Surveying soil faunal communities using a direct molecular approach

Soil faunal communities are often phylogenetically diverse and the accurate assessment of the taxonomic structure of these communities is both time-consuming and requires a high level of taxonomic expertise. Here we describe a DNA sequence-based methodology for characterizing soil micro- and mesofaunal communities that is similar to the molecular approaches commonly used to survey soil microbial diversity. The technique involves the direct extraction of faunal DNA from soil, PCR amplification of the extracted DNA with metazoan-specific primers, followed by the construction of clone libraries and direct sequencing of individual PCR products. We used this technique to characterize micro- and mesofaunal community composition from six individual soils representing two land-use types. The technique captured the more abundant faunal groups in the soils (nematodes, Collembola, Acari, tardigrades, enchytraeids) and provided sufficient taxonomic resolution to describe the overall structure of the communities. We compared the results obtained using this molecular approach to results obtained using a traditional, microscopy-based approach and found that the results were broadly similar. However, since biases are inherent in both methods it remains unclear which method provides a more accurate assessment of soil faunal community composition. Although this molecular approach has some distinct disadvantages over the more widely-used direct extraction methods, one advantage is that the taxonomic identification it can provide will be more accurate and consistent across research groups, facilitating effective comparisons of mesofaunal surveys.     

Modification of Marine Natural Product Ningalin B and SAR Study Lead to Potent P-Glycoprotein Inhibitors

In this study, new marine ningalin B analogues containing a piperazine or a benzoloxy group at ring C have been synthesized and evaluated on their P-gp modulating activity in human breast cancer and leukemia cell lines. Their structure-activity relationship was preliminarily studied. Compounds 19 and 20 are potent P-gp inhibitors. These two synthetic analogues of permethyl ningalin B may be potentially used as effective modulators of P-gp-mediated drug resistance in cancer cells.     

Detection of bovine viral diarrhea virus (BVDV) in seropositive cattle

Detection of bovine virus diarrhoea virus (BVDV) in one vaccinated beef cattle and three non-vaccinated dairy herds was investigated on peripheral blood leukocytes (PBL) with or without previous treatment followed by a capture ELISA (cELISA). Using the combination of PHA and polycation treatment, PBL from 229 seropositive cattle were studied and could be classified in four different states of BVDV infection. Lysed PBL from four animals were directly positive in cELISA (Category I), PBL of 17 animals were positive after PHA stimulation (Category II), 15 animals were positive only after PHA stimulation plus polycation treatment (Category III), while virus could not be detected in 193 seropositive cattle. Wild-type BVDV strains were isolated by co-culture on polycation-treated MDBK cells from 11 of these seropositive animals. BVDV antibodies of these same animals were able to neutralize their own virus, indicating that virus persists in PBL in spite of strain-specific antibodies. No apparent change of leukocyte subpopulations could be detected in any category of virus-positive animals. Thus, BVDV may be present in the PBL of some cattle, even in the presence of a specific active immune response.     

Oestradiol Induced Inhibition of Neuroendocrine Marker Expression in Leydig Cells of Adult Rats

The objectives of this work were to determine the changes in the expression of neuroendocrine markers in Leydig cell by oestradiol treatment, and to determine whether testosterone is able to recover partially the effects of hormonal suppression induced by oestradiol. Adult male rats were injected daily with either 50 microg of oestradiol or oestradiol plus testosterone propionate (25 mg every 3 days) for 15 days. The animals were sacrificed and testicles were dissected and processed by routine histological protocols. FSH and LH serum levels were determined by radioimmunoassay. The visualization of antigens was achieved by the streptavidin-peroxidase immunohistochemical method. Antibodies against chromogranin A (CrA), S-100 protein (S-100), P substance (PS), synaptofisin (SYN), neurofilament protein (NF), gliofibrillary acidic protein (GFAP) and neuron specific enolase (NSE) were used. The mean LH and FSH serum concentrations were consistently suppressed with hormonal treatments. Intermediate filaments (NF and GFAP) showed no difference in their expression. The expression of S-100, NSE and SYN was significantly lower in both hormone-treated groups. In oestradiol-treated rats, the immunoreactivity of CrA and SP decreased significantly but was restored after testosterone supplementation. Although the nature and functions of many of these substances in Leydig cells remain unknown, these results are consistent with the hypothesis that the expression of some neuroendocrine markers is hormonally controlled.     

