Comparison of expression of key sporulation,solventogenic and acetogenic genes in C. beijerinckii NRRL B-598 and its mutant strain overexpressing spo0A

Applied Microbiology and Biotechnology - The production of acetone, butanol and ethanol by fermentation of renewable biomass has potential to become a valuable industrial process. Mechanisms of...     

Pseudomonas syringae naturally lacking the canonical type III secretion system are ubiquitous in nonagricultural habitats,are phylogenetically diverse and can be pathogenic

The type III secretion system (T3SS) is an important virulence factor of pathogenic bacteria, but the natural occurrence of variants of bacterial plant pathogens with deficiencies in their T3SS raises questions about the significance of the T3SS for fitness. Previous work on T3SS-deficient plant pathogenic bacteria has focused on strains from plants or plant debris. Here we have characterized T3SS-deficient strains of Pseudomonas syringae from plant and nonplant substrates in pristine nonagricultural contexts, many of which represent recently described clades not yet found associated with crop plants. Strains incapable of inducing a hypersensitive reaction (HR⁻) in tobacco were detected in 65% of 126 samples from headwaters of rivers (mountain creeks and lakes), snowpack, epilithic biofilms, wild plants and leaf litter and constituted 2 to 100% of the P. syringae population associated with each sample. All HR⁻ strains lacked at least one gene in the canonical hrp/hrc locus or the associated conserved effector locus, but most lacked all six of the genes tested (hrcC, hrpL, hrpK1, avrE1 and hrpW1) and represented several disparate phylogenetic clades. Although most HR⁻ strains were incapable of causing symptoms on cantaloupe seedlings as expected, strains in the recently described TA-002 clade caused severe symptoms in spite of the absence of any of the six conserved genes of the canonical T3SS according to PCR and Southern blot assays. The phylogenetic context of the T3SS variants we observed provides insight into the evolutionary history of P. syringae as a pathogen and as an environmental saprophyte.     

High-throughput Titration of Luciferase-expressing Recombinant Viruses

Standard plaque assays to determine infectious viral titers can be time consuming, are not amenable to a high volume of samples, and cannot be done with viruses that do not form plaques. As an alternative to plaque assays, we have developed a high-throughput titration method that allows for the simultaneous titration of a high volume of samples in a single day. This approach involves infection of the samples with a Firefly luciferase tagged virus, transfer of the infected samples onto an appropriate permissive cell line, subsequent addition of luciferin, reading of plates in order to obtain luminescence readings, and finally the conversion from luminescence to viral titers. The assessment of cytotoxicity using a metabolic viability dye can be easily incorporated in the workflow in parallel and provide valuable information in the context of a drug screen. This technique provides a reliable, high-throughput method to determine viral titers as an alternative to a standard plaque assay.     

Malaria transmission in a central area of Futa Djalon (Guinea): results of a parasitological survey during the 1989 rainy season.

A malaria survey based on household surveys and dispensary visits without notice was carried out during the rainy season of 1989 in three selected areas of central Futa Djalon (Republic of Guinea). Preliminary entomological evidence showed that Anopheles gambiae was the main vector in the area with a CS positive rate of 7.6% and a human blood index of 78% in August 1989. Indoor resting densities were highest in the rural village, lowest in Labé and intermediate in the town of Timbi Madina (median density per room of 18, 2 and 4.5, respectively). The household survey showed different parasite rates in the three areas: 78.2% of children had parasitaemia in Sombili, 45.8% in Timbi Madina and 16.7% in the urban sector of Labé. Prevalence and levels of P. falciparum antisporozoite antibodies were lowest in the urban area (seroprevalence of 51.9% and median titre of 2.4 arbitrary units), intermediate in the town (70.1% and 5.2) and highest in the rural village (78.9% and 5.8). Serological findings produced by dispensary visits were similar to those obtained with the household surveys except in the rural area, while parasitological data obtained with the two sampling methods were different both in rural and in urban areas. Routine malaria diagnosis made on a presumptive basis in the health services in the survey period was able to detect 30.1% of cases with parasitaemia and 53.2% of cases with hyperparasitaemia (sensitivity) and malaria was correctly not suspected in 82.8% of non parasitized people (specificity).     

An Intact U5-Leader Stem Is Important for Efficient Replication of Simian Immunodeficiency Virus

Previous work has shown that four deletions in simian immunodeficiency virus (SIV), termed SD1a, SD1b, SD1c, and SD6, which eliminated sequences at nucleotide positions 322 to 362, 322 to 370, 322 to 379, and 371 to 379, respectively, located downstream of the primer binding site, impaired viral replication capacity to different extents. Long-term culturing of viruses containing the SD1a, SD1b, and SD6 deletions led to revertants that possessed wild-type replication kinetics. We now show that these revertants retained the original deletions in each case but that novel additional mutations were also present. These included a large deletion termed D1 (nt +216 to +237) within the U5 region that was shown to be biologically relevant to reversion of both the SD1a and SD1b constructs. In the case of SD6, two compensatory point mutations, i.e., A+369G, termed M1, located immediately upstream of the SD6 deletion, and C+201T, termed M2, within U5, were identified and could act either singly or in combination to restore viral replication. Secondary structure suggests that an intact U5-leader stem is important in SIV for infectiousness and that the additional mutants described played important roles in restoration of this motif.     

