Melanin biosynthesis in Pyricularia oryzae: Site of tricyclazole inhibition and pathogenicity of melanin-deficient mutants

Tricyclazole (5-methyl-1,2,4-triazolo[3,4-b]benzothiazole) inhibits melanin synthesis in Pyricularia oryzae at concentrations less than 0.01 μg/ml. The primary site of inhibition in the biosynthetic pathway occurs between scytalone and vermelone. Accumulation of several metabolites derived from melanin precursors along branch pathways is associated with inhibition of melanin biosynthesis. At low tricyclazole concentrations (0.01–1 μg/ml), predominant accumulation of 2-hydroxyjuglone and 3,4-dihydro-3,4,8-trihydroxy-1-(2H)-naphthalenone (3,4,8-DTN) occurs as a result of the primary block between 1,3,8-trihydroxynaphthalene and vermelone. As the concentration of tricyclazole is increased from 1 to 10 μg/ml, flaviolin accumulation is markedly enhanced, whereas that of 3,4,8-DTN and 3,4-dihydro-4,8-dihydroxy-1-(2H)-naphthalenone is depressed, indicating possible secondary sites of inhibition in the main and branch pathways. Five melanin-deficient mutants of P. oryzae that phenotypically resemble the tricyclazole-treated wild-type strain were nonpathogenic or rarely infected two rice varieties. Three of the mutants studied were genetically defective in the melanin biosynthetic pathway at the site blocked by tricyclazole in the wild type. The wild-type strain converted both scytalone and vermelone to melanin; whereas the three mutants and the tricyclazole-treated wild type converted only vermelone to melanin. The data suggest a relationship between melanin biosynthesis and pathogenicity in P. oryzae.     

Caprine toxoplasmosis in Southern Brazil: a comparative seroepidemiological study between the indirect immunofluorescence assay,the enzyme-linked immunosorbent assay,and the modified agglutination test

Tropical Animal Health and Production - This study investigated the prevalence of caprine toxoplasmosis in goat herds from Southern Brazil by the indirect immunofluorescence assay (IFA), and...     

Fucose-Rich Sulfated Polysaccharides from Two Vietnamese Sea Cucumbers Bohadschia argus and Holothuria (Theelothuria) spinifera: Structures and Anticoagulant Activity

Fucosylated chondroitin sulfates (FCSs) FCS-BA and FCS-HS, as well as fucan sulfates (FSs) FS-BA-AT and FS-HS-AT were isolated from the sea cucumbers Bohadschia argus and Holothuria (Theelothuria) spinifera, respectively. Purification of the polysaccharides was carried out by anion-exchange chromatography on DEAE-Sephacel column. Structural characterization of polysaccharides was performed in terms of monosaccharide and sulfate content, as well as using a series of non-destructive NMR spectroscopic methods. Both FCSs were shown to contain a chondroitin core [→3)-β-d-GalNAc-(1→4)-β-d-GlcA-(1→]_n bearing sulfated fucosyl branches at O-3 of every GlcA residue in the chain. These fucosyl residues were different in pattern of sulfation: FCS-BA contained Fuc2S4S, Fuc3S4S and Fuc4S at a ratio of 1:8:2, while FCS-HS contained these residues at a ratio of 2:2:1. Polysaccharides differed also in content of GalNAc4S6S and GalNAc4S units, the ratios being 14:1 for FCS-BA and 4:1 for FCS-HS. Both FCSs demonstrated significant anticoagulant activity in clotting time assay and potentiated inhibition of thrombin, but not of factor Xa. FS-BA-AT was shown to be a regular linear polymer of 4-linked α-L-fucopyranose 3-sulfate, the structure being confirmed by NMR spectra of desulfated polysaccharide. In spite of considerable sulfate content, FS-BA-AT was practically devoid of anticoagulant activity. FS-HS-AT cannot be purified completely from contamination of some FCS. Its structure was tentatively represented as a mixture of chains identical with FS-BA-AT and other chains built up of randomly sulfated alternating 4- and 3-linked α-L-fucopyranose residues.     

