Molecular characterization of Corynebacterium pseudotuberculosis isolates using ERIC-PCR

Caseous lymphadenitis is an infectious sheep and goats disease caused by Corynebacterium pseudotuberculosis and characterized by abscesses in superficial and visceral lymph nodes. C. pseudotuberculosis strains isolated from these hosts have been shown to be very difficult to type by the existing methods. The aim of this study is evaluating the potential of the Enterobacterial Repetitive Intergenic Consensus (ERIC-PCR) as a tool for molecular typing of C. pseudotuberculosis strains isolated in sheep. One hundred and twenty seven isolates of C. pseudotuberculosis were isolated from lesions suspected to have had caseous lymphadenitis collected from sheep at the slaughterhouse. Animals were from 24 flocks in 13 municipalities of the Minas Gerais State, Brazil. Species identification of the isolates was performed by routine biochemical tests and mPCR. Fingerprint was performed by RAPD using ERIC-1R, ERIC-2 and ERIC-1R+ERIC-2 primers. Seventeen different genotypes were generated by ERIC 1-PCR, 21 genotypes by ERIC 2-PCR and 21 genotypes by ERIC 1+2-PCR. Hunter-Gaston Discrimination Index (HGDI) found for ERIC 1, ERIC 2, ERIC 1+2 PCR were 0.69, 0.87, and 0.84, respectively. For most herds evaluated observed at most three different genotypes among isolates from animals of these property, in all ERIC-PCR assays. However a few flocks observed between four and nine genotypes per flock. The W Kendall value found for correlation among the three techniques of ERIC-PCR was 0.91 (P<5.0 x 10(-6)). The results show that ERIC-PCR has good discriminatory power and advantages over other DNA-based typing methods, making it a useful tool to discriminate C. pseudotuberculosis isolates.     

Archaeal communities in the sediments of three contrasting mangroves

Purpose

This study evaluates the presence and diversity of 16S rRNA (rrs) and amoA genes from archaea in three mangrove sediments under different stages of preservation (one pristine mangrove, one affected by anthropogenic activity, and another contaminated by an oil spill) in the state of São Paulo (Brazil).

Materials and methods

A combination of DGGE, coupled with ordination analysis, and clone libraries of both targeted genes (rrs and amoA) was used to infer the diversity and phylogeny of archaeal communities in the mangrove analyzed samples.

Results and discussion

The DGGE combined with multivariate analysis revealed, based on the ribosomal gene, differences in archaeal communities according to environmental parameters such as mangrove location, anthropogenic activity, and oil contamination. The profiles based on the amoA gene were more similar than those obtained for the gene rrs, with the only difference statistically observed for the community found in the mangrove under anthropogenic pressure. Furthermore, phylogenetic analysis revealed most archaeal groups affiliated to sequences belonging to the Thaumarchaeota (53.1%, 24 OTUs) and Euryarchaeota (29.6%, 14 OTUs) phyla, in addition to 14 sequences affiliated to an unclassified Archaea (16.1%, 8 OTUs). Concerning the analysis of the amoA gene, mangroves harbored sequences affiliated with those previously described in water column and soil/sediment samples, besides two possible clusters specifically found in mangroves.

Conclusions

The findings are that the mangroves act as a reservoir for archaeal diversity, are possibly involved in nitrogen transformation in this ecosystem, and are affected by distinct pressures caused by anthropogenic activities.

     

Effect of diet and ration on the relationship between plasma GH and IGF-1 concentrations in Arctic charr, Salvelinus alpinus (L.)

