Predicting frequency distribution and influence of sociodemographic and behavioral risk factors of Schistosoma mansoni infection and analysis of co-infection with intestinal parasites.

Geospatial analysis was used to study the epidemiology of Schistosoma mansoni, intestinal parasites and co-infections in an area (Ilha das Flores) in Sergipe, Brazil. We collected individually georeferenced sociodemographic, behavioral and parasitological data from 500 subjects, analyzed them by conventional statistics, and produced risk maps by Kernel estimation. The prevalence rates found were: S. mansoni (24.0%), Trichuris trichiura (54.8%), Ascaris lumbricoides (49.2%), Hookworm (17.6%) and Entamoeba histolytica (7.0%). Only 59/500 (11.8%) individuals did not present any of these infections, whereas 279/500 (55.8%) were simultaneously infected by three or more parasites. We observed associations between S. mansoni infection and various variables such as male gender, being rice farmer or fisherman, low educational level, low income, water contact and drinking untreated water. The Kernel estimator indicated that high-risk areas coincide with the poorest regions of the villages as well as with the part of the villages without an adequate sewage system. We also noted associations between both A. lumbricoides and hookworm infections with low education and low income. A. lumbricoides infection and T. trichiura infection were both associated with drinking untreated water and residential open-air sewage. These findings call for an integrated approach to effectively control multiple parasitic infections.

Thirty years of viable but nonculturable state research: unsolved molecular mechanisms.

Viable but nonculturable (VBNC) cells were recognized 30 years ago; and despite decades of research on the topic, most results are disperse and apparently incongruous. Since its description, a huge controversy arose regarding the ecological significance of this state: is it a degradation process without real significance for bacterial life cycles or is it an adaptive strategy of bacteria to cope with stressful conditions? In order to solve the molecular mechanisms of VBNC state induction and resuscitation, researchers in the field must be aware and overcome common issues delaying research progress. In this review, we discuss the intrinsic characteristic features of VBNC cells, the first clues on what is behind the VBNC state's induction, the models proposed for their resuscitation and the current methods to prove not only that cells are in VBNC state but also that they are able to resuscitate.

Characterization of two resuscitation promoting factors of Listeria monocytogenes.

In actinobacteria, resuscitation promoting factor (Rpf) proteins have been described as having the ability to increase the viable count of dormant cultures and stimulate growth of vegetative cells through lag phase reduction. Recently, it was suggested that proteins Lmo0186 and Lmo2522 of Listeria monocytogenes are equivalent to Rpf proteins based on their genomic context and conserved domain architecture. It was proposed that they have evolved through non-orthologous displacement of the Rpf domain found in actinobacteria. Here we present biological and biochemical data supporting a function of Lmo0186 and Lmo2522 as Rpfs. These proteins are collectively dispensable for growth but a lmo0186 lmo2522 double mutant exhibits an extended lag phase when diluted in minimal medium. This phenotype could be partially complemented by medium supplementation with fM to nM concentrations of purified hexahistidine-tagged versions of Lmo0186 and Lmo2522, showing that these proteins can stimulate growth. Gel filtration analysis and cross-linking experiments suggest that the recombinant proteins in solution are elongated monomers. Both proteins display muralytic activity against crude cell wall preparations and are active against an artificial lysozyme substrate. Our study thus supports the hypothesis that Lmo0186 and Lmo2522 are functional equivalents of actinobacteria Rpf proteins and represents the first characterization of two Rpf homologues from firmicutes.

