Examining serological diversity of “cowpea” rhizobia by the ELISA technique

To appraise the usefulness of the enzyme-linked immunosorbent assay (ELISA) technique for examining the serological diversity of slow-growing rhizobia, twelve diverse strains from three countries were examined with four antisera. Eleven of the strains were from the cowpea miscellany, and the twelfth was a Rhizobium japonicum strain. Some cowpea strains showed no antigenic relatedness with each other while others were closely related, and some showed a greater affinity with the R. japonicum strain than with other cowpea strains. All of the strains showed antigenic homology to an isolate from a wild Arachis sp., while two strains isolated from adjacent plants of the same cultivar had little homology. These patterns ofrelatedness and diversity clearly demonstrated the utility of the ELISA method, and so it was used to examine 53 strains isolated from cowpeas grown at three West African locations, Maradi (Niger), Ibadan and Onne (Nigeria). Broad ranges of serological diversity were found in the rhizobia at each location, moreover each population had its own general characteristics. Maradi strains were highly reactive with the five antisera used, Onne strains less so, and Ibadan strains even less so. ELISA reactivity correlated with colony morphology but not with nodulation potential.     

Cowpea Rhizobia Producing Dark Nodules: Use in Competition Studies

During a program of screening rhizobia from West Africa, it was found that some strains produced nodules of unusually dark appearance on cowpeas, but not on peanuts, soybeans, pigeon peas, or mung beans. The dark pigmentation was in the bacteroid zone, was not correlated with nodule effectiveness, and was additional to the leghemoglobin pigment. Only rhizobial strains with a nongummy (“dry”) colony morphology produced dark nodules. Visually distinguishable pink and dark nodules formed on the same root when a mixture of pink and dark strains was applied as inoculum. The dark-nodule phenotype was therefore appraised as a marker and found to be useful for studying nodulation competition with strains of the orthodox pink-nodule type. The competitiveness of 10 pink-nodule strains was examined relative to a black-nodule strain, IRc 256; a range of competitiveness was obtained of less competitive than, equally competitive to, or more competitive than IRc 256. Patterns of primary (early) nodulation were generally the same as patterns of secondary (later) nodulation. Mixed infections by dark and pink strains produced piebald nodules, the frequency of occurrence of which was much greater among primary than among secondary nodules.     

Effect of inoculation of lactic acid bacteria on proteolytic activity of psychrotrophic Gram-negative bacteria in fresh farmed sea bass (Dicentrarchus labrax) fillets during storage at 4 °C under vacuum-packed conditions

The effect of two strains of lactic acid bacteria (LAB) (Lactococcus lactis and Carnobacterium piscicola) on the proteolytic activity of four strains of Psychrotrophic Gram-negative bacteria [Psy G(?)] (Pseudomonas fluorescens, Aeromonas hydrophila, Pseudomonas putida and Photobacterium damselae) has been determined using sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE), in fresh vacuum-packed farmed sea bass (Dicentrarchus labrax) fillets artificially contaminated, during 21 days of chilled storage. The profiles of sarcoplasmic (SP) and myofibrillar (MP) proteins indicated that the major changes were produced with Pseudomonas fluorescens, Aeromonas hydrophila, and Pseudomonas putida starters. The results also showed that LAB strains presented a weak proteolytic activity against MP and SP proteins in muscle of fresh sea bass. In fact, we noted the less pronounced degradation of protein fractions in samples inoculated with LAB combination. Moreover, a significant bacteriostatic effect of LAB strains was demonstrated against all microflora, particularly mesophilic aerobic plate counts (MAPC) and psychrotrophic bacterial counts (PBC), with fillets remaining unspoiled until the end of storage, against values of 7 and 8 log CFU/g, respectively; control fish fillets exceeded the upper acceptability limit.     

Enhancement of bacterial efficiency for metal removal using mutation techniques

The objective of the present study was to obtain by mutation and selection techniques bacterial strains capable of removing heavy metals at high efficiency. Four of the bacteria most promising in metal uptake, Staphylococcus aureus, Bacillus Sphaericus, B. licheniformis and Arthrobacter sp. were selected after isolation from water heavily polluted with heavy metals. Two mutagenic agents were used: U.V. irradiation at 245nm (physical) and 1% ethidium bromide (chemical). Optimum conditions for metal removal by most of the tested bacteria were: pH 9, 50°C and 200rev/min agitation speed. Induction of mutation both physically or chemically resulted in mutants that were superior over their wild types in removing heavy metals under investigation. The highest removal efficiencies (REs) achieved were in the following order: Cd(89.9–100%); Cr(87.3–99.7%); Zn(47.7–100%); Cu(40.8–84.7%); Pb(40.2–51%); Fe(17.5–28.7%); Ni(13.8–23.9%) and finally Co(17.2–18.4%). Using mixed cultures of the wild and the selected mutants enhanced the RE(s) of some metals compared to those obtained by individual species, and the time required to achieve the highest RE was reduced.     

