Survey and molecular characterization of Cryptosporidium and Giardia spp. in owned companion animal, dogs and cats, in Japan

Compared with other countries, surveys of these parasites have been rarely performed in companion animals of Japan in spite of their significance for public health. Here, we investigated pet dogs and cats in Japan for the first time, and genetically analyzed the isolates to evaluate the risk of zoonotic infections. Seventy-seven fecal samples were collected from privately owned dogs and 55 samples from owned cats in Osaka city, Japan. Cryptosporidium oocysts were identified in 3/77 dogs (3.9%) and 7/55 cats (12.7%), and Giardia infection in 2/77 dogs (2.6%) and 1/55 cats (1.8%). Amplification of the target regions for genotyping was successful, Cryptosporidium isolates in dogs and cats were identified as C. canis and C. felis, respectively, and those of Giardia in dogs and cats were G. intestinalis Assemblages D and F. The discharge period of the oocysts varied within 3-16 weeks and that of the cysts was 12 weeks. To date, zoonotic types of both parasites have been identified in other animals in Japan, and further large-scale studies are needed to determine the distribution of zoonotic genotypes in these animals, especially those closely associated with humans.     

Induction of apoptotic lesions in liver and lymphoid tissues and modulation of cytokine mRNA expression by acute exposure to deoxynivalenol in piglets

Six 1-month-old piglets were intravenously injected with deoxynivalenol (DON) at the concentration of 1 mg/kg body weight, with three pigs each necropsied at 6 and 24 h post-injection (PI) for investigation of hepatotoxicity and immunotoxicity with special attention to apoptotic changes and cytokine mRNA expression. Histopathological examination of the DON-injected pigs revealed systemic apoptosis of lymphocytes in lymphoid tissues and hepatocytes. Apoptosis of lymphocytes and hepatocytes was confirmed by the TdT-mediated dUTP-biotin nick end-labeling (TUNEL) method and immunohistochemical staining against single-stranded DNA and cleaved caspase-3. The number of TUNEL-positive cells in the thymus and Peyer''s patches of the ileum was increased at 24 h PI compared to 6 h PI, but the peak was at 6 h PI in the liver. The mRNA expression of interleukin (IL)-1β, IL-6, IL-18, and tumor necrosis factor (TNF)-α in the spleen, thymus and mesenteric lymph nodes were determined by semi-quantitative RT-PCR, and elevated expression of IL-1β mRNA at 6 h PI and a decrease of IL-18 mRNA at 24 h PI were observed in the spleen. IL-1β and IL-6 mRNA expressions increased significantly at 6 h PI in the thymus, but TNF-α decreased at 6 h PI in the mesenteric lymph nodes. These results show the apoptosis of hepatocytes suggesting the hepatotoxic potential of DON, in addition to an immunotoxic effect on the modulation of proinflammatory cytokine genes in lymphoid organs with extensive apoptosis of lymphocytes induced by acute exposure to DON in pigs.     

Fibrolipoma of a cryptorchid testis in a young bull

A six-month-old Japanese Black bull was found to have no left testis in the scrotum. A fist-sized mass was palpated per rectum. Two months later, hCG was injected and blood samples were collected before and after injection. No testosterone response to hCG was observed. On the cut surface of the excised mass, most of the mass was composed of homogeneous adipose-like tissue. The rest of the surface was composed of a well-circumscribed testicular parenchyma-like tissue (18 × 16 × 15 mm). Histology revealed diffusely distributed mature adipocytes and septa of fibrous connective tissue. Neither germ cells nor spermatozoa were observed in the seminiferous tubules. A diagnosis was made of fibrolipoma and aspermatogenesis of the left cryptorchid testis.     

Development of a new selective medium for isolation of methicillin-resistant Staphylococcus aureus

A new selective medium containing cephem antibiotics was developed for isolation of methicillin-resistant Staphylococcus aureus (MRSA). MRSA colonies on a medium containing ceftazidime (CAZ) were most easily identifiable and a medium containing cefoperazone (CPZ) was superior in suppressing the growth of other bacteria. With the medium containing a couple of CAZ and CPZ, MRSA and methicillin-resistant coagulase-negative staphylococci (MRCNS) were detected from 2 and 1 of 15 chicken meat samples respectively. The MRSA and MRCNS recovery test showed that the medium was effective for MRSA isolation, suppressing the growth of other bacteria efficiently. These results suggested that the medium containing a couple of CAZ and CPZ was useful for MRSA detection from foods and animals.     

