Evaluation of serological tests for the diagnosis of Neospora caninum infection in dogs: optimization of cut off titers and inhibition studies of cross-reactivity with Toxoplasma gondii

Diagnosis of Neospora caninum infection in dogs is based on serological assays such as the indirect fluorescent antibody test (IFAT) and enzyme-linked immunosorbent assays (ELISA). This study evaluated two serological tests (IFAT and ELISA) for the detection of IgG antibodies to N. caninum in 300 serum samples of dogs through the optimization of cut off titers by using the two-graph receiver-operating characteristic (TG-ROC) curve. In addition, the identification of major cross-reactive antigens with Toxoplasma gondii was investigated by inhibition ELISA and immunoblotting (IB) assays. IFAT and ELISA results showed 74% agreement, with a good negative concordance (P(neg)=0.83), but a poor positive concordance (P(pos)=0.42). The great majority (86%) of sera with positive concordant results (IFAT+/ELISA+) recognized at least two out of three N. caninum immunodominant antigens, particularly the 29-32 and 35-37 kDa bands. Optimization of cut off titers in IFAT and ELISA was performed considering the reactivity to at least two out of three N. caninum immunodominant antigens as infection markers, obtaining a titer of 50 for IFAT and 200 for ELISA. Seropositivity to N. caninum was significantly associated with T. gondii-seropositive samples, particularly in ELISA (55.4%). Inhibition ELISA curves for N. caninum showed a partial heterologous inhibition, indicating some degree of cross-reactivity between N. caninum and T. gondii antigens. Inhibition IB assays showed a moderate heterologous inhibition for N. caninum antigens above 45-50 kDa. These results indicate that ELISA should be used critically when crude tachyzoite antigen preparations are employed, due to possible cross-reactivity with other related parasites as T. gondii. Also, the cut off dilution of 1:50 in IFAT showed to be the most appropriated for N. caninum serology in dogs. Therefore, we suggest that N. caninum immunodominant antigens, specially the 17 and 29-32 kDa proteins, should be selected markers in serological assays for canine neosporosis.     

Comparison of estimates of hip dysplasia genetic parameters in Estrela Mountain Dog using linear and threshold models

Genetic parameters, breeding values, and genetic trends of hip dysplasia in Estrela Mountain Dogs were estimated using a linear model (LM) and a threshold model (TM). A database with 313 animals was used. Right and left hip joints were individually scored, according to the Fédération Cynologic Internationale grading rules of the canine hip dysplasia system, as normal (1), borderline (2), slight (3), moderate (4), and severe (5 and 6). The estimate of repeatability was lower in LM (0.86) than in TM (0.90). The same tendency was verified with the heritability because its estimate in LM was 0.38 and in TM was 0.43. However, these results did not establish any statistical differences between the models. The genetic trend of canine hip dysplasia for LM and TM showed a similarity in shape, but considerable individual differences were found in the EBV ranking lists. Therefore, the selection of breeding animals would not be the same with the 2 methodologies. To select the best method for genetic evaluation of hip dysplasia, further studies using more data and other dog breeds are required.     

Broodstock conditioning of the Portuguese oyster (Crassostrea angulata,Lamarck, 1819): influence of different diets

The Portuguese oyster Crassostrea angulata shows great potential in oyster farming. The conservation of pure populations of this species is important for production diversification and biodiversity preservation. In this way, the zootechnological development for seed hatchery production is extremely important. Broodstock conditioning is a key step in the process of rearing bivalves in a hatchery. Many factors regulate the reproductive cycle, being food one of the most important ones. To evaluate the effect of different diets on C. angulata reproductive performance, broodstock were conditioned with different food regimes formulated fundamentally by flagellates (Diet 1 – Pavlova lutheri and Isochrysis galbana clone T‐ISO; Diet 2 – P. lutheri, T‐ISO and Skeletonema costatum) and constituted fundamentally by diatoms (Diet 3 – S. costatum and Chaetoceros calcitrans; Diet 4 – P. lutheri, S. costatum and C. calcitrans). During conditioning, samples of oysters were collected to evaluate condition index, gonadal development and biochemical composition. At the end of the conditioning period, oysters were induced to spawn to evaluate reproductive output (fecundity, fertilization rate and D‐larvae development). The diets had an impact on the gametogenesis process, energy storage and reproductive output performance, being the best results those obtained in broodstock fed with the diatoms‐predominant diets. However, those fed with diets majority flagellates had an unsuccessful performance. Holistic approaches incorporating all results in this study reveal and reinforce the idea that the diatom species used presented the nutritional requirements to C. angulata broodstock, being essential in the conditioning phase.     