Sensibilização a aeroalérgenos em pacientes com suspeita de alergia respiratória atendidos na rede pública e privada no município de Aracaju / Sensitization to inhalant allergens in patients with suspected respiratory allergy treated at public and private health care clinics in the city of Aracaju

OBJETIVO: Identificar e comparar os padrões de sensibilização a alérgenos inalantes entre pacientes com suspeita de alergias respiratórias atendidos em ambulatórios públicos e privados do município de Aracaju.MÉTODOS: Foram analisados 1.514 resultados de testes cutâneos (TC) de punctura, realizados de abril de 2006 a setembro de 2009. Destes, 872 TC foram realizados em consultório de setor privado, e 642 nos ambulatórios do setor público. Os extratos alergênicos utilizados em TC foram: Dermatophagoides pteronyssinus, Dermatophagoides farinae, Blomia tropicalis, Periplaneta americana, Blattella germanica, penas, mistura de fungos e epitélios de cão e de gato. RESULTADOS: Asensibilização a pelo menos um alérgeno foi de 55% (58,4% no setor privado vs 50,3% no setor público; p < 0,05). Houve predomínio de TC positivos para ácaros domiciliares (total 53%; 57,3% vs. 47,2%; p < 0,0001), seguido de baratas (total 19,9%; 22,1% vs. 16,8%; p < 0,05) e animais domésticos (total 18,4%; 24,4% vs. 10,3%; p < 0,0001). Foram encontradas frequências significativamente maiores de TC positivos no setor privado quando comparado ao setor público para D. farinae (50,1% vs. 37,5%; p < 0,0001), D. pteronyssinus (47,7% vs. 35,7%; p < 0,0001), B. tropicalis (50,8% vs. 35,5%; p < 0,0001), epitélio de gato (18,1% vs. 6,2%; p < 0,0001), epitélio de cão (12,0% vs. 4,2%; p < 0,0001), penas (4,8% vs. 2,6%; p = 0,03) e B. germanica (16,1% vs. 11,5%; p = 0,01). Não se observou diferença significante quanto à sensibilização a fungos do ar e a P. americana.CONCLUSÃO: Ácaros domiciliares, baratas e animais domésticos foram as principais fontes de sensibilização alérgica na população estudada. Indivíduos atendidos no setor público apresentaram menor frequência de sensibilização a alérgenos inalantes quando comparados àqueles atendidos em clínica privada na cidade de Aracaju.

Role of bacteriophage SPP1 tail spike protein gp21 on host cell receptor binding and trigger of phage DNA ejection.

Bacteriophages recognize and bind specific receptors to infect suitable hosts. Bacteriophage SPP1 targets at least two receptors of the Bacillus subtilis cell envelope, the glucosylated wall teichoic acids and the membrane protein YueB. Here, we identify a key virion protein for YueB binding and for the trigger of DNA ejection. Extracts from B. subtilis-infected cells applied to a YueB affinity matrix led to preferential capturing of gp21 from SPP1. To assess the significance of this interaction, we isolated mutant phages specifically affected in YueB binding. The mutants exhibited a very low inactivation rate and a strong defect to eject DNA when challenged with YueB. The phenotype correlated with presence of a single amino acid substitution in the gp21 carboxyl terminus, defining a region involved in YueB binding. Immunoelectron microscopy located the gp21 N-terminus in the SPP1 cap and probably in the adjacent tail spike region whereas the gp21 C-terminus was mapped further down in the spike structure. Antibodies against this part of gp21 interfered with the interaction of YueB with SPP1 and triggered DNA ejection. The gp21 C-terminal region thus plays a central role in two early key events that commit the virus to deliver its genome into host cells.

Longevity in untreated congenital growth hormone deficiency due to a homozygous mutation in the GHRH receptor gene.

CONTEXT: Reduced longevity observed in hypopituitarism has been attributed to GH deficiency (GHD). It is, however, unclear whether GHD or other confounding factors cause this early mortality. OBJECTIVE: The aim was to study longevity in subjects from a large kindred with untreated, lifetime isolated GHD (IGHD) due to a homozygous mutation in the GHRH receptor gene and in heterozygous carriers of the mutation. DESIGN, SETTING, AND PARTICIPANTS: We carried out a retrospective cohort study on three groups. We first compared mortality risk of 65 IGHD individuals and their 128 unaffected siblings from 34 families. We then compared mean age of death of the IGHD to the general population. A transversal study was carried out to compare the rate of heterozygosity for the mutation in two groups of young (20-40 yr old) and old (60-80 yr old) normal-appearing subjects from the same county. MAIN OUTCOME MEASURE: We measured longevity. RESULTS: The risk of death of IGHD subjects was not different from their siblings. Life span in IGHD individuals was shorter than the general population. When stratified by sex, this difference persisted only in females, due to a high frequency of IGHD deaths in females aged 4-20. There was no significant difference in life span between IGHD subjects and siblings or the general population when analyzing subjects who reached age 20. The prevalence of heterozygosity did not differ in young and old groups, suggesting no survival advantage or disadvantage. CONCLUSIONS: In a selected genetic background, lifelong untreated IGHD does not affect longevity.

