Milk vetch dwarf virus infection in the Solanaceae and Caricaceae families in Southeast Asia

Milk vetch dwarf virus (MDV) is an important member of the genus Nanovirus and is transmitted by the aphid Aphis craccivora. MDV has multiple single-stranded DNA genome components, each approximately 1 kb, and two or three alpha-satellite molecules. It mainly infects plants of the legume family Fabaceae. Recently, papaya (Carica papaya) collected in Yesan, South Korea, displaying symptoms of leaf yellowing and dwarfism, was identified as a new host for MDV. To examine the geographical distribution of MDV, papaya samples with symptoms were harvested in South Korea, Vietnam, and Taiwan in August 2018, along with tomato and pepper samples grown in adjacent fields in Vietnam. The results revealed the presence of MDV not only in papaya but also in pepper and tomato. This MDV infection in members of the Solanaceae family was confirmed by Southern blot hybridization performed using a PCR product of segment S as a probe. Based on sequence analysis of three MDV components (M, S, and C3), we verified the presence of three different isolates of MDV in these three countries and homology between sequences of isolates from papaya and from members of the Solanaceae from Vietnam. Taken together, our results clearly demonstrate MDV infection in Vietnam and Taiwan for the first time and confirm that MDV can infect economically important pepper and tomato.     

Resistance to Streptococcus iniae and its genetic associations with traits of economic importance in Asian seabass (Lates calcarifer)

Streptococcus iniae is one of the most serious aquatic pathogens, causing significant economic losses in marine and freshwater species, including Asian seabass (Lates calcarifer). Controlling this gram‐positive bacterial pathogen has been an issue in aquaculture systems, due to the combined effects of aquaculture intensification and climatic impacts. To date, there have not been any genetic parameter estimates for S. iniae resistance in Asian seabass. The main aim of this study was to examine genetic variation in S. iniae resistance and its genetic correlations with growth and cannibalism in Asian seabass families produced from a breeding programme for high growth in 2016 and 2017. The study included a total of 5,835 individual fish that were offspring of 41 sires and 60 dams (31 half‐sib and 34 full‐sib families). The experimental fish were challenged by intraperitoneal injection with a volume containing 10⁵ CFU (colony‐forming unit)/fish. Resistance to S. iniae was measured as survival rate at 6 hr, 3, 5, 7, 10 and 15 days post‐challenge test. There were significant variations in S. iniae resistance among families at different observation periods (ranging from 24.4% to 80%). Restricted maximum‐likelihood method and mixed model analysis were applied to estimate heritability for S. iniae resistance. The heritability for S. iniae resistance ranged from 7% to 18% across different statistical models used. The common full‐sib effects accounted for 0.1%–2% of the total variation in resistance to S. iniae. Genetic correlations of the S. iniae resistance at 6 hr and 3 days with later post‐challenge test periods were low to moderate. However, these estimates for S. iniae resistance between successive measurement times (5, 7, 10 and 15 days) were high and close to 1. The genetic correlations of resistance with body weights at 180, 270 and 360 days post‐hatch were not significant as well with cannibalism. It is concluded that there is substantial additive genetic variation in resistance to S. iniae, suggesting there is potential for genetic improvement of Asian seabass for resistance to S. iniae through selective breeding.     

Development of a continuous cell line from larval Atlantic cod (Gadus morhua) and its use in the study of the microsporidian,Loma morhua

In vitro cell culture methods are crucial for the isolation, purification and mass propagation of intracellular pathogens of aquatic organisms. Cell culture infection models can yield insights into infection mechanisms, aid in developing methods for disease mitigation and prevention, and inform commercial‐scale cultivation approaches. This study details the establishment of a larval cell line (GML‐5) from the Atlantic cod (Gadus morhua) and its use in the study of microsporidia. GML‐5 has survived over 100 passages in 8 years of culture. The line remains active and viable between 8 and 21°C in Leibovitz‐15 (L‐15) media with 10% foetal bovine serum and exhibits a myofibroblast phenotype as indicated by immuno‐positive results for vimentin, α‐smooth muscle actin, collagen I and S‐100 proteins, while being desmin‐negative. GML‐5 supports the infection and development of two microsporidian parasites, an opportunistic generalist (Anncaliia algerae) and cod‐specific Loma morhua. Using GML‐5, spore germination and proliferation of L. morhua was found to require exposure to basic pH and cool incubation temperatures (8°C), in contrast to A. algerae, which required no cultural modifications. Loma morhua‐associated xenoma‐like structures were observed 2 weeks postexposure. This in vitro infection model may serve as a valuable tool for cod parasitology and aquaculture research.     