Biochemical comparison of lectins among three different color strains of the red alga Kappaphycus?alvarezii

The red alga Kappaphycus alvarezii is economically important as an edible species and as a source of carrageenan, and has been extensively cultivated in many tropical countries. For this species, different color strains, which differ from each another in growth rate and carrageenan content, have been reported for decades. In this study, lectins from brown, red, and green strains of K. alvarezii cultivated in Vietnam were isolated and characterized for evaluation of their biochemical properties and contents. The results showed that each color strain contained in common the three lectins, named KAA-1, KAA-2, and KAA-3, which shared the hapten-inhibition profile of hemagglutination, 20 N-terminal amino acid sequence, and equivalent molecular mass within a range of 28,016 ± 1.2 to 28,021 ± 1.8 Da, but differed in their yields, with the highest yield of KAA-2. These properties of the three isolectins were also comparable among the three different color strains. However, the sum of the yields of the three isolectins decreased in the order: red (21.4 mg) to green (15.9 mg) to brown strains (15.1 mg), from 500 g fresh alga. Thus, this algal species can be a good source of useful lectins, irrespective of color strain.     

Phytoavailability of lead altered by two Pelargonium cultivars grown on contrasting lead-spiked soils

Purpose

This study assesses the potential of two contrasted fragrant Pelargonium cultivars to induce pH and dissolved organic carbon (DOC) changes in the soil solution, Pb speciation, and their subsequent effects on rhizosphere phytoavailable Pb.

Materials and methods

Rooted plantlets were grown in special devices, floating on aerated nutrient solution in PVC tanks. This setup allows roots to be physically separated, through a mesh, from a 3-mm soil matrix layer that can be considered as rhizosphere soil. Two contrasted soils, each spiked with Pb-rich particles, emitted from a battery recycling industry, were used at total burdens of 500 and 1500 mg Pb kg^?1 in addition to a control unspiked soil. Soil solution pH, phytoavailable Pb, DOC, Pb adsorption, precipitation on roots, and Pb phases in soil and plant were investigated.

Results and discussion

Attar of Roses (Attar) cultivar acidified its rhizosphere by 0.4 pH units in both spiked soils. Concolor Lace (Concolor) was unable to change soil solution pH on soil-1 and increased it by 0.7 units on soil 2. Concentrations of Pb in soil solution from Attar plants were always higher than those of Concolor ones. DOC contents of both unspiked soil-1 and soil-2 without plants were not significantly different. In the case of spiked samples, DOC contents in the rhizosphere soil were increased by three and two times for Attar and Concolor, respectively, compared to the unspiked soil without plant. Both cultivars were able to increase DOC contents, independent of soil type and level of contamination. Accumulation of Pb in shoots and roots was higher in Attar as compared to Concolor due to enhanced available Pb as a result of pH and DOC modifications of the rhizosphere soil. Significant amounts of Pb were adsorbed on roots of both cultivars. X-ray elemental analysis of precipitates on roots revealed the association of Pb with P in cylinder-like structures. Extended X-ray absorption fine structure (EXAFS) spectroscopy revealed that Pb was present, to a major extent in the inorganic form, mainly as PbSO₄ in the soil, whereas it was complexed with organic species within plant tissues. The conversion of Pb into organic species could decrease toxicity, may enhance plant tolerance, and could increase translocation.

Conclusions

Plant-induced changes were responsible for the modification of lead phases within the soil. Immobile forms present in the source leaded particles as well as in the soils were converted into soluble species, ultimately improving the phytoavailable or soil solubilized Pb.