Abundance and distribution of bacterioplankton in the Gambia River,West Africa

Four ecological zones of the Gambia River were sampled during four different hydrologic seasons for determination of microbial, nutrient, and physical parameters. A Greco-Latin Square experimental design was used to define the particular transect, station, depth, and tide/time-of-day of samples taken. Ranges of total bacterioplankton densities (10⁶ cells/ml) were similar to those of tropical and temperate environments. Numbers of free bacteria were similar temporally, whereas attached bacteria numbers were greater during periods of high stream flows when suspended solids concentrations were higher. Free bacteria were usually twice as numerous in the freshwater zones than in the estuarine zones. Attached bacterial densities were approximately four times greater in the estuarine zones than in the freshwater zones. Uptake of³H-glucose on both a sample volume and per-cell basis increased from the early stages of the flood (6.95±SE 1.37 ng/liter/hour and 3.8 pg/hour/10⁶ cells, respectively) and reached observed annual maximums during the dry season (21.01±SE 3.05 ng/ liter/hour and 13.0 pg/hour/10⁶ cells, respectively). Spatially,³H-glucose uptake per sample volume and per cell was highest in the upper river zone and lowest in the lower estuary zone. The lower estuary zone consistently acted out of concert with the other river zones in terms of³H-glucose and¹⁴C-bicarbonate uptake. Analysis of variance (ANOVA) indicated that free and attached bacterioplankton densities were not homogeneous among transects, stations, depths, and tide/time-of-day at the different zones during the four hydrologic seasons. The results suggested that heterotrophy overshadowed autotrophy in the river and that the bacterial abundance, distribution, and glucose uptake activity in this tropical floodplain river were greatly influenced by the annual flood and the presence of extensive mangrove forests in the estuary.     

Retrospective study of maternal HIV-1 and HIV-2 infections and child survival in Abidjan,C?te d'Ivoire.

OBJECTIVES--To compare the effects of maternal HIV-1 and HIV-2 infections on outcome of pregnancy, infant mortality, and child survival, and to measure serological concordance between mothers and children. DESIGN--Retrospective cohort study with cross sectional study of concordance for HIV antibodies. SETTING--Hospital, tuberculosis clinic, and maternal and child health centre in Abidjan, Côte d''Ivoire, west Africa. SUBJECTS--986 women who had had a total of 2758 pregnancies since 1980. The last born children of 194 of these women. MAIN OUTCOME MEASURES--Pregnancy outcomes; mortality for all children born since 1980; and outcome for last born children. Serological concordance between mothers and last born children. RESULTS--Women with HIV-1 and HIV-2 infections had higher rates of spontaneous abortion and stillbirth than uninfected women (86/769 in HIV-1 positive women, 48/421 in HIV-2 positive, 31/234 in dually reactive, and 96/1131 in uninfected). Compared with children born to uninfected mothers (mortality 10.3%), greater proportions of children of HIV-1 positive (20.6%) and dually reactive (20.3%) mothers had died; mortality in children of HIV-2 infected women (13.1%) was not significantly increased. Infant mortalities for the last born children of HIV-1 positive, dually reactive, HIV-2 positive, and seronegative women were, respectively, 133, 82, 32, and 40 per 1000 live births. Nine of 77 last born children of HIV-1 positive mothers were concordantly seropositive compared with none of 21 children of HIV-2 infected mothers. CONCLUSIONS--Maternal HIV-2 infection has less influence on child survival than infection with HIV-1, probably because of a lower vertical transmission rate.     

Insecticide resistance status and mechanisms in Aedes aegypti populations from Senegal

Aedes aegypti is the main epidemic vector of arboviruses in Africa. In Senegal, control activities are mainly limited to mitigation of epidemics, with limited information available for Ae. aegypti populations. A better understanding of the current Ae. aegypti susceptibility status to various insecticides and relevant resistance mechanisms involved is needed for the implementation of effective vector control strategies. The present study focuses on the detection of insecticide resistance and reveals the related mechanisms in Ae. aegypti populations from Senegal.Bioassays were performed on Ae. aegypti adults from nine Senegalese localities (Matam, Louga, Barkedji, Ziguinchor, Mbour, Fatick, Dakar, Kédougou and Touba). Mosquitoes were exposed to four classes of insecticides using the standard WHO protocols. Resistance mechanisms were investigated by genotyping for pyrethroid target site resistance mutations (V1016G, V1016I, F1534C and S989P) and measuring gene expression levels of key detoxification genes (CYP6BB2, CYP9J26, CYP9J28, CYP9J32, CYP9M6, CCEae3a and GSTD4).All collected populations were resistant to DDT and carbamates except for the ones in Matam (Northern region). Resistance to permethrin was uniformly detected in mosquitoes from all areas. Except for Barkédji and Touba, all populations were characterized by a susceptibility to 0.75% Permethrin. Susceptibility to type II pyrethroids was detected only in the Southern regions (Kédougou and Ziguinchor). All mosquito populations were susceptible to 5% Malathion, but only Kédougou and Matam mosquitoes were susceptible to 0.8% Malathion. All populations were resistant to 0.05% Pirimiphos-methyl, whereas those from Louga, Mbour and Barkédji, also exhibited resistance to 1% Fenitrothion. None of the known target site pyrethroid resistance mutations was present in the mosquito samples included in the genotyping analysis (performed in > 1500 samples). In contrast, a remarkably high (20-70-fold) overexpression of major detoxification genes was observed, suggesting that insecticide resistance is mostly mediated through metabolic mechanisms. These data provide important evidence to support dengue vector control in Senegal.