Update on seroprevalence of anti-Trypanosoma cruzi antibodies among blood donors in northeast Mexico.

Chagas disease has become frequent in non-endemic areas, where it can be transmitted by blood transmission. Therefore, we explored seroprevalence of anti-Trypanosoma cruzi antibodies among blood donors at the Cardiology Hospital, Mexican Institute of Social Security at Monterrey, Nuevo León, by both an enzyme-linked immunosorbent assay and indirect hemagglutination. Blood samples from 1,000 healthy blood donors were selected. A seropositivity of 2.8% was shown among the studied population, of which 2.59% (21/809) were inhabitants of Nuevo León, whereas 3.07% (2/65) and 3.96% (5/126) were from Coahuila and Tamaulipas, respectively. Our result is higher than that of a previous study from 1998, where a prevalence of 0.5% was reported. This once again corroborates the importance of installing a surveillance program to detect and prevent the transfusion of T. cruzi from asymptomatic blood donors in blood banks located in urban cities recognized as non-endemic.

A single amino acid substitution in one of the lipases of Aspergillus nidulans confers resistance to the antimycotic drug undecanoic acid.

A plausible approach to evaluate the inhibitory action of antifungals is through the investigation of the fungal resistance to these drugs. We describe here the molecular cloning and initial characterization of the A. nidulans lipA gene, where mutation (lipA1) conferred resistance to undecanoic acid, the most fungitoxic fatty acid in the C(7:0)-C(18:0) series. The lipA gene codes for a putative lipase with the sequence consensus GVSIS and WIFGGG as the catalytic signature. Comparison of the wild-type and LIP1 mutant strain nucleotide sequences showed a G --> A change in lipA1 allele, which results in a Glu(214) --> Lys substitution in LipA protein. This ionic charge change in a conserved LipA region, next to its catalytic site, may have altered the catalytic properties of this enzyme resulting in resistance to undecanoic acid.

Phage SPP1 reversible adsorption to Bacillus subtilis cell wall teichoic acids accelerates virus recognition of membrane receptor YueB.

Bacteriophage SPP1 targets the host cell membrane protein YueB to irreversibly adsorb and infect Bacillus subtilis. Interestingly, SPP1 still binds to the surface of yueB mutants, although in a completely reversible way. We evaluated here the relevance of a reversible step in SPP1 adsorption and identified the receptor(s) involved. We show that reversible adsorption is impaired in B. subtilis mutants defective in the glucosylation pathway of teichoic acids or displaying a modified chemical composition of these polymers. The results indicate that glucosylated poly(glycerolphosphate) cell wall teichoic acid is the major target for SPP1 reversible binding. Interaction with this polymer is characterized by a fast adsorption rate showing low-temperature dependence, followed by a rapid establishment of an equilibrium state between adsorbed and free phages. This equilibrium is basically determined by the rate of phage dissociation, which exhibits a strong dependence on temperature compatible with an Arrhenius law. This allowed us to determine an activation energy of 22.6 kcal/mol for phage release. Finally, we show that SPP1 reversible interaction strongly accelerates irreversible binding to YueB. Our results support a model in which fast SPP1 adsorption to and desorption from teichoic acids allows SPP1 to scan the bacterial surface for rapid YueB recognition.