The purpose of the study was to investigate whether dietary ration or diet composition influence the relationship between plasma growth hormone (GH) and insulin‐like growth factor‐1 (IGF‐1) in Arctic charr (Salvelinus alpinus L.). The pattern of changes in plasma GH and IGF‐1 concentrations was examined in fish fed at different ration levels (0%, 0.35% and 0.70% BW day⁻¹) for 5 weeks, and in fish fed diets containing different lipid:crude protein (LCP) ratios. Ration level significantly affected plasma GH and IGF‐1 concentrations; at 5 weeks the levels of both hormones in the food‐deprived group were significantly lower than in fish fed the 0.70% BW day⁻¹ ration. Also, plasma IGF‐1 levels in fish of each ration treatment group were significantly correlated with individual final body weight; no such correlation was found for GH. To examine the effects of dietary LCP ratios, fish were fed for up to 18 weeks, with one of four formulated diets that had LCP ratios (dry matter basis) of 0.35 (Diet 1), 0.43 (Diet 2), 0.51 (Diet 3) or 0.59 (Diet 4), or a commercial diet (Diet 5) which had an LCP ratio of 0.38. Statistical differences in plasma GH and IGF‐1 concentrations were found only after 18 weeks. Growth hormone was significantly lower in fish fed Diets 1 and 2 compared with Diets 3 and 5, and IGF‐1 was significantly lower in fish fed Diet 1 compared with Diets 2 and 5. Significant correlations between plasma GH and IGF‐1 concentrations were found only for fish fed Diets 1 and 5, suggesting that the influence of diet composition on the relationship between GH and IGF‐1 varies with the dietary LCP ratio in this species. The decline in plasma IGF‐1 concentrations during food deprivation is similar to that described in other species; however, the unexpected decrease in plasma GH during food deprivation in this study may represent a species‐specific response.     

Bioavailability and Tolerability of Topical and Oral Diclofenac Sodium Administration in Healthy Ponies

Aiming to characterize the bioavailability and assess the safety of topical and oral treatment of diclofenac sodium in healthy ponies, 18 animals were divided in three groups: one treated with topical (group I, n = 6), the second with oral diclofenac (group II, n = 6) at 2.5 mg/kg for 3 days, and the third group received 2.2 mg/kg oral phenylbutazone (group III, n = 6) also for 3 days to serve as positive control. To evaluate bioavailability, blood samples were collected before and at 0, 0.5, 1, 3, 6, 9, 12, 24, 36, 48, 60, 72, 96, 120, and 144 hours after starting treatment. To evaluate diclofenac sodium concentration in the synovial fluid, samples of six ponies (group I, n = 3; group II, n = 3) were collected at 6, 12, and 24 hours after starting the treatment.     

New Invertebrate Vectors for PST,Spirolides and Okadaic Acid in the North Atlantic

The prevalence of poisoning events due to harmful algal blooms (HABs) has declined during the last two decades through monitoring programs and legislation, implemented mainly for bivalves. However, new toxin vectors and emergent toxins pose a challenge to public health. Several locations on the Portuguese coast were surveyed between 2009 and 2010 for three distinct biotoxin groups [saxitoxin (PST), spirolide (SPX) and okadaic acid (OA)], in 14 benthic species of mollusks and echinoderms. Our main goals were to detect new vectors and unravel the seasonal and geographical patterns of these toxins. PSTs were analyzed by the Lawrence method, SPXs by LC-MS/MS, and OA by LC-MS/MS and UPLC-MS/MS. We report 16 new vectors for these toxins in the North Atlantic. There were differences in toxin contents among species, but no significant geographical or seasonal patterns were found. Our results suggest that legislation should be adjusted to extend the monitoring of marine toxins to a wider range of species besides edible bivalves.     

Growth and Saxitoxin Production by Cylindrospermopsis raciborskii (Cyanobacteria) Correlate with Water Hardness

The cosmopolitan and increasing distribution of Cylindrospermopsis raciborskii can be attributed to its ecophysiological plasticity and tolerance to changing environmental factors in water bodies. In reservoirs in the semi-arid region of Brazil, the presence and common dominance of C. raciborskii have been described in waters that are considered hard. We investigated the response of a Brazilian C. raciborskii strain to water hardness by evaluating its growth and saxitoxin production. Based on environmental data, a concentration of 5 mM of different carbonate salts was tested. These conditions affected growth either positively (MgCO₃) or negatively (CaCO₃ and Na₂CO₃). As a control for the addition of cations, MgCl₂, CaCl₂ and NaCl were tested at 5 or 10 mM, and MgCl₂ stimulated growth, NaCl slowed but sustained growth, and CaCl₂ inhibited growth. Most of the tested treatments increased the saxitoxin (STX) cell quota after six days of exposure. After 12 days, STX production returned to concentrations similar to that of the control, indicating an adaptation to the altered water conditions. In the short term, cell exposure to most of the tested conditions favored STX production over neoSTX production. These results support the noted plasticity of C. raciborskii and highlight its potential to thrive in hard waters. Additionally, the observed relationship between saxitoxin production and water ion concentrations characteristic of the natural environments can be important for understanding toxin content variation in other harmful algae that produce STX.     