     

Modeling aerobic decomposition of rice straw during the off-rice season in an Andisol paddy soil in a cold temperate region of Japan: Effects of soil temperature and moisture

Submerged rice paddies are a major source of methane (CH₄) which is the second most important greenhouse gas after carbon dioxide (CO₂). Accelerating rice straw decomposition during the off-rice season could help to reduce CH₄ emission from rice paddies during the single rice-growth season in cold temperate regions. For understanding how both temperature and moisture can affect the rate of rice straw decomposition during the off-rice season in the cold temperate region of Tohoku district, Japan, a modeling incubation experiment was carried out in the laboratory. Bulk soil and soil mixed with 2% of δ¹³C-labeled rice straw with a full factorial combination of four temperature levels (?5 to 5, 5, 15, 25°C) and two moisture levels (60% and 100% WFPS) were incubated for 24 weeks. The daily change from ?5 to 5°C was used to model the freezing–thawing cycles occurring during the winter season. The rates of rice straw decomposition were calculated by (i) CO₂ production; (ii) change in the soil organic carbon (SOC) content; and (iii) change in the δ¹³C value of SOC. The results indicated that both temperature and moisture affected the rate of rice straw decomposition during the 24-week aerobic incubation period. Rates of rice straw decomposition increased not only with high temperature, but also with high moisture conditions. The rates of rice straw decomposition were more accurately calculated by CO₂ production compared to those calculated by the change in the SOC content, or in its δ¹³C value. Under high moisture at 100% WFPS condition, the rates of rice straw decomposition were 14.0, 22.2, 33.5 and 46.2% at ?5 to 5, 5, 15 and 25°C temperature treatments, respectively. While under low moisture at 60% WFPS condition, these rates were 12.7, 18.3, 31.2 and 38.4%, respectively. The Q₁₀ of rice straw decomposition was higher between ?5 to 5 and 5°C than that between 5 and 15°C and that between 15 and 25°C. Daily freezing–thawing cycles (from ?5 to 5°C) did not stimulate rice straw decomposition compared with low temperature at 5°C. This study implies that to reduce CH₄ emission from rice paddies during the single rice-growth season in the cold temperate regions, enhancing rice straw decomposition during the high temperature period is very important.     

Entry Inhibition of Influenza Viruses with High Mannose Binding Lectin ESA-2 from the Red Alga Eucheuma serra through the Recognition of Viral Hemagglutinin

Lectin sensitivity of the recent pandemic influenza A virus (H1N1-2009) was screened for 12 lectins with various carbohydrate specificity by a neutral red dye uptake assay with MDCK cells. Among them, a high mannose (HM)-binding anti-HIV lectin, ESA-2 from the red alga Eucheuma serra, showed the highest inhibition against infection with an EC₅₀ of 12.4 nM. Moreover, ESA-2 exhibited a wide range of antiviral spectrum against various influenza strains with EC₅₀s of pico molar to low nanomolar levels. Besides ESA-2, HM-binding plant lectin ConA, fucose-binding lectins such as fungal AOL from Aspergillus oryzae and AAL from Aleuria aurantia were active against H1N1-2009, but the potency of inhibition was of less magnitude compared with ESA-2. Direct interaction between ESA-2 and a viral envelope glycoprotein, hemagglutinin (HA), was demonstrated by ELISA assay. This interaction was effectively suppressed by glycoproteins bearing HM-glycans, indicating that ESA-2 binds to the HA of influenza virus through HM-glycans. Upon treatment with ESA-2, no viral antigens were detected in the host cells, indicating that ESA-2 inhibited the initial steps of virus entry into the cells. ESA-2 would thus be useful as a novel microbicide to prevent penetration of viruses such as HIV and influenza viruses to the host cells.     

Estimating Distance Sampling Detection Functions When Distances Are Measured With Errors

Distance sampling methods assume that distances are known but in practice there are often errors in measuring them. These can have substantial impact on the bias and precision of distance sampling estimators. In this paper we develop methods that accommodate both systematic and stochastic measurement errors. We use the methods to estimate detection probability in two surveys with substantial measurement error. The first is a shipboard line transect survey in the North Sea in which information on measurement error comes from photographically measured distances to a subset of detections. The second is an aerial cue-counting survey off Iceland in which information on measurement error comes from pairs of independently estimated distances to a subset of detections. Different methods are required for measurement error estimation in the two cases. We investigate by simulation the properties of the new estimators and compare them to conventional estimators. They are found to perform better than conventional estimators in the presence of measurement error, more so in the case of cue-counting and point transect estimators than line transect estimators. An appendix on the asymptotic distributions of conditional and full likelihood estimators is available online.