Phylogenetic identification of Cystoisospora spp. from dogs, cats, and raccoon dogs in Japan

Cystoisospora spp. from feces in dogs, cats, and raccoon dogs were isolated, sequenced at the small subunit ribosomal RNA gene locus and compared to other Cystoisospora spp. Cystoisospora oocysts from dogs and raccoon dogs were morphologically similar with those of C. ohioensis, and cat isolates were similar with those of C. felis. The sequences from dogs and raccoon dogs, and cats have a homology with C. ohioensis and C. felis, respectively. Phylogenetic analysis of the DNA sequences showed that the dog and raccoon dog isolates were nested in a clade with other Cystoisospora spp. including C. ohioensis, C. belli, and C. orlovi. The cat isolate formed a sister group with C. felis that was a separate clade from the dog and raccoon dog group. We report sequence variation in these Cystoisospora sequences and have identified raccoon dogs as another carnivore host for Cystoisospora spp. infecting dogs.     

Characterization of monoclonal antibodies against canine P-selectin glycoprotein ligand-1 (PSGL-1)

Thirteen different monoclonal antibodies against canine P-selectin glycoprotein ligand-1 (cPSGL-1) were obtained by immunization of rats with cells of a canine lymphoma cell line (Ema). O-sialoglycoprotein endopeptidase treatment of Ema cells showed that all of these antibodies recognized O-glycosylated peptides of canine PSGL-1. Experiments using deletion or point mutants of cPSGL-1 indicated that these antibodies could be categorized into several groups based on their cPSGL-1 recognition characteristics. These anti-cPSGL-1 monoclonal antibodies will be useful for analysis of the canine P-selectin and PSGL-1 system.     

Characterization of canine BCL6 cDNA and detection systems for its protein expression

BCL6 is known to be a key molecule in germinal center (GC) formation of lymph nodes, and its expression profiles have been implicated in the prognosis of diffuse large B-cell lymphoma in humans. The present study was carried out to characterize canine BCL6 cDNA and to indicate the technical methods for detection of the BCL6 protein in dog tissues. The deduced amino acid sequence of canine BCL6 showed close homology to that of human BCL6 (96.3%), especially in the zinc-finger motifs and POZ (poxvirus and zinc finger) domain with complete identity. Immunoblot analysis of a canine lymph node with an anti-human BCL6 monoclonal antibody revealed a band of 80 kDa. Immunohistochemical staining using the same antibody produced positive reactions in the cells exclusively localized in the GC of a canine lymph node. This study will be useful for the molecular classification of canine B-cell lymphomas with different prognoses.     

Aluminum oxyhydroxide polymorphs and some micromorphological characteristics in sclerotium grains

The presence of aluminum oxyhydroxide and its micromorphological characteristics in sclerotium grains were examined by SEM, TEM, and EDX analyses. A spherical fraction with a diameter of 100 nm, which contained C and Al as the predominant elements, was recognized as the unit particle of the matrix of sclerotium grains. The Al polymorphs and the Si spherical structures found in the ignited grains were assumed to correspond to boehmite and opaline silica particles, respectively. SEMI analysis revealed the presence of a needle ball structure inside the hollow part of sclerotium grains. The relationship between the inner structure of sclerotium and biochemical processes of the host fungi was discussed.     

Changes in radiocesium concentration in a blueberry (Vaccinium virgatum Aiton) orchard resulting from radioactive fallout

We investigated changes in the radiocesium concentration in a blueberry (Vaccinium virgatum Aiton) orchard in Fukushima prefecture to clarify the radiocesium contamination for 3 years after the Tokyo Electric Power Company’s Fukushima Daiichi nuclear power plant accident occurred in mid-March 2011. In the aboveground part of blueberry bushes, the concentrations of radiocesium in branches that were directly affected by radioactive fallout were the highest among the samples investigated, and the concentrations in fruits were the lowest. The concentration of radiocesium decreased exponentially in the fruits and leaves over 3 years. The radiocesium concentrations in soils were higher in the surface layer. The amount of radiocesium in organic matter on the soil surface under the canopy greatly decreased from 2011 to 2012. The transfer factor of the radiocesium from soil to blueberry fruits decreased annually. These results suggest that radiocesium adhered directly to the aboveground organs of blueberry bushes in March 2011, and it is possible that the accumulation of radiocesium in fruits is mainly due to radiocesium transfer from the branches and trunk to fruits for several years after the nuclear power plant accident.