Detection of anti-neutrophil cytoplasmic antibodies in dogs with immune-mediated inflammatory diseases.

Anti-neutrophil cytoplasmic antibody (ANCA) is a type of autoantibody against neutrophil cytoplasm. In veterinary medicine, few studies have reported the detection of ANCA in dogs, and most of these studies were performed in dogs with inflammatory bowel disease (IBD). The aim of this study was to evaluate whether ANCA is detected in dogs with immune-mediated inflammatory diseases (IMIDs) other than IBD. Serum samples were collected before treatment initiation from 40 client-owned dogs with various diseases and 22 healthy beagle dogs; the dogs were classified into two groups: IMID group (n = 16) and control group (n = 46). ANCA was detected using the indirect fluorescent antibody test. Of the 16 dogs in the IMID group, 13 (81.3%) tested positive for ANCA. In contrast, of the 46 dogs in the control group, 13 (28.03%) tested positive for ANCA. Moreover, a significant association between ANCA positivity and IMIDs was identified in the IMID group than in the control group (P = 0.0003). In the control group, however, dogs with bacterial infection showed a relatively high rate of ANCA positivity. Therefore, ANCA positivity was observed in dogs with both IMIDs and bacterial infection. This suggested that ANCA positivity is associated with diseases accompanied by neutrophil activation and infiltration.

First detection of porcine circovirus type 3 in Japan.

Porcine circovirus associated diseases (PCVAD) have multiple manifestations that have been attributed to porcine circovirus type 2 (PCV2). Recently, a novel porcine circovirus, PCV type 3 (PCV3), was identified in pigs with systemic inflammation of unknown etiology. In this study, we tried to detect the PCV3 genome in tissue samples collected from Japanese pig herds in 2016. The PCV3 genome was detected by PCR in 7 of 73 samples. The homology between each Japanese strain was 99.5% for the full-length sequence and 98.9 to 99.2% for the open reading frame 2. These results suggest that PCV3 has already invaded Japanese pig farms.

Prevalence of swine Torque teno virus genogroups 1 and 2 in Japanese swine with suspected post-weaning multisystemic wasting syndrome and porcine respiratory disease complex.

Torque teno virus (TTV) was first isolated from a human hepatitis patient in 1997. TTV was also identified in several animals, including pigs, cattle, sheep, cats and dogs. In this study, we analysed the prevalence of swine TTV genogroups 1 (TTV1) and 2 (TTV2) in Japanese swine populations with suspected post-weaning multisystemic wasting syndrome and porcine respiratory disease by using a nested polymerase chain reaction method. Of 153 serum samples from 16 different herds in Japan, TTV1 was detected in 46 samples (30%), TTV2 in 47 samples (31%) and both in 15 samples (10%). There was no significant difference in the detection rate among geographical regions. The overall prevalence rate of TTV genogroups was significantly lower in < or = 30-day-old pigs (11%) compared to that in older age groups (54-82%). These results suggest that swine TTV may be widespread in post-weaning pigs and could play aetiological roles in pig diseases in Japan. This is the first report on the prevalence of swine TTV in Japan.

Multiplex PCR and multiplex RT-PCR for inclusive detection of major swine DNA and RNA viruses in pigs with multiple infections.

Multiplex PCR and multiplex RT-PCR were developed to identify nine viruses in pigs with multiple infections. These viruses are: porcine circovirus type 2 (PCV2), suid herpesvirus 1, porcine parvovirus (PPV), porcine reproductive and respiratory syndrome virus (PRRSV), Japanese encephalitis virus, porcine rotavirus A (PoRV-A), porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis virus (TGEV), and Getah virus. These methods were shown to be high specificity and sensitivity. In the clinical application, a total of 75 field samples were examined by our methods and previously reported methods for PCV2, PRRSV, TGEV, and PEDV. As a result, the detection rates of our multiplex PCR and multiplex RT-PCR were higher than those of the previously reported methods. Furthermore, it was confirmed that 24 PCV2 positive samples were co-infected with other viruses, 11 with PRRSV, 10 with PPV, 2 with PoRV-A, and 1 with TGEV by a combination of multiplex PCR and multiplex RT-PCR. PPV and PoRV-A were newly detected by multiplex PCR and multiplex RT-PCR. These results suggest that the combination of our multiplex PCR and multiplex RT-PCR is useful for rapid and accurate identification of nine major pathogenic viruses in pigs with multiple infections.

Yeast mutant with efficient secretion identified by a novel secretory reporter, Cluc.

Yeast is an important host for the production of pharmaceutical or industrial proteins by virtue of its genetic information and easy handling. A number of heterologous proteins have been produced and purified from yeast cell cultures as secreted forms. Here, we describe a novel screening system of Saccharomyces cerevisiae and its application to improve the secretion efficiency of yeast. In our system, a natural secretory luciferase from Cypridina noctiluca is used as a reporter enzyme. The accumulation of enzymatically active luciferase in culture medium makes it possible to screen many samples simultaneously in a simple and sensitive assay. Using this system, we have discovered that the deletion mutant of MON2, which encoded a scaffold protein for vesicle formation located at the late Golgi, secreted luciferase highly efficiently to the extracellular space. Thus, we conclude that this new reporter assay is useful for the improvement and screening of yeast secretory strains.