Esterification and hydrolytic activities of Candida rugosa lipase isoform 1 (LIP1) immobilized on celite 545, duolite A7, and sephadex G-25

The esterification and hydrolytic activities of free and immobilized Candida rugosa lipase isoform 1 (LIP1) were investigated. Esterification activity was determined by reacting caprylic acid with glycerol in the presence of molecular sieves (30%, w/w), and the volume of 1.0 M NaOH consumed by the reaction products upon titration was used to calculate esterification activity. Caprylic acid was also reacted with cottonseed oil, and the amount of caprylic acid incorporated after 12 h of reaction was determined. Results indicated that LIP1 had little esterification activity, which was not significantly improved upon immobilization. Hydrolytic activity was determined by incubating tricaprylin emulsion (15%, w/w) with the respective lipases for 60 min, and the reaction products were titrated against 0.5 M NaOH. LIP1 showed hydrolytic activity comparable to Lipozyme RM IM. The hydrolytic activity improved significantly upon immobilization. Immobilization on Celite 545 produced the highest increase in hydrolytic activity.     

An evaluation of an oral formulation of moxidectin against selected anthelmintic-resistant and -susceptible strains of nematodes in lambs

The efficacy of an oral formulation of the newly developed parasiticide, moxidectin, was tested against benzimidazole-resistant Haemonchus contortus, Trichostrongylus colubriformis, and Nematodirus spathiger, levamisole-resistant Ostertagia circumcincta, and susceptible Cooperia curticei infections in weaned lambs. Thirty-two lambs were experimentally infected with mixed doses of the above strains of nematodes. They were allocated into four treatment groups by stratified randomisation using liveweights and faecal egg counts 28 days later. One group received moxidectin at 0.2 mg/kg liveweight, one group oxfendazole at 4.5 mg/kg liveweight, one group levamisole at 7.5 mg/kg liveweight and the last group remained untreated as the control. Worm burdens in the lambs at slaughter 10 days after oral treatment confirmed the resistance status of the nematode strains used, and showed that moxidectin had a greater than 99.9% efficacy (p<0.01) against all of them. No adverse effects due to treatment with moxidectin were observed in any of the animals.     

Modulation of broilers’ caecal microflora and metabolites in response to a potential probiotic Bacillus amyloliquefaciens

Studies have found that a dietary supplement of Bacillus amyloliquefaciens improved the growth performance, increased the nutrient digestibility of hosts and modulated the intestinal microflora. A total of 360 1‐day‐old Ross broilers were randomly divided into three treatments: a control group with a basal diet, an antibiotic group with a basal diet and added colistin sulphate, and a probiotics group with a basal diet and added Bacillus amyloliquefaciens. The HiSeq high‐throughput sequencing analysis of 16S rRNA was used to investigate the differences in birds’ caecal microflora, and metabolomics was used to analyse changes in caecal metabolites. Results showed that the supplementation of Bacillus amyloliquefaciens significantly improved the BW and ADG compared with the control birds. Results of sequencing indicated that (i) 645, 670, 596 unique operational taxonomic units (OTUs) were found in birds supplemented with Bacillus amyloliquefaciens on day 7, 21 and 42, separately, (ii) due to the diversity and relative abundance of the birds’ caecal microflora, the OTUs of the caecal microflora clustered according to age and treatment, except on day 42, (iii) among the six predominate families (Ruminococcaceae, Lachnospiraceae, Enterobacteriaceae, Erysipelotrichaceae, Lactobacillaceae and Rikenellaceae), the supplementation of Bacillus amyloliquefaciens significantly increased Enterobacteriaceae on day 42, (iv) Bacillus amyloliquefaciens increased the relative abundance of Faecalibacterium and Ruminococcus on day 21, increased the Faecalibacterium and Blautia and decreased the Ruminococcus on day 42. The metabolomics of caecal metabolites showed that the dietary Bacillus amyloliquefaciens changed the caecal metabolites involved of amino acid metabolism and glyceride metabolism, and the antibiotics changed the caecal metabolites that were related to carbohydrates and amino acid metabolism on day 21.     

Anticipating the species jump: surveillance for emerging viral threats

Zoonotic disease surveillance is typically triggered after animal pathogens have already infected humans. Are there ways to identify high‐risk viruses before they emerge in humans? If so, then how and where can identifications be made and by what methods? These were the fundamental questions driving a workshop to examine the future of predictive surveillance for viruses that might jump from animals to infect humans. Virologists, ecologists and computational biologists from academia, federal government and non‐governmental organizations discussed opportunities as well as obstacles to the prediction of species jumps using genetic and ecological data from viruses and their hosts, vectors and reservoirs. This workshop marked an important first step towards envisioning both scientific and organizational frameworks for this future capability. Canine parvoviruses as well as seasonal H3N2 and pandemic H1N1 influenza viruses are discussed as exemplars that suggest what to look for in anticipating species jumps. To answer the question of where to look, prospects for discovering emerging viruses among wildlife, bats, rodents, arthropod vectors and occupationally exposed humans are discussed. Finally, opportunities and obstacles are identified and accompanied by suggestions for how to look for species jumps. Taken together, these findings constitute the beginnings of a conceptual framework for achieving a virus surveillance capability that could predict future species jumps.