Genetic diversity of <Emphasis Type="Italic">Ralstonia solanacearum</Emphasis> strains causing bacterial wilt of solanaceous crops in Myanmar

In 2013 and 2014, an extensive survey of bacterial wilt in Myanmar was performed, and 70 strains of Ralstonia solanacearum (Rs) were collected from wilting plants of tomato, potato, chili and eggplant. Myanmar Rs strains were characterized by traditional and molecular methods. Polymerase chain reaction (PCR) test using Rs-specific primer set amplified one specific band (281-bp) from template DNA of all strains. Pathogenicity tests on the four solanaceous plants differentiated the strains into six pathogenic groups. Biovar determination tests showed that biovar 3 strains predominated (63%) among all Rs strains. Biovar 4 strains (7%) were obtained from both tomato and chili strains, whereas biovar 2 (30%) strains were isolated only from potato. Multiplex-PCR analysis indicated that tomato, eggplant and chili strains belonged to phylotype I, whereas potato strains comprised phylotype I and phylotype II. Strains in phylotype I, which was suggested to have originated from Asia, were the most prevalent in all surveyed areas. Phylogenetic analysis based on the endoglucanase (egl) gene sequences revealed that Myanmar strains partitioned into two major clusters that corresponded to phylotype I and II. Strains in phylotype I were further divided into seven subclusters, each corresponding to a distinct sequevar (15, 17, 46, 47, 48, unknown 1 or unknown 2). All strains in phylotype II belonged to sequevar 1. This is the first comprehensive report of the presence of diverse Rs strains in Myanmar.     

Effect of long‐distance transportation on serum metabolic profiles of steer calves

Long‐distance transportation is sometimes inevitable in the beef industry because of the geographic separation of major breeding and fattening areas. Long‐distance transportation negatively impacts production and health of cattle, which may, at least partly, result from the disturbance of metabolism during and after transportation. However, alteration of metabolism remains elusive in transported cattle. We investigated the effects of transportation on the metabolomic profiles of Holstein steer calves. Non‐targeted analysis of serum concentrations of low molecular weight metabolites was performed by gas chromatography mass spectrometry. Transportation affected 38 metabolites in the serum. A pathway analysis suggested that 26, 10, and 10 pathways were affected immediately after transportation, and 3 and 7 days after transportation, respectively. Some pathways were disturbed only immediately after transportation, likely because of feed and water withdrawal during transit. Nicotinate and nicotinamide metabolism, and citric acid cycle were affected for 3 days after transportation, whereas propionate metabolism, phenylalanine and tyrosine metabolism were affected throughout the experiment. Four pathways were not affected immediately after transportation, but were altered thereafter. These results suggested that many metabolic pathways had marked perturbations during transportation. Metabolites such as citric acid, propionate, tyrosine and niacin can be candidate supplements for mitigating transportation‐induced adverse effects.     

Effect of ginkgo extract supplementation on in vitro rumen fermentation and bacterial profiles under different dietary conditions

Ginkgo extract was applied to a batch culture study and evaluated for its potential as a feed additive for ruminant animals under different forage‐to‐concentrate (F:C) ratios (1:9, 3:7, 5:5, 7:3 and 9:1). Rumen fluid was mixed with respective diet and incubated at 39°C for 24 h with and without ginkgo extract (1.6% fruit equivalent in culture). Methane production was significantly decreased by ginkgo extract, with the greatest reductions found in the 5:5 (41.9%) followed by the 7:3 ratios (36.7%). Total short chain fatty acid and ammonia levels were not affected by ginkgo extract supplementation in any of the five different diets. However, ginkgo extract increased propionate proportion and decreased acetate proportion in all dietary conditions tested. The levels of total bacteria, Ruminococcus flavefaciens, Ruminococcus albus and Fibrobacter succinogenes were decreased by ginkgo extract. The levels of Selenomonas ruminantium, Anaerovibrio lipolytica, Ruminobacter amylophilus, Succinivibrio dextrinosolvens and Megasphaera elsdenii were increased by ginkgo extract supplementation, possibly contributing to the higher propionate production. These results suggest that rumen modulation by ginkgo extract can be achieved at a wide range of F:C ratios with no adverse impact on feed digestion. Moreover, F:C ratios of 5:5 and 7:3 may be optimal when methane mitigation is expected.     

Pink root rot of squash caused by Setophoma terrestris in Japan

Pink root rot of squash (Cucurbita moschata) caused by Setophoma terrestris was found in Maebashi, Gunma Prefecture, Japan in July 2007. Cucumber grafted on the squash first developed wilt and finally blight. These symptoms followed a severe pink root rot of the squash rootstock. The fungal isolates from diseased roots were identified as S. terrestris based on morphological characteristics and nucleotide sequences. One isolate induced a similar pink root rot but not entire wilting of the cucumber vine. We propose the name “pink root rot” (koshoku-negusare-byo in Japanese) of squash for the new disease.     

