A novel genetic marker to differentiate feline herpesvirus type 1 field isolates

Five recent field isolates of feline herpesvirus type 1 (FHV-1) were compared by digestion with a restriction endonuclease, SalI or MluI. The SalI digestion showed a potentially useful difference in one isolate 00-035 that had an approximately 3.0 kbp fragment instead of a 2.6 kbp fragment in the other strains. After cloning the 3.0 and 2.6 kbp fragments, the nucleotide sequences were analyzed. The result showed that the 3.0 kbp fragment of 00-035 included a complete open reading frame of the herpes simplex virus 1 (HSV-1) homologue of the UL17 gene and a 5'-part of UL16 gene and that only one nucleotide substitution was found in the 5'-region of UL17 gene where the SalI site of the 2.6 kbp fragment locates. Based on these nucleotide sequences, two PCR primers were designed to amplify the region around the SalI site in the UL17 gene and the PCR was carried out using 78 field isolates from various parts of Japan. The SalI digestion of the PCR products revealed an interesting profile in that the genotype without the SalI site in UL17 gene was dominant in Tottori and Yamagata prefectures (69% and 75%, respectively) but minor in the other regions of Japan (0-10%). These results suggest that the SalI digestion method described in the present study can be used as a genetic marker to differentiate some FHV-1 field isolates and this is the first report that showed different distributions of FHV-1 genotypes using the novel genetic marker.     

Microconidia act the role as spermatia in the sexual reproduction of <Emphasis Type="Italic">Botrytis cinerea</Emphasis>

In the sexual reproductive cycle of Botrytis cinerea, large numbers of microconidia were observed in all the crosses that formed sexual bodies. To clarify the role of the microconidia in sexual reproduction, they were separated using a sucrose density gradient and then used in crossing tests with fungal sclerotia. Sexual bodies were formed in all the crosses in the five mating combinations, demonstrating that microconidia are able to function as spermatia during sexual reproduction of B. cinerea.     

Cloning of bovine MAIL and its mRNA expression in white blood cells of Holstein cows

Molecule possessing ankyrin-repeats induced by lipopolysaccharide (MAIL) is known as an IkappaB protein induced after administration of bacterial lipopolysaccharide (LPS) to mice. In the present study, we cloned bovine MAIL cDNA and examined its mRNA expression in white blood cells isolated from Holstein cows. Bovine MAIL had more than 80% amino acid identities with murine and human MAILs, highly conserved ankyrin-repeat motifs and PEST-like sequences. Bovine MAIL mRNA was undetectable in isolated peripheral white blood cells, but rapidly induced (<1h) after stimulation by LPS and lipid A in vitro in a dose-dependent manner. The lipid A-induced MAIL mRNA expression was found in polymorphonuclear cells, monocytes/macrophages and total lymphocytes, but not in T-lymphocytes. MAIL mRNA was also induced in vivo in peripheral blood leukocytes of cows after intramammary injection of Escherichia coli derived from coliform mastitis. Thus, bovine MAIL, as rodent MAILs, is induced by inflammatory stimuli in specific immune cells in vitro and in vivo, suggesting a role in inflammatory responses to bacterial infection in cattle.     

Cardiovascular effects of a phosphodiesterase III inhibitor in the presence of carvedilol in dogs

The aim of this study was to determine whether dobutamine, dopamine, or milrinone (a phosphodiesterase [PDE] III inhibitor) would support cardiac function that had been attenuated by administration of the beta-blocker, carvedilol (0.2, 0.4, or 0.8 mg/kg). Hemodynamic and cardiac parameters including the heart rate (HR), left-ventricular fractional shortening (FS), and arterial pressure were measured in six healthy dogs without cardiac disease. Carvedilol did not affect FS or arterial pressure, but decreased the HR significantly. The positive inotropic and chronotropic responses to dobutamine and dopamine were attenuated by carvedilol, whereas arterial pressure was unaffected. Milrinone did not affect the HR and decreased arterial pressure, whereas FS was significantly greater both in the control and carvedilol-treated groups. Although milrinone affect the negative chronotropic effects of carvedilol, milrinone increased FS and prevented the decrease in arterial pressure. These results suggest that inhibition of PDE III preserves cardiac contractility and hemodynamic function in the presence of carvedilol.     