The lactococcal abortive infection protein AbiP is membrane-anchored and binds nucleic acids.

AbiP, a lactococcal abortive phage infection system, has previously been shown to arrest phage bIL66M1 DNA replication around 10 min after infection and to inhibit the switch off of phage early transcripts. We report here the functional characterization and implication in the abortive infection phenotype of two domains identified in the AbiP sequence. We show that AbiP is a protein anchored to the membrane by an N-terminal membrane-spanning domain. Our results further suggest that membrane localization may be required for the anti-phage activity of AbiP. The remainder of the protein, which contains a putative nucleic acid binding domain, is shown to be located on the cytosolic side. Purified AbiP is shown to bind nucleic acids with an approximately 10-fold preference for RNA relative to ssDNA. AbiP interaction with both ssDNA and RNA molecules occurs in a sequence-independent manner. We have analyzed the effect of substitutions of aromatic and basic residues on the surface of the putative binding fold. In vitro and in vivo studies of these AbiP derivatives indicate that the previously reported effects on phage development might be dependent on the nucleic acid binding activity displayed by the membrane-bound protein.

Pressure built by DNA packing inside virions: enough to drive DNA ejection in vitro, largely insufficient for delivery into the bacterial cytoplasm.

Tailed bacteriophage particles carry DNA highly pressurized inside the capsid. Challenge with their receptor promotes release of viral DNA. We show that addition of the osmolyte polyethylene glycol (PEG) has two distinct effects in bacteriophage SPP1 DNA ejection. One effect is to inhibit the trigger for DNA ejection. The other effect is to exert an osmotic pressure that controls the extent of DNA released in phages that initiate ejection. We carried out independent measurements of each effect, which is an essential requirement for their quantitative study. The fraction of phages that do not eject increased linearly with the external osmotic pressure. In the remaining phage particles ejection stopped after a defined amount of DNA was reached inside the capsid. Direct measurement of the size of non-ejected DNA by gel electrophoresis at different PEG concentrations in the latter sub-population allowed determination of the external osmotic pressure that balances the force powering DNA exit (47 atm for SPP1 wild-type). DNA exit stops when the ejection force mainly due to repulsion between DNA strands inside the SPP1 capsid equalizes the force resisting DNA insertion into the PEG solution. Considering the turgor pressure in the Bacillus subtilis cytoplasm the energy stored in the tight phage DNA packing is only sufficient to power entry of the first 17% of the SPP1 chromosome into the cell, the remaining 83% requiring application of additional force for internalization.

The minor capsid protein gp7 of bacteriophage SPP1 is required for efficient infection of Bacillus subtilis.

Gp7 is a minor capsid protein of the Bacillus subtilis bacteriophage SPP1. Homologous proteins are found in numerous phages but their function remained unknown. Deletion of gene 7 from the SPP1 genome yielded a mutant phage (SPP1del7) with reduced burst-size. SPP1del7 infections led to normal assembly of virus particles whose morphology, DNA and protein composition was undistinguishable from wild-type virions. However, only approximately 25% of the viral particles that lack gp7 were infectious. SPP1del7 particles caused a reduced depolarization of the B. subtilis membrane in infection assays suggesting a defect in virus genome traffic to the host cell. A higher number of SPP1del7 DNA ejection events led to abortive release of DNA to the culture medium when compared with wild-type infections. DNA ejection in vitro showed that no detectable gp7 is co-ejected with the SPP1 genome and that its presence in the virion correlated with anchoring of released DNA to the phage particle. The release of DNA from wild-type phages was slower than that from SPP1del7 suggesting that gp7 controls DNA exit from the virion. This feature is proposed to play a central role in supporting correct routing of the phage genome from the virion to the cell cytoplasm.

Leuconostoc pseudoficulneum sp. nov., isolated from a ripe fig.