Association of Verticillium dahliae and Verticillium longisporum with Chinese cabbage yellows and their distribution in the main production areas of Japan

Chinese cabbage yellows caused by Verticillium dahliae and V. longisporum is one of the most economically important diseases in Japan. Verticillium isolates from infected Chinese cabbage in Japan were identified as V. dahliae and V. longisporum based on morphological characteristics and newly developed molecular genotyping methods using group I intron of 18S rDNA, mitochondrial-SSU rDNA and cob gene. Although the two species were equally virulent to Chinese cabbage in the greenhouse, V. longisporum was more virulent in the field compared with V. dahliae. Among the 67 Verticillium isolates from Chinese cabbage, 53 were V. longisporum and considered to be the major pathogen. The isolation frequency of V. longisporum was 98?% in Ibaraki and eastern Gunma where Chinese cabbage was cropped from autumn to winter. In contrast, the frequency was 48?% in Nagano and north to west Gunma where the plant was cropped in cool summer and highland conditions and the rests were V. dahliae. This is the first report to show the different geographical distribution of V. longisporum and V. dahliae in Chinese cabbage production area of Japan.     

Milrinone treatment of bovine oocytes during in vitro maturation benefits production of nuclear transfer embryos by improving enucleation rate and developmental competence

In the production of cattle nuclear transfer embryos, the production efficiency is affected by the oocyte developmental competence and successful enucleation rate. This study investigated the effect of treating oocytes with milrinone, a phosphodiesterase inhibitor, on these two characteristics. When cumulus-oocyte complexes (COCs) were cultured for 19 h with 0, 50 or 100 μM of milrinone, the enucleation rate was significantly improved by 100 μM milrinone. However, milrinone treatment during in vitro maturation (IVM) also delayed meiotic progression by at least 2 h, which would affect the examination of enucleation rate and developmental competence of oocytes. Thus, in the second experiment, meiotic resumption was temporarily inhibited with butyrolactone I (BL-I; 100 μM, 18 h) to decrease the delayed maturation caused by milrinone; this enabled a more accurate comparison of the effects of milrinone after oocyte maturation. In nuclear transfer embryo production, oocytes treated with milrinone (100 μM, 20 h) showed a significantly higher rate of enucleation compared with that of control oocytes. This improved enucleation rate was associated with a closer location of the metaphase plate to the first polar body in the treated oocytes compared with that in control oocytes. Furthermore, milrinone improved the frequency of development to the blastocyst stage in the resulting embryos. In conclusion, milrinone supplementation during IVM improved enucleation rates by rendering the metaphase plate in close proximity to the first polar body, and this treatment also improved oocyte developmental competence. These benefits additively improved the yield of cloned embryos that developed to the blastocyst stage.     

Molecular analyses of a potentially novel Anaplasma species closely related to Anaplasma phagocytophilum detected in sika deer (Cervus nippon yesoensis) in Japan

An Anaplasma species closely related to Anaplasma phagocytophilum detected in sika deer in Hokkaido, Japan was molecularly analyzed using 16S rRNA, citrate synthase (gltA), and heat-shock operon (groEL) gene sequences. Genome walking was performed to determine its complete gltA and groEL sequences (1233 bp and 1650 bp, respectively). Percent identities to the closest A. phagocytophilum sequences from the US and European strains were 98.6-98.8%, 76.5%, and 80.3-80.8% for 16S rRNA, gltA, and groEL genes, respectively. For deduced amino acid sequences, percent identities to the closest A. phagocytophilum sequences were 66.7% and 97.6% for gltA and groEL genes, respectively. Phylogenetic analyses revealed divergence from any known A. phagocytophilum strain. The lower identities and the divergent phylogenetic position of the Anaplasma sp. detected from sika deer in Japan with established A. phagocytophilum strains provide evidence of its potential novelty.     

Efficacy of soluble recombinant FliC protein from Salmonella enterica serovar enteritidis as a potential vaccine candidate against homologous challenge in chickens

FliC, the flagellin antigen of Salmonella Enteritidis, was tested as a vaccine candidate for protective effect against a homologous challenge in chickens. After immunization with recombinant FliC (rFliC) or administration of phosphate-buffered saline (PBS) at 56 days old, the chickens were challenged with 10(9) colony-forming units of Salmonella Enteritidis at 76 days old. The vaccinated birds showed significantly decreased bacterial counts in the liver and cecal contents compared to those administered PBS at 7 days postchallenge, but the protection was partial. The replication experiment also showed a similar result. In both experiments, vaccination induced an increased level of serum anti-rFliC IgG, which was also reactive to the native flagella. The intestinal IgA level was slightly higher in the vaccinated birds than in the control. However, neither the proliferative response nor interferon-gamma secretion of splenic cells upon stimulation with rFliC was induced. Therefore, the effect of rFliC as a vaccine is limited, and further improvement is needed.     

Feline demodicosis caused by an unnamed species

A case of feline demodicosis is described in this report. A 13-year-old spayed female domestic short hair cat weighing 4.5 kg was being treated with cefovecin and alternately with prednisone or methylprednisolone. On further physical examination, the cat showed mild erythema and hair loss on the bridge of the nose, around the eyes, on the chin, on the side part of the breast and on the abdomen. A large number of Demodex mites were found in deep skin scrapings from the affected areas. The cat was then treated with ivermectin at 600 μg/kg administered SC daily. After 4 weeks of treatment, the cat was clinically normal with no mites detected in the skin scrapings from the face or breast areas. The mite responsible may represent a previously seen but as yet unnamed new species. This is third report that describes a case of feline demodicosis caused by a different, unnamed mite species that has different morphological characteristics to those of known Demodex mites and may represent a previously seen but as yet unnamed species.