Evaluation of non-chemical seed treatment methods for the control of Alternaria dauci and A. radicina on carrot seeds

The current study was initiated to evaluate the efficacy of physical methods (hot water, aerated steam, electron treatment) and agents of natural origin (resistance inducers, plant derived products, micro-organisms) as seed treatments of carrots for control of Alternaria dauci and A. radicina. Control of both Alternaria species by seed treatment with the resistance inducers was generally poor. Results were also not satisfactory with most of the formulated commercial micro-organism preparations. Based on the average of five field trials, one of these, BA 2552 (Pseudomonas chlororaphis), provided a low but significant increase in plant stand. Among the experimental micro-organisms, the best results were obtained with Pseudomonas sp. strain MF 416 and Clonostachys rosea strain IK726. A similar level of efficacy was provided by seed treatment with an emulsion (1%) of thyme oil in water. Good and consistent control was generally achieved with the physical methods aerated steam, hot water and electron treatment. Aerated steam treatment was, apart from the thiram-containing chemical standard, the best single treatment, and its performance may at least partially be due to extensive pre-testing, resulting in dosages optimally adapted to the respective seed lot. In some of the experiments the effect of the hot water treatment, which was tested at a fixed, not specifically adapted dosage, was significantly improved when combined with a Pseudomonas sp. MF 416 or C. rosea IK726 treatment. The results are discussed in relation to the outcome of experiments in which the same seed treatment methods and agents were tested in other seed-borne vegetable pathosystems.     

Tissue distribution of Red Spotted Grouper Nervous Necrosis Virus (RGNNV) genome in experimentally infected juvenile European seabass (Dicentrarchus labrax)

The distribution of viral genome in the tissues of juvenile European seabass (Dicentrarchus labrax) during the course of a Red Spotted Grouper Nervous Necrosis Virus (RGNNV) infection has not yet been described. The present study addresses this and indicates which target organs may be involved in viral replication. This information should enable more accurate detection of virus in asymptomatic carriers, and in turn help to control the spread of the disease. The aim of this study was to examine the pattern of expression of viral genomic segments RNA1 and RNA2, using two absolute real-time PCRs (RT-qPCR), over the course of a RGNNV infection after administering the virus by intramuscular injection. In situ hybridization was also used to locate the RNA2 viral segment in different organs throughout the infection. The experimental challenge provoked an acute form of viral nervous necrosis (VNN), with a resulting cumulative mortality of 37%. The RT-qPCRs designed allowed the detection of both genomic segments in all the organs tested (nervous and non-nervous tissues) at all sampling times examined. The highest viral RNA copy number was found in eyes, although viral replication appeared to begin in the brain. Viral replication was also recorded in pooled internal organs and in caudal fin. However, the increase in the viral RNA copy number in these organs did not result in an increased viral titre, which may indicate that a productive infection does not take place in non-nervous tissues, possibly due to a failure in a viral post-replication step.     

Pseudomonas fluorescens contamination of a feline packed red blood cell unit and studies of canine units

Background: While screening programs have reduced the risk of infectious disease transmission by donors in human and veterinary blood banking, bacterial contamination of blood products has emerged as a major complication in human medicine. Objectives: To describe a Pseudomonas fluorescens (Pf)‐contaminated feline packed RBC (pRBC) unit and experimentally investigate Pf‐contaminated canine pRBCs. Methods: Canine pRBCs were inoculated with Pf‐rich pRBCs from the sentinel feline unit and stored at 4°C or 20°C for 72 hours. Aliquots from the pRBCs were serially evaluated by microscopy, culture, and a eubacterial 16S rRNA real‐time PCR assay. Results: One Pf‐contaminated feline unit turned black after 22 days of storage and was removed from the blood bank; a source was not found, and no other contaminated units were identified. Canine pRBCs spiked with 5 or 25 μL of the sentinel unit became culture‐ and/or 16S PCR‐positive at ≥8 hours at 20°C and 48 hours at 4°C and developed a color change at ≥24 hours. Sensitivity studies indicated that without incubation, inoculation of ≥100 μL Pf‐rich pRBCs was necessary for a positive 16S PCR test result. Conclusions: P. fluorescens grows in stored pRBCs slowly at 4°C and rapidly at 20°C. Screening of blood products for color change, estimating bacterial concentration with microscopy, and 16S PCR testing are simple and fast ways to detect bacteria in stored blood. Aseptic collection, temperature‐controlled storage, and regular visual monitoring of stored units is recommended. Discolored units should not be transfused, but examined for bacterial contamination or other blood product quality problems.     