Six strains of lactic acid bacteria (LAB) were isolated from a ripe fig. These strains constituted a highly homogeneous, but distinct, cluster that was separate from other LAB species in a polyphasic approach including dot-blot DNA-DNA hybridization, SDS-PAGE whole-cell protein profiling, carbohydrate fermentation ability, growth characteristics, enzymic profiling, pulsed-field gel electrophoresis macrorestriction analysis and RFLPs. Phylogenetic analysis based on 16S rRNA gene sequencing positioned a representative strain, LC51(T), in a distinct line of descent within the recently described clade comprising Leuconostoc ficulneum, Leuconostoc fructosum and Leuconostoc durionis; L. ficulneum was its closest neighbour (98 % sequence similarity). DNA-DNA hybridization values and chemotaxonomic and biochemical characteristics, including enzymic profiles detected with API ZYM microtubes, confirmed that this group of strains is distinct from L. ficulneum and represents a novel species within the genus Leuconostoc. Taking into account the common origin and phylogenetic proximity, the name Leuconostoc pseudoficulneum sp. nov. is proposed. Strain LC51(T) (=DSM 15468(T) = CECT 5759(T)) is the type strain; the DNA G + C content of this strain is 44.5 mol%.

Expression of the anaphylatoxin receptors C3aR and C5aR is increased in fatal asthma.

BACKGROUND: The mechanisms leading to death from asthma are not completely understood. Recent studies suggest the involvement of the anaphylatoxins C3a and C5a, generated during complement activation, and their receptors C3aR and C5aR in the pathogenesis of asthma. OBJECTIVE: The aim of our study was to investigate the expression of C3aR and C5aR in fatal asthma. METHODS: We analyzed lung tissue from 14 subjects who died of asthma (fatal asthma; FA) and 14 subjects who died of nonpulmonary causes (controls) and bronchial biopsy specimens from 16 subjects with mild intermittent asthma (MIA). C3aR and C5aR expression was evaluated by immunohistochemistry, and a semiquantitative analysis of the intensity of staining was performed according to a visual analogue scale (score, 0-3). RESULTS: C3aR was expressed on airway epithelium, smooth muscle, submucosal, and parenchymal vessels. C5aR was expressed on myeloid cells infiltrating the submucosa and on airway epithelium. Statistical analysis demonstrated higher expression of C3aR on submucosal vessels in FA compared with controls and MIA (median [minimum-maximum], controls, 0.24 [0-1.48]; MIA, 0.0 [0-1.00]; FA, 1.56 [0.13-3]; P = .002). C3aR was also increased on parenchymal vessels in FA (controls, 0.56 [0-2.00]; FA, 1.81 [0.5-3]; P = .0004). C5aR expression on airway epithelium was increased in FA compared with controls and MIA (controls, 1.25 [0.25-3]; MIA, 1.00 [0-2.00]; FA, 3.00 [1.13-3.00]; P = .001). CONCLUSION: The results of our study suggest a role of complement in FA.

Bacillus subtilis operon encoding a membrane receptor for bacteriophage SPP1.

The results reported here have identified yueB as the essential gene involved in irreversible binding of bacteriophage SPP1 to Bacillus subtilis. First, a deletion in an SPP1-resistant (pha-2) strain, covering most of the yueB gene, could be complemented by a xylose-inducible copy of yueB inserted at amyE. Second, disruption of yueB by insertion of a pMutin4 derivative resulted in a phage resistance phenotype regardless of the presence or absence of IPTG (isopropyl-beta-D-thiogalactopyranoside). YueB homologues are widely distributed in gram-positive bacteria. The protein Pip, which also serves as a phage receptor in Lactococcus lactis, belongs to the same family. yueB encodes a membrane protein of approximately 120 kDa, detected in immunoblots together with smaller forms that may be processed products arising from cleavage of its long extracellular domain. Insertional inactivation of yueB and the surrounding genes indicated that yueB is part of an operon which includes at least the upstream genes yukE, yukD, yukC, and yukBA. Disruption of each of the genes in the operon allowed efficient irreversible adsorption, provided that yueB expression was retained. Under these conditions, however, smaller plaques were produced, a phenotype which was particularly noticeable in yukE mutant strains. Interestingly, such reduction in plaque size was not correlated with a decreased adsorption rate. Overall, these results provide the first demonstration of a membrane-bound protein acting as a phage receptor in B. subtilis and suggest an additional involvement of the yukE operon in a step subsequent to irreversible adsorption.

