Purification and characterization of chymotrypsin from viscera of vermiculated sailfin catfish, Pterygoplichthys disjunctivus, Weber, 1991

Pterygoplichthys disjunctivus viscera chymotrypsin was purified by fractionation with ammonium sulfate (30–70 % saturation), gel filtration, affinity, and ion exchange chromatography. Chymotrypsin molecular weight was approximately 29 kDa according to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), shown a single band in zymogram. Electrofocusing study suggested being an anionic enzyme (pI ≈ 3.9), exhibiting maximal activity at pH 9 and 50 °C, using Suc-Ala-Ala-Pro-Phe-p-nitroanilide (SAAPNA) as substrate. Enzyme was effectively inhibited by phenyl methyl sulfonyl fluoride (PMSF) (99 %), and N-tosyl-l-phenylalanine chloromethyl ketone (TPCK) (94 %). Enzyme activity was affected by the following ions in decreasing order: Hg²⁺, Fe²⁺, Cu²⁺, Li¹⁺, Mg²⁺, K¹⁺, Mn²⁺, while Ca²⁺ had no effect. Chymotrypsin activity decreased continuously as NaCl concentration increased (from 0 to 30 %). K _m and V _max values were 0.72 ± 1.4 mM and 1.15 ± 0.06 μmol/min/mg of protein, respectively (SAAPNA as substrate). Results suggest the enzyme has a potential application where low processing temperatures are needed, such as in fish sauce production.     

Rubrivivax gelatinosus biomass as an immunostimulant for pacu Piaractus mesopotamicus

Rubrivivax gelatinosus is a bacterium present in the environment successfully used for treatment of fish industry effluent. Besides cleaning the effluent, this bacterium provides a biomass rich in proteins and carotenoid as a metabolic product. This study describes for the first time the use of this biomass as an immunostimulant feed. Haematological, immunological, biochemical and growth parameters in pacu Piaractus mesopotamicus fed control diet or diets supplemented with 0.5 or 1.5 g kg^?1 of the R. gelatinosus biomass during 60 days were assessed. Fish were sampled at 0, 30 and 60 days. The inclusion of increasing levels of biomass in feed increased thrombocytes and neutrophils (P < 0.05) and decreased lymphocytes (P < 0.05) as well as activated the complement system (P < 0.05). Glucose levels, hepatosomatic index and weight were positively affected by the inclusion of R. gelatinosus biomass (P < 0.05). The use of R. gelatinosus biomass in feed improved the immune response of fish through the enhancement of some phagocytic cells, reduced time to activation of the complement system and increased growth parameters. This study reveals a promising use for a by‐product resulted in the wastewater treating in aquaculture.     

Field and laboratory pilot rearing experiments with early ontogenic stages of Concholepas concholepas (Gastropoda: Muricidae)

The carnivorous gastropod Concholepas concholepas, known in Chile as “loco”, is a species present only along the Chilean coast line and in central-southern Perú showing high economic and ecological importance. Its fishery, which in the past decades has ranged between 828 (2001) and 24,828 (1980) metric tons per year [SERNAP, 2005. Servicio Nacional de Pesca: Anuarios estadísticos. (http://www.sernap.cl/paginas/publicaciones/anuarios/index_anuario.php)], is based exclusively on the exploitation of wild stocks. So far, there has been limited interest in tackling the biological and technical feasibility of cultivation of C. concholepas. Hence, the knowledge about its early ontogenetic stages (i.e., larval, early postmetamorphic and small juvenile) cultivation is still deficient. In this study we investigated, under laboratory and field conditions, C. concholepas survivorship, growth rates, feeding rates and the onset of sexual reproduction. Competent loco's larvae were collected in the field and metamorphosed in the laboratory to assess growth rates and survivorship during the first six months of postmetamorphic life. Moreover, using small juvenile C. concholepas of ca. 20 mm of peristomal length, collected in the field, we monitored for the body size and live weight traits in laboratory and field rearing conditions. The feeding of the early postmetamorphics and small juveniles was exclusively based in mono diets of the mussel Semimytilus algosus. The rearing of small juveniles was conducted in two consecutive phases, using two specially designed rearing apparatus. The biochemical content of our laboratory cultivated specimens was compared with values obtained from specimens collected in natural habitats. Their rapid growth and good survivorship makes this species suitable for rearing of cocktail-size specimens (50–60 mm) in less than a year. According to the growing rates reported in this study the legal commercial size of C. concholepas: 100 mm of PL would be reached under field and laboratory conditions in 1.65 and 2.64 years respectively. Moreover, it was found that in our rearing conditions C. concholepas reach sexual maturity in less than a year of benthic life. In summary, considering the findings about C. concholepas high growth rates, and low mortalities, we suggest that our rearing methodologies may be scaled up and implemented for the commercial aquaculture of this unique and valuable muricid.     

Mercury Concentration and Monomethylmercury Production in Sediment: Effect of Dredged Sediment Reuse on Bioconcentration for Ragworms

The effect of dredged sediment reuse on the production and bioconcentration of monomethylmercury (MMHg) was investigated by examining sediments and ragworms found in dredge material banks and surrounding sites in the Venice Lagoon, Italy. Total Hg concentrations in the surface 20 cm of sediments were higher in the banks than in the surrounding sites, but MMHg concentrations were similar, which suggests reduced MMHg production in the banks. Monomethylmercury content in ragworms was significantly lower (p?<?0.05) in the banks than in the surrounding sites. In pore water, concentrations of both sulfate and Fe decreased with depth in the upper 20 cm of the dredge banks. In contrast, sulfate concentrations were constant with depth and large amounts of dissolved Fe occurred in the upper 20 cm of sediments of surrounding sites. Continuous sulfate reduction and possible precipitation of iron sulfide may decrease the production and bioconcentration of MMHg in the dredge material banks compared to the surrounding sites. Overall, the production of MMHg in sediments and its bioconcentration in benthic organisms were connected to the process of sediment diagenesis of organic matter through the control of pore water and sediment geochemistry involving iron and sulfur.     