Plum pox virus: diagnosis and spread inhibition by weed control

Journal of Plant Diseases and Protection - Plum pox virus (PPV) is a plant virus (genus Potyvirus, family Potyviridae) infecting stone fruit trees. Since the first report from Bulgaria in 1917, PPV...     

Application of nox-restriction fragment length polymorphism for the differentiation of Brachyspira intestinal spirochetes isolated from pigs and poultry in Australia.

Sixty-nine intestinal spirochetes isolated from pigs and poultry in eastern Australia were selected to evaluate the effectiveness of a species-specific PCR-based restriction fragment length polymorphism (RFLP) analysis of the Brachyspira nox gene. For comparative purposes, all isolates were subjected to species-specific PCRs for the pathogenic species Brachyspira hyodysenteriae and Brachyspira pilosicoli, and selected isolates were examined further by sequence analysis of the nox and 16S ribosomal RNA genes. Modifications to the original nox-RFLP method included direct inoculation of bacterial cells into the amplification mixture and purification of the PCR product, which further optimized the nox-RFLP for use in a veterinary diagnostic laboratory, producing sufficient product for both species identification and future comparisons. Although some novel profiles that prevented definitive identification were observed, the nox-RFLP method successfully classified 45 of 51 (88%) porcine and 15 of 18 (83%) avian isolates into 5 of the 6 recognized species of Brachyspira. This protocol represents a significant improvement over conventional methods currently used in veterinary diagnostic laboratories for rapid specific identification of Brachyspira spp. isolated from both pigs and poultry.     

Generation of reactive oxygen species by equine spermatozoa

OBJECTIVE: To characterize generation of reactive oxygen species (ROS) by equine spermatozoa. SAMPLE POPULATION: Multiple semen samples collected from 9 stallions. PROCEDURE: Equine spermatozoa were separated from seminal plasma on a discontinuous polyvinylpyrrolidone (PVP)-coated silica gradient and resuspended in a modified Tyrode albumin-lactate-pyruvate medium. Amount of hydrogen peroxide (H2O2) generated was assayed by use of a 1-step fluorometric assay, using 10-acetyl-3,7-dihydroxyphenoxazine as a probe for detection of H2O2 in a microplate assay format. Concentration of H2O2 was determined by use of a fluorescence microplate reader. RESULTS: Amount of H2O2 generated increased significantly with time and spermatozoa concentration for live and flash-frozen spermatozoa, and amount of H2O2 generated was significantly greater for flash-frozen than for live spermatozoa. Addition of the reduced form of nicotinamide adenine dinucleotide phosphate (NADPH) significantly increased generation of H2O2 by live and flash-frozen spermatozoa. Addition of a calcium ionophore also significantly increased the amount of H2O2 generated by live spermatozoa but did not have an effect on amount of H2O2 generated by flash-frozen spermatozoa. Abnormal equine spermatozoa generated significantly greater amounts of H2O2 than did normal spermatozoa. CONCLUSIONS AND CLINICAL RELEVANCE: Equine spermatozoa generate ROS in vitro, possibly via a NADPH-oxidase reaction. Spermatozoa damaged during flash-freezing or morphologically abnormal spermatozoa generated significantly greater amounts of ROS than did live or morphologically normal spermatozoa. Damaged and abnormal spermatozoa generate greater amounts of ROS that may contribute to reduced fertility or problems related to semen preservation.     

Silk genes support the single origin of orb webs

The orb web is a spectacular evolutionary innovation that enables spiders to catch flying prey. This elegant, geometric structure is woven with silk fibers that are renowned for their superior mechanical properties. We used silk gland expression libraries to address a long-standing controversy concerning the evolution of the orb-web architecture. Contrary to the view that the orb-web design evolved multiple times, we found that the distribution and phylogeny of silk proteins support a single, ancient origin of the orb web at least 136 million years ago. Furthermore, we substantially expanded the repository of silk sequences that can be used for the synthesis of high-performance biomaterials.