Widespread Expression of Erythropoietin Receptor in Brain and Its Induction by Injury

Erythropoietin (EPO) exerts potent neuroprotective, neuroregenerative and procognitive functions. However, unequivocal demonstration of erythropoietin receptor (EPOR) expression in brain cells has remained difficult since previously available anti-EPOR antibodies (EPOR-AB) were unspecific. We report here a new, highly specific, polyclonal rabbit EPOR-AB directed against different epitopes in the cytoplasmic tail of human and murine EPOR and its characterization by mass spectrometric analysis of immuno-precipitated endogenous EPOR, Western blotting, immunostaining and flow cytometry. Among others, we applied genetic strategies including overexpression, Lentivirus-mediated conditional knockout of EpoR and tagged proteins, both on cultured cells and tissue sections, as well as intracortical implantation of EPOR-transduced cells to verify specificity. We show examples of EPOR expression in neurons, oligodendroglia, astrocytes and microglia. Employing this new EPOR-AB with double-labeling strategies, we demonstrate membrane expression of EPOR as well as its localization in intracellular compartments such as the Golgi apparatus. Moreover, we show injury-induced expression of EPOR. In mice, a stereotactically applied stab wound to the motor cortex leads to distinct EpoR expression by reactive GFAP-expressing cells in the lesion vicinity. In a patient suffering from epilepsy, neurons and oligodendrocytes of the hippocampus strongly express EPOR. To conclude, this new analytical tool will allow neuroscientists to pinpoint EPOR expression in cells of the nervous system and to better understand its role in healthy conditions, including brain development, as well as under pathological circumstances, such as upregulation upon distress and injury.     

Changes in physical,chemical and microbial parameters during the composting of municipal sewage sludge

Changes in physical, chemical and microbial parameters were investigated during the composting of municipal sewage sludge. Raw sewage sludge (30% dry matter) was mixed with compost from sewage sludge (85% dry matter) in 3:1 ratio (v/v). The mixture was divided into 4 windrows which were composted under the same conditions except the turning factor. The turning was every 7, 10, 15 days and according to the temperature which must be (55–65°C) for windrow 1 (W1), windrow 2 (W2), windrow 3 (W3) and windrow 4 (W4), respectively. Water was added to adjust the moisture content (40–60%). The composting process consisted of 2 periods; fermentation (12 weeks) and maturation (4 weeks). The results showed that the temperature reached the maximum after 12 weeks for W1 and 11 weeks for W2, W3 and W4 and then decreased. The final compost was nearly odourless and black, especially in case of W4. The general trend indicates a decrease in organic matter, organic carbon and nitrogen (N), whereas ash, potassium (K) and phosphorus (P) increased and consequently C/K and C/P ratios decreased. There was a slight increase in C/N ratio. The pH increased and then decreased to near neutrality at the end. The mesophilic bacteria increased during the fermentation period and decreased after that, whereas the thermophilic ones increased with increasing of temperature, decreased after 2 weeks and increased again during the fermentation period and then decreased. The mesophilic and thermophilic fungi were present during the first week and disappeared after that. The final compost was pathogens-free as indicated by the counts of coliforms and Salmonella.     

The Combined Deletion of S6K1 and Akt2 Deteriorates Glycemic Control in a High-Fat Diet

Signaling downstream of mechanistic target of rapamycin complexes 1 and 2 (mTORC1 and mTORC2) controls specific and distinct aspects of insulin action and nutrient homeostasis in an interconnected and as yet unclear way. Mice lacking the mTORC1 substrate S6 kinase 1 (S6K1) maintain proper glycemic control with a high-fat diet. This phenotype is accompanied by insulin hypersensitivity, Akt- and AMP-activated kinase upregulation, and increased lipolysis in adipose tissue and skeletal muscle. Here, we show that, when S6K1 inactivation is combined with the deletion of the mTORC2 substrate Akt2, glucose homeostasis is compromised due to defects in both insulin action and β-cell function. After a high-fat diet, the S6K1(-/-) Akt2(-/-) double-mutant mice do not become obese, though they are severely hyperglycemic. Our data demonstrate that S6K1 is required for pancreatic β-cell growth and function during adaptation to insulin resistance states. Strikingly, the inactivation of two targets of mTOR and phosphatidylinositol 3-kinase signaling is sufficient to reproduce major hallmarks of type 2 diabetes.     