Plasma adrenomedullin concentration in dogs with myxomatous mitral valvular disease

Adrenomedullin (AM), a peptide identified to have vasodilating and natriuretic effects, is involved in the regulation of the cardiovascular system. To evaluate plasma AM concentration in dogs with myxomatous mitral valvular disease (MMVD), and to investigate the associations between the concentrations of plasma AM and natriuretic peptides and the echocardiographic data, we evaluated plasma AM concentrations in 31 healthy control dogs and 57 dogs with MMVD. Plasma AM concentrations in dogs with MMVD were higher than that in the control subjects. The plasma AM concentration increased in conjunction with the severity of heart failure according to the International Small Animal Cardiac Health Council (ISACHC). The AM concentrations were 25.1 ± 5.0 fmol/ml (ISACHC class Ia), 29.9 ± 11.0 fmol/ml (ISACHC class Ib), 43.4 ± 19.8 fmol/ml (ISACHC class II) and 73.5 ± 21.7 fmol/ml (ISACHC class III) and 7.5 ± 5.1 fmol/ml (control group), respectively. The receiver operating characteristic curve indicated an area of 0.93 (95% CI, 0.8801-0.9889; <0.0001), a cutoff value of 30.5 fmol/ml, a sensitivity of 87.1%, and a specificity of 82.5% for the determination of congestive heart failure. Plasma AM concentrations correlated with atrial natriuretic peptide concentrations, LA/Ao ratio, and left ventricular diameter. In conclusion, AM may be a potential diagnostic marker for canine MMVD and possibly plays a pathophysiological role in collaboration with the other neurohumoral factors such as natriuretic peptides.     

Spontaneous Malignant T Cell Lymphoma in a Young Male Common Marmoset (Callithrix jacchus)

We histopathologically and immunohistochemically investigated a case of malignant lymphoma that spontaneously developed in a male common marmoset at two years of age. Beginning at two years four months of age, the animal had an enlargement of the submandibular and inguinal lymph nodes, small subcutaneous nodules near the right breast and an approximately fivefold increase in peripheral lymphocyte count compared with the previous examination value. The postmortem findings at two years eight months of age showed lymphadenopathy with enlargement of the thymus and spleen. Small- to intermediate-sized neoplastic lymphocytes had diffusely proliferated in the enlarged nodes. The neoplastic cells were pleomorphic and had irregularly shaped nuclei. The nuclear chromatin staining revealed hyperchromatism in the small-sized cells, and the intermediate-sized cells exhibited vesicular staining. An immunohistochemical examination indicated that the neoplastic lymphocytes were positive for CD3 and negative for CD20, thus suggesting that they had originated from T cells. In addition, the proliferation of high endothelial venules and reactive epithelioid histiocytes was observed. Scattered tingible body-laden macrophages were infrequently detected. Neoplastic lymphocytes were also observed in the thymus, spleen, heart, lungs, liver, kidneys, adrenal glands and femoral and sternal bone marrow. This malignant lymphoma in a young male common marmoset was considered to fit the category of “peripheral T-cell lymphoma, not otherwise specified (PTCL-NOS)” according to the new WHO system of classification.     

Effect of trehalose supplementation in milk replacer on the incidence of diarrhea and fecal microbiota in preweaned calves

Trehalose, a nonreducing disaccharide consisting of d-glucose with α,α-1,1 linkage, was evaluated as a functional material to improve the gut environment in preweaned calves. In experiment 1, 173 calves were divided into two groups; the trehalose group was fed trehalose at 30 g/animal/d with milk replacer during the suckling period, and the control group was fed nonsupplemented milk replacer. Medication frequency was lower in the trehalose group (P < 0.05). In experiment 2, calves (n = 20) were divided into two groups (control group [n = 10] and trehalose group [n = 10]) based on their body weight and reared under the same feeding regimens as in experiment 1. Fresh feces were collected from individual animals at the beginning of the trial (average age 11 d), 3 wk after trehalose feeding (experimental day 22), and 1 d before weaning, and the fecal score was recorded daily. Fecal samples were analyzed for fermentation parameters and microbiota. The fecal score was significantly lower in the trehalose group than in the control group in the early stage (at an age of 14 to 18 d; P < 0.05) of the suckling period. Calves fed trehalose tended to have a higher proportion of fecal butyrate on day 22 than calves in the control group (P = 0.08). Population sizes of Clostridium spp. were significantly lower (P = 0.036), whereas those of Dialister spp. and Eubacterium spp. tended to be higher in the feces of calves in the trehalose group on day 22 (P = 0.060 and P = 0.083). These observations indicate that trehalose feeding modulated the gut environment and partially contributed to the reduction in medication frequency observed in experiment 1.     