Seroprevalence and risk factors for Neospora caninum in sheep in the state Minas Gerais, southeastern Brazil

The aim of this study was to determine the prevalence and risk factors associated with infection due to Neospora caninum in serum samples from 488 sheep originating from 63 farms in 63 municipalities distributed across eight of the twelve mesoregions of the state Minas Gerais, southeastern Brazil. For detection of N. caninum the sheep serum samples were subjected to the indirect fluorescence antibody test (IFAT ≥ 50). To identify the risk factors associated with infection due to N. caninum a questionnaire was filled out for each herd by interviewing, the individual responsible for the herd, demanding information on the general characteristics of the property. Sixty-four sheep sera (13.1%; 95% CI=10.3-16.4) presented IgG-specific anti-N. caninum antibodies with the following titers: 50 (49; 76.6%), 100 (7; 10.9%), 200 (4; 6.2%), 400 (3; 4.7%) and 800 (1; 1.6%). The prevalence of infected sheep per mesoregion ranged from 0 to 28.1%. Out of the 63 farms sampled, 31 (49.2%; 95% CI=36.4-62.1) presented at least one seropositive sheep. No significant association was found between the presence of anti-N. caninum antibodies and the risk factors evaluated on the farms, except for the mesoregion variable (p=0.004; OR=0.429; CI95%=0.182-1.008). These results indicate that there is a need for additional research to define the epidemiological importance of this parasite as a cause of reproductive problems in sheep herds in Minas Gerais.     

Effects of vaccination against brucellosis and clostridia on the intake,performance, feeding behavior,blood parameters,and immune responses of dairy heifers calves

The aim of this study was to identify possible effects of different vaccination strategies (concomitantly or not) against brucellosis and clostridia on intake, performance, feeding behavior, blood parameters, and immune responses of dairy heifers calves. Fifty heifers calves were enrolled [38 Gyr (Zebu, Bos taurus indicus) and 12 5/8 Holstein Gyr]. At 120 d of age, animals were randomly distributed among 3 groups: B (n = 18), vaccinated against brucellosis; C (n = 14), vaccinated against clostridia and CB (n = 18), vaccinated concomitantly for both. Rectal and thermographic temperatures were evaluated on days 1, 0, 1, 2, 3, 5, 7,10, 14, and 28 relatives to the vaccination day. Feed and water intake, body weight (BW), and feeding behavior were monitored daily by an electronic feeding system. Blood was sampled on days 0, 3, 7, 14, and 28, relative to the vaccination day for determination of glucose and -hydroxybutyrate (BHBA) concentrations. Blood sampled on day 0 (prevaccination) and on days 28 and 42 were used to evaluate the immune response against Brucella abortus and clostridia. There was an increase in rectal temperature between the first and the third day postvaccination in the 3 groups. The thermography revealed an increase of local temperature for 7 d on groups B and CB. Group C had increased local temperature for a longer period, lasting for up to 14 d. Dry mater intake was reduced for groups B and CB, but no alteration was observed for group C. No alterations regarding initial BW, final BW, average daily weight gain, and feed efficiency were observed. No differences were observed for the 3 vaccination groups for blood parameters throughout the evaluation period. The concomitant vaccination against brucellosis and clostridia led to lower neutralizing antibody titers against epsilon toxin of Clostridium perfringens and botulinum toxin type C of C. botulinum (C > CB > B). When cellular proliferation assay and serological tests to B. abortus were evaluated, no differences were observed between groups B and CB. The present results indicate that the concomitant vaccination against brucellosis and clostridia has no relevant impact on the intake, performance, and feeding behavior of dairy calves. However, the concomitant vaccination of vaccines against these 2 pathogens impacts animal immunity against clostridial infections.