Abnormal alveolar attachments with decreased elastic fiber content in distal lung in fatal asthma.

Small airway disease is thought to contribute significantly to functional impairment caused by asthma. Functional evidence of airway-parenchyma uncoupling in asthma, such as loss of deep breath bronchodilator effect in bronchoconstrictive episodes and enhanced airway closure, has been previously demonstrated. Elastic fibers are essential to maintain adequate elastic recoil of the lungs. In this study, we hypothesized that alveolar attachments could be abnormal and that elastic fibers could be damaged in the distal lungs of patients with fatal asthma. For this purpose, we measured the number of abnormal alveolar attachments and quantified the content of elastic fibers in the adventitial layer of small airways and in the peribronchial and distal alveolar septa of 15 patients who died of asthma (FA) and 9 control subjects (CTRL). Our data (geometric mean [range]) showed an increased proportion of abnormal alveolar attachments per centimeter of basement membrane perimeter in fatal asthma (FA, 0.18 [0.03-4.00]; CTRL, 0.00 [0.00-0.12]; p < 0.001) and decreased elastic fiber content in the small airway adventitial layer (FA, 4.08 [2.22-11.46] microm; CTRL, 6.79 [5.62-10.0] microm; p = 0.01) and in the peribronchial alveoli (FA, 1.08 [0.46-1.91] microm; CTRL, 1.81 [1.22-1.74] microm; p = 0.003), but not in the distal alveoli. We propose that structural alterations at the peribronchiolar level might contribute to the pathogenesis of some functional abnormalities observed in patients with severe asthma.

The high-resolution functional map of bacteriophage SPP1 portal protein.

An essential component in the assembly of nucleocapsids of tailed bacteriophages and of herpes viruses is the portal protein that is located at the unique vertex of the icosahedral capsid through which DNA movements occur. A library of mutations in the bacteriophage SPP1 portal protein (gp6) was generated by random mutagenesis of gene 6. Screening of the library allowed identification of 67 single amino acid substitutions that impair portal protein function. Most of the mutations cluster within stretches of a few amino acids in the gp6 carboxyl-terminus. The mutations were divided into five classes according to the step of virus assembly that they impair: (1) production of stable gp6; (2) interaction of gp6 with the minor capsid protein gp7; (3) incorporation of gp6 in the procapsid structure; (4) DNA packaging; and (5) sizing of the packaged DNA molecule. Most of the mutations fell in classes 3 and 4. This is the first high-resolution functional map of a portal protein, in which its function at different steps of viral assembly can be directly correlated with specific regions of its sequence. The work provides a framework for the understanding of central processes in the assembly of viruses that use specialized portals to govern entry and exit of DNA from the viral capsid.

Physical map of the genome of Oenococcus oeni PSU-1 and localization of genetic markers.

A physical map of the chromosome of Oenococcus oeni PSU-1 was constructed. This represents the first map for a strain of this species. A total of 37 restriction sites for the rare-cutting endonucleases Ascl, Fsel, Notl and Sfil were mapped on the chromosome, which was found to be circular with an estimated size of 1857 kb. Fragment order was determined using several approaches: analysis of partial and double digestions, two-dimensional pulsed-field gel electrophoresis, isolation of linking clones, and Southern hybridization with labelled restriction fragments both from PSU-1 and from O. oeni strain GM. Oenococcal genes alsS/alsD, mleA and mir, two phage attachment sites and recurrent sequences such as IS1165-like elements and rrn loci were located on the physical map. Specific fragments hybridizing with gene probes from Lactococcus lactis, Leuconostoc mesenteroides and Bacillus subtilis were also identified. The two ribosomal operons have been precisely located and their transcription direction determined.