Etiologic study of upper respiratory infections of household dogs

Infectious tracheobronchitis (ITB), also known as the kennel cough, is a respiratory syndrome of dogs and usually appears to be contagious among dogs housed in groups. Etiologic agent of ITB is multiple and sometimes complex. In the present study, 68 household dogs showing clinical signs of respiratory infection were examined, and 20 dogs (29.4%) were found to be positive for either of following agents. Bordetella bronchiseptica (B.b.) was most frequently detected from nasal and oropharynx sites of 7 dogs (10.3%). Among the viruses examined, canine parainfluenza virus (CPIV) was detected with the highest frequency (7.4%). Other pathogens included in the order of frequency group 1 canine coronavirus (4.4%), canine adenovirus type 2 (2.9%), group 2 canine respiratory coronavirus (1.5%), and canine distemper virus (1.5%). Only 2 cases showed mixed infections. Neither influenza A virus nor canine bocavirus (minute virus of canines) was found in any dogs examined. These results indicate that both B.b. and CPIV are likely to be the principal etiologic agents of canine ITB in Japan, and they may be considered as the target for prophylaxis by vaccination.     

Virulence factors of Botrytis cinerea

Botrytis cinerea is responsible for gray mold disease in more than 200 host plant species. The infection of host plants is mediated by numerous extracellular enzymes, proteins and metabolites. Each of these compounds may play a role in different stages of the infection process. Cell wall-degrading enzymes may facilitate the penetration into the host surface, while toxins, oxalic acid and reactive oxygen species may contribute to killing of the host cells. Cell wall-degrading enzymes contribute to the conversion of host tissue into fungal biomass. On the other hand, B. cinerea infection induces biosynthesis of phytoalexins. Therefore, the ability to overcome a wide spectrum of phytoalexins contributes to the pathogenicity of the fungus with a broad host range. The cloning of the corresponding genes has facilitated studies on gene expression and targeted mutagenesis. This review gives an overview of the research performed on virulence factors that play the roles in pathogenesis.     

N2O and NO emissions from an Andisol field as influenced by pelleted poultry manure

Animal manures from intensive livestock operations can be pelleted to improve handlings and recyclings of embodied nutrients. The aim of this study was to evaluate the influence of pelleted poultry manure on N₂O and NO fluxes from an Andisol field. In autumn 2006 and summer 2007, poultry manure (PM), pelleted poultry manure (PP), and chemical fertilizer (CF) were applied at a rate of 120 kg N ha⁻¹ in each cultivation period to Komatsuna (Brassica rapa var. peruviridis). Nitrous oxide and NO fluxes were measured using an automated monitoring system. A soil incubation experiment was also conducted to determine the influence of intact and ground pelleted manure on N₂O, NO, and CO₂ production with a water-filled pore space (WFPS) of 30 or 50%. In the field measurements, N₂O emission rates from the organic fertilizer treatments were larger than that from the CF treatment, possibly because organic C stimulated denitrification. The highest N₂O flux was observed from the PP treatment after a rainfall following fertilization, and the cumulative emission rate (2.72 ± 0.22 kg N ha⁻¹ y⁻¹) was 3.9 and 7.1 times that from the PM and CF treatments, respectively. In contrast, NO emission rates were highest from the CF treatment. The NO/N₂O flux ratio indicated that nitrification was the dominant process for NO and N₂O production from the CF treatment. Cumulative N₂O emission rates from all treatments were generally higher during the wetter cultivation period (autumn 2006) than during the drier cultivation period (summer 2007). In contrast, NO emission rates were higher in the drier than in the wetter cultivation period. The incubation experiment results showed a synergistic effect of soil moisture and the pelleted manure form on N₂O emission rates. The intact pelleted manure with the 50% WFPS treatment produced the highest N₂O and CO₂ fluxes and resulted in the lowest soil NO₃⁻ content after the incubation. These results indicate that anaerobic conditions inside the pellets, caused by rainfall and heterotrophic microbial activities, led to denitrification, resulting in high N₂O fluxes. Controlling the timing of N application by avoiding wet conditions might be one mitigation option to reduce N₂O emission rates from the PP treatment in this study field.