Babesia bovis contains an abundant parasite-specific protein-free glycerophosphatidylinositol and the genes predicted for its assembly

Autonomous glycosylphosphatidylinositol (GPI) molecules (also protein-free GPIs or free GPIs) have been reported to be particularly abundant in some parasitic protozoa and mediate strong immunomodulatory effects on the host immune system. In the work at hand we have investigated the existence of free GPIs in Babesia bovis. Comparative thin layer chromatographic analysis of the protein-free glycolipid fraction of in vitro cultured B. bovis merozoites and erythrocyte membranes demonstrated the presence of an abundant parasite-specific band. Its chemical analysis revealed a GPI species containing a chain of two mannose residues, N-glucosamine and non-acylated inositol. The lipid moiety linked to inositol was diacylglycerol. The total fatty acid composition showed predominantly long-carbon chain molecules (12% of C_22:0 and 45% of C_24:0). The potential of B. bovis to assemble the presented free GPI species was verified by the existence of seven genes in its genome that putatively encode the following GPI biosynthetic enzymes: PI N-acetyl-GlcN-transferase (PIG-A and GPI-1), N-acetyl-GlcN-PI-de-N-acetylase (PIG-L), acyltransferase (PIG-W), dolichyl-phosphate mannosyl transferase (DPM-1), GPI mannosyltransferase I (PIG-M), and GPI mannosyltransferase II (PIG-V). GPI biosynthesis is vital for the intraerythrocytic parasite stage as mannosamine, an inhibitor of GPI biosynthesis, impaired in vitro growth of B. bovis merozoites. Absence of the vast majority of N-glycan metabolism encoding genes in the B. bovis genome underscores that the growth inhibitory effect of mannosamine is attributable to its interference with GPI biosynthesis and not with assembly of N-linked oligosaccharides, as has been described for higher eukaryotes. Elucidation of the structure and biosynthesis of GPI may allow to facilitate the development of future immune interventions against bovine babesiosis.     

The activity of a selected extract of <Emphasis Type="Italic">Drimys winteri</Emphasis> bark and polygodial on settling and probing behavior of the lettuce aphid <Emphasis Type="Italic">Nasonovia ribisnigri</Emphasis>

An n-hexane extract and the sesquiterpene polygodial isolated from the bark of Drimys winteri J.R. Forster & G. Forster (Winteraceae) were tested to assess their ability to inhibit settling of the aphid Nasonovia ribisnigri Mosley on lettuce leaves; assessments were made using a choice and no-choice leaf-disk assay. The n-hexane extract and the polygodial both significantly inhibited settling in the choice and no-choice tests, but their application on lettuce leaves did not result in any aphid mortality. Probing behavior of N. ribisnigri on polygodial-treated plants was assessed by means of the electrical penetration graph technique. Polygodial interfered with aphid probing behavior by reducing the total probing time, the number of probes, and the number of intracellular punctures. Aphids did not reach the sieve elements of polygodial-treated plants during the 2-h access period. Both tested compounds have the potential to be used in the development of lettuce aphid control agents to reduce the risk of virus transmission.     

Impacts of woodchip amendments and soil nutrient availability on understory vegetation establishment following thinning of a ponderosa pine forest

Mechanical thinning for fire mitigation has become increasingly widespread in recent years throughout the western United States. A common practice in fire-mitigation procedures is the conversion of slash into chipped mulch (referred to as “woodchips”) that is spread on-site. Here, we investigated: (1) the effect of woodchip amendments on soil nitrogen availability, and (2) the influence of potential interactions between woodchip amendments and soil nitrogen availability on patterns of understory plant establishment in a thinned montane forest in the Front Range of Colorado.     

Short‐term effect of the inoculation of probiotics in mature bioflocs: Water quality parameters and abundance of heterotrophic and ammonia‐oxidizing bacteria

The aim of this study was to evaluate the short‐term effect of probiotic inoculation on the abundance of heterotrophic and ammonia‐oxidizing bacteria in mature biofloc, as well as on total suspended solids (TSS), chlorophyll‐a and nitrogenous compounds in water. A completely randomized design consisting of five treatments (three commercial probiotics, one native consortia and one control) was performed. At the beginning of the experiment (day 1), each treatment was inoculated with the respective probiotic: PondPlus^® (PP), Efinol^® PT (EF) and Epicin^® ponds (EP), native consortia UE, whereas the control was not inoculated. Water parameters and bacterial abundance were evaluated at 0, 2, 4, 6 and 8 days. The addition of probiotics, either native or commercial, did not show any significant effect on the TSS, Chl‐a and colony‐forming unit (CFU) of heterotrophic bacteria when they were added to the systems containing mature biofloc. A significant increase in ammonium oxidizing bacteria was registered with the probiotics PP and EP, although the levels of total ammonia nitrogen, NO₃‐N and NO₂‐N were statistically similar among all treatments. Modifications on most of the parameters measured were associated with the factor of time, rather than the inclusion of probiotics. Results suggest that the bacterial conglomerates in mature stage contain well‐established bacterial communities that are difficult to be affected by the addition of probiotics.