeding have yielded conflicting results.Methodology/Principal findingsTo resolve these discrepancies, we examined blood-feeding physiology and behavior in virgin and mated females and in virgins injected with male accessory gland extracts (MAG), which induce post-mating changes in female behavior. We controlled adult nutritional status prior to blood feeding by using water- and sugar-fed controls. Our data show that neither mating nor injection with MAG affect Ae. aegypti blood intake, digestion, or feeding avidity for an initial blood meal. However, sugar feeding, a common supplement in laboratory settings but relatively rare in nature, significantly affected all aspects of feeding and may have contributed to conflicting results among previous studies. Further, mating, MAG injection, and sugar intake induced declines in subsequent feedings after an initial blood meal, correlating with egg production and laying. Taking our evaluation to the field, virgin and mated mosquitoes collected in Colombia were equally likely to contain blood at the time of collection.Conclusions/SignificanceMating, MAG, and sugar feeding impact a mosquito’s estimated ability to transmit pathogens through both direct and indirect effects on multiple aspects of mosquito biology. Our results highlight the need to consider natural mosquito ecology, including diet, when assessing their physiology and behavior in the laboratory.     

The Nuclease Activity of Mre11 Is Required for Meiosis but Not for Mating Type Switching, End Joining, or Telomere Maintenance

The Saccharomyces cerevisiae MRE11 gene is required for the repair of ionizing radiation-induced DNA damage and for the initiation of meiotic recombination. Sequence analysis has revealed homology between Mre11 and SbcD, the catalytic subunit of an Escherichia coli enzyme with endo- and exonuclease activity, SbcCD. In this study, the purified Mre11 protein was found to have single-stranded endonuclease activity. This activity was absent from mutant proteins containing single amino acid substitutions in either one of two sequence motifs that are shared by Mre11 and SbcD. Mutants with allele mre11-D56N or mre11-H125N were partially sensitive to ionizing radiation but lacked the other mitotic phenotypes of poor vegetative growth, hyperrecombination, defective nonhomologous end joining, and shortened telomeres that are characteristic of the mre11 null mutant. Diploids homozygous for the mre11-H125N mutation failed to sporulate and accumulated unresected double-strand breaks (DSB) during meiosis. We propose that in mitotic cells DSBs can be processed by other nucleases that are partially redundant with Mre11, but these activities are unable to process Spo11-bound DSBs in meiotic cells.     

Experimental evidence for predation risk sensitive oviposition by a mosquito, Culiseta longiareolata

1. Females should choose to oviposit in habitats where risk of predation and competition are low. The ovipositional responses of a mosquito, Culiseta longiareolata, to a predator and to species sharing the same trophic level as this mosquito (controphic species) were assessed experimentally in outdoor artificial pools. 2. The predator, larval Anax imperator, which strongly reduced larval C. longiareolata survival, resulted in a 52% reduction of C. longiareolata egg rafts. The controphic species (primarily Daphnia magna), which had a small but statistically significant negative effect on the survival of C. longiareolata larvae, did not have a statistically significant influence on the number of egg rafts. 3. Laboratory trials indicated that only a small fraction of the reduced number of egg rafts seen in predator pools may be due to consumption of the egg rafts by A. imperator. 4. The experimental evidence indicates that the reduced number of C. longiareolata egg rafts found in the presence of A. imperator is due largely to oviposition habitat selection, i.e. C. longiareolata females choose pools with low risk of predation for their offspring.     

Expression of a Recombinant Toxoplasma gondii ROP2 Fragment as a Fusion Protein in Bacteria Circumvents Insolubility and Proteolytic Degradation

A 268-amino-acid-residue carboxy-terminal antigenic fragment of the Toxoplasma gondii rhoptry protein ROP2 (recROP2_t, residues 196–464) was expressed in Escherichia coli. This recombinant fragment was produced at low concentration and in a highly insoluble form. By contrast, the level of recROP2_t production was drastically greater when the same coding sequence was fused to the C-terminus of thioredoxin (TRX) or to the maltose-binding protein (MBP) gene. While both fusion proteins were found to be mainly insoluble, solubilization could be achieved without significant degradation. MBP was more efficient than TRX in increasing the recovery of soluble protein with more than 10% of total MBP–recROP2_t being readily expressed in a soluble form. Moreover, the insoluble form of MBP–recROP2_t could be correctly refolded with a recovery of more than 80%. Both forms of MBP–recROP2_t were purified to homogeneity by amylose chromatography. In contrast, the refolding of TRX–recROP2_t promoted aggregation of the protein, which was prevented by the use of zwitterionic detergent during the one-step purification by gel filtration. Subsequent proteolytic cleavages of purified TRX–recROP2_t and of MBP–recROP2_t led respectively to the complete degradation or to the truncation of the recROP2_t moiety. However, recROP2_t, despite the presence of the fusion partners, adopted a suitable conformation recognized by human serum-derived antibodies from T. gondii-seropositive individuals. Finally, both fusion proteins were able to induce specific humoral and cell-mediated immune response to the ROP2 fragment. Such fusions could represent an alternative to study the immunogenicity of T. gondii proteins which are difficult to produce because of insolubility and degradation.