A Natural Variant of Obestatin,Q90L,Inhibits Ghrelin's Action on Food Intake and GH Secretion and Targets NPY and GHRH Neurons in Mice

Background

Ghrelin and obestatin are two gut-derived peptides originating from the same ghrelin/obestatin prepropeptide gene (GHRL). While ghrelin stimulates growth hormone (GH) secretion and food intake and inhibits γ-aminobutyric-acid synaptic transmission onto GHRH (Growth Hormone Releasing Hormone) neurons, obestatin blocks these effects. In Humans, GHRL gene polymorphisms have been associated with pathologies linked to an unbalanced energy homeostasis. We hypothesized that one polymorphism located in the obestatin sequence (Q to L substitution in position 90 of the ghrelin/obestatin prepropeptide, rs4684677) may impact on the function of obestatin. In the present study, we tested the activity of native and Q90L obestatin to modulate ghrelin-induced food intake, GH secretion, cFos activity in GHRH and Neuropeptide Y (NPY) neurons and γ-aminobutyric-acid activity onto GHRH neurons.

Methodology/Principal findings

Food intake, GH secretion and electrophysiological recordings were assessed in C57BL/6 mice. cFos activity was measured in NPY-Renilla-GFP and GHRH-eGFP mice. Mice received saline, ghrelin or ghrelin combined to native or Q90L obestatin (30 nmol each) in the early light phase. Ghrelin stimulation of food intake and GH secretion varied considerably among individual mice with 59–77% eliciting a robust response. In these high-responders, ghrelin-induced food intake and GH secretion were reduced equally by native and Q90L obestatin. In contrast to in vivo observations, Q90L was slightly more efficient than native obestatin in inhibiting ghrelin-induced cFos activation within the hypothalamic arcuate nucleus and the nucleus tractus solitarius of the brainstem. After ghrelin injection, 26% of NPY neurons in the arcuate nucleus expressed cFos protein and this number was significantly reduced by co-administration of Q90L obestatin. Q90L was also more potent that native obestatin in reducing ghrelin-induced inhibition of γ-aminobutyric-acid synaptic transmission onto GHRH neurons.

Conclusions/Significance

These data support the hypothesis that Q90L obestatin partially blocks ghrelin-induced food intake and GH secretion by acting through NPY and GHRH neurons.     

Keratinophilic fungi on the hair of goats from the West Bank of Jordan

The mycoflora of the hair in 178 goats from the West Bank of Jordan was analysed and the frequency of occurrence and the relative importance value for the different keratinophilic fungi found were calculated. One hundred and seven species which belong to 38 genera were isolated. Thirty six of these species were either well recognised agents of mycoses (Trichophyton mentagrophytes, T. verrucosum, and M. nanum), or have been frequently isolated from human and animal lesions (Arthroderma spp., Acremonium kiliense, Alternaria alternata, Aspergillus flavus, Cladosporium carionii, and several other species). These potentially pathogenic fungal species comprised 66.9% of all keratinophilic fungi found on the hair of goats. The role of this animal as a reservoir for dermatophytes and other potentially pathogenic fungi is discussed.     

Vermifiltration as a stage in reuse of swine wastewater: Monitoring methodology on an experimental farm

Vermifiltration is a new technology using earthworms to process organically polluted water. A pilot associated with a swine facility (piggery) with 66 swine was developed to treat diluted manure, produce earthworms and vermicompost, and reduce air pollution. The aim of the experiment reported here was to devise an integrated method – biological, chemical and physical – for further research and development of vermifiltration in diluted swine manure, and provide some preliminary results. The earthworm population increased by 30% in 4 weeks, indicating the acclimation of the earthworms. A reduction in ammonia emission was observed of about 50% for the whole system. Higher water (+100%), carbon (+70%), and total nitrogen (+80%) gaseous losses were observed compared to conventional breeding on a slatted floor. This methodology can be used for further studies to develop vermifiltration for earthworm and vermicompost production from diluted animal manure, without pollution transfer.