Pea Protein Provides a Promising Matrix for Microencapsulating Iron

Worldwide, the most prevalent nutritional deficiency is iron. The strategies for iron supplementation often fail due to poor adherence to supplementation methods contributed to unpleasant sensory characteristics. An alternative is the use of microencapsulated nutrients for home fortification in order to mask undesirable tastes and to allow its release in strategic sites of the gastrointestinal tract. Toward this end, pea protein concentrate was tested as a natural, edible and alternative material and the spray-drying technique was utilized for the preparation of microparticles containing ferrous sulfate. Their physical and chemical characteristics were evaluated. The microparticles had a spherical shape and grooves with an average size ranging between 2 and 3 μm. Analysis by in vitro assays tested the release of iron in simulated salivary and gastric fluids and its intestinal absorption in Caco-2 cells. No dissolution of iron occurred in the salivary medium whereas the sensory analysis showed good acceptance of a product which incorporated 5.5 mg of iron per 100 g portion of food. Thus, the effectiveness of microencapsulation was demonstrated by utilizing a plant protein as an encapsulating matrix for the controlled release of iron and capable of preserving the bioaccessibility of ferrous sulfate.     

Ammonia and nitrite toxicity to false clownfish <Emphasis Type="Italic">Amphiprion ocellaris</Emphasis>

False clownfish, Amphiprion ocellaris, is one of the most commercialized fish species in the world, highly produced to supply the aquarium market. The high stocking densities used to maximize fish production can increase ammonia and nitrite to toxic levels. In this study, A. ocellaris juveniles (1.20 ± 0.34 g) were exposed to six concentrations of ammonia ranged from 0.23 to 1.63 mg/L NH₃-N and eight concentrations of nitrite (26.3–202.2 mg/L NO₂ ^?-N). The LC₅₀- 24, LC₅₀-48, LC₅₀-72 and LC₅₀-96 h were estimated to be 1.06, 0.83, 0.75 and 0.75 mg/L for NH₃-N and 188.3, 151.01, 124.1 and 108.8 mg/L for NO₂ ^?-N. Analysis of gill lesions caused by sublethal concentrations of these nitrogenous compounds showed that both nitrogenous compounds induced tissue lesions such as hyperplasia of epithelium cells, hypertrophy of chloride cells and lamellar lifting to all concentrations tested. However, histopathological alterations were more conspicuous accordingly the increase of ammonia or nitrite in fish exposed to 0.57 mg/L NH₃-N or 100 mg/L NO₂ ^?-N. Based on our results, we recommend to avoid concentrations higher than 0.57 mg/L of NH₃-N and 25 mg/L of NO₂-N in water.     

Inheritance of resistance to <Emphasis Type="Italic">Meloidogyne enterolobii</Emphasis> and individual selection in segregating populations of <Emphasis Type="Italic">Psidium</Emphasis> spp

Interaction between the phytonematode Meloidogyne enterolobii and the fungus Fusarium solani has caused direct and indirect losses in the entire guava production chain and consequent extermination of guava plantations throughout Brazil. The combined action of these two pathogens is known as “guava decline”. In order to obtain and assess Psidium spp. interspecific hybrids for resistance to the nematode M. enterolobii, interspecific crosses of P. guineense (susceptible araçá) x P. cattleyanum (resistant araçá); P.guineense (susceptible araçá) x P. guajava (susceptible guava) and P. cattleyanum (resistant araçá) x P. guajava (susceptible guava) were conducted. These crosses resulted in hybrid immune, susceptible and resistant to Meloidogyne enterolobii. The chi-square test rejected the hypothesis of monogenic inheritance with incomplete dominance, which corroborates that this trait has polygenic action. Predictions of genetic values ??and parameters were obtained by the REML / BLUP procedure, at individual level. Finally, the 30 selected individuals (immune and resistant) were obtained, which will be backcrossed with guava for the recovery of the agronomic traits desired and subsequent release of a new cultivar.