Yumi Kirino

Evaluation of the natural perinatal transmission of bovine leukaemia virus

The perinatal transmission of bovine leukaemia virus (BLV) plays a critical role in the spread and persistence of BLV infection in cattle herds. The purpose of this study was to examine the frequency of perinatal infections in an area in Japan and investigate some risk factors associated with infection. Altogether, 129 calves born to BLV-infected cows in a herd in Japan were tested for infection immediately after birth and again at one month of age using nested PCR. Twenty-four calves (18.6 per cent) were infected with BLV, of which 14 (10.8 per cent) and 10 (7.7 per cent) calves were infected via the transplacental and the birth canal routes, respectively. Maternal viral loads, breed, the presence or absence of assistance during parturition and the number of births per dam were evaluated to investigate risk factors associated with infection. Maternal viral load was significantly correlated with the frequency of perinatal infection, and more than 40 per cent of newborn calves born to dams with high viral loads were infected with BLV. The results of this study could contribute towards developing effective eradication programmes by providing necessary data for replacement of breeding cow in the field.

Horizontal transmission and phylogenetic analysis of bovine leukemia virus in two districts of Miyazaki, Japan

Horizontal transmission is recognized as a major infection route for bovine leukemia virus (BLV), and cattle with high viral loads are considered to be a major infectious source in a herd. However, a correlation between viral loads and the risk of infection has been insufficient to use as a foundation for BLV control strategies. In this report, we examined the epidemiology of BLV infection and the infectious source in a local area. In 2013–2014, BLV infection was investigated in 1,823 cattle from 117 farms in two adjacent districts, Miyazaki, Japan. Seropositive samples for BLV were detected with 88 cattle and in 14 farms. Phylogenetic analysis revealed that 94% of the isolates clustered into genotype I and the remaining isolate into genotype III. Among genotype I, genetically distinct strains were spread at each farm, and cattle infected with less than 3 copies/100 cells did not transmit BLV to other cattle for more than thirty months. This is the first report of concrete data of viral load in relation to viral horizontal transmission under the field condition. The data facilitate farmers and veterinarians understanding the status of BLV infected cattle. This research contributes to BLV infection control and the development of effective BLV eradication programs.

Identification of O serotypes, genotypes, and virulotypes of Shiga toxin-producing Escherichia coli isolates, including non-O157 from beef cattle in Japan.

Bovines are recognized as an important reservoir of Shiga toxin-producing Escherichia coli (STEC). Although STEC strains are significant foodborne pathogens, not all of the STEC held by cattle are pathogenic, and which type of STEC that will become epidemic in humans is unpredictable. Information about the prevalence of serotype and virulence gene distribution in beef cattle is insufficient to develop monitoring and controlling activities for a food safety and security program. Thus, this study investigated the prevalence of O157 and non-O157 STEC in Japanese beef cattle and characterized the isolates by the type of O antigen and several virulence markers to help predict the pathogenicity. In this study, 64.2% (176 of 274) of enrichment cultures of fecal samples collected from an abattoir and farms were stx1 and/or stx2 positive by PCR. STEC strains were isolated from 22.1% (39 of 176) of the positive fecal samples, and these isolates represented 17 types of O antigen (O1, O2 or O50, O5, O8, O55, O84, O91, O109, O113, O136, O150, O156, O157, O163, O168, O174, and O177). Two selective media targeting major STEC groups, cefixime-tellurite sorbitol MacConkey agar and CHROMagar O26/O157, allowed isolation of a variety of STEC strains. The most frequently isolated STEC was O113 (8 of 39), which has previously been reported as a cause of foodborne infections. Although most of the O113 STEC isolated from infected patients possessed the enterohemolysin (hlyA) gene, none of the O113 STEC cattle isolates possessed the hlyA gene. The second most common isolate was O157 (6 of 39), and all these isolates contained common virulence factors, including eae, tir, lpf1, lpf2, and hlyA. This study shows the prevalence of O157 and non-O157 STEC in Japanese beef cattle and the relationship of O antigen and virulotypes of the isolates. This information may improve identification of the source of infection, developing surveillance programs or the current understanding of virulence factors of STEC infections.

Campylobacter and Salmonella are prevalent in broiler farms in Kyushu, Japan: results of a 2‐year distribution and circulation dynamics audit

To elucidate the distribution and circulation dynamics of Campylobacter and Salmonella in Japanese chicken broiler flocks.

Infection of a group of boar-hunting dogs with Paragonimus westermani in Miyazaki Prefecture, Japan.

A 5-year-old male mixed-bred boar-hunting dog with a Plott hound background weighing 23 kg was brought to the Veterinary Teaching Hospital, University of Miyazaki, in October 2002. The dog was diagnosed with active infection with the lung fluke Paragonimus westermani by serological testing and also by detection of parasite eggs in his feces. Subsequent examination of four other dogs working with this dog as a boar-hunting team revealed that all five dogs were infected with P. westermani.

A seroepidemiological survey for paragonimosis among boar-hunting dogs in central and southern Kyushu, Japan

A seroepidemiological survey for paragonimosis among boar-hunting dogs was conducted throughout central and southern Kyushu, Japan. Among a total of 224 sera collected from boar-hunting dogs kept by 38 owners, 147 (65.6%) were positive for IgG antibodies against Paragonimus westermani antigen by ELISA. In the 147 seropositive dogs, 83 (56.5%) dogs demonstrated high antibody titers (OD>0.700), which appeared to indicate active infection. There were no obvious differences noted in age, sex and the genetic background of the dogs. The seroprevalence was dependent on the feeding manner of the owners and was extremely variable (0-100%). The majority (34/38) of owners fed their dogs uncooked boar meat and/or allowed them free-access to the residues of hunted boars after dissection, resulting in dogs with seropositive results. The dogs that were never fed wild boar meat were entirely seronegative. These results clearly demonstrate that boar-hunting dogs play an important role as a definitive Paragonimus host and that wild boars serve as a reservoir host for the maintenance of the Paragonimus life-cycle, presumably P. westermani, in the mountainous areas of central and southern Kyushu, Japan.

Increase of Clostridium perfringens in association with Eimeria in haemorrhagic enteritis in Japanese beef cattle

A coprological survey with detailed clinical observation of naturally occurring haemorrhagic enteritis (HE) cases was conducted to understand the pathophysiology of HE by clarifying the infection status of Eimeria and enteropathogenic bacteria in cattle. Faecal samples from 55 cases of HE and 26 clinically normal animals were collected, and a quantitative examination of Eimeria and potential enteropathogenic bacteria was performed. The number of Eimeria species oocysts per gram of faeces (OPG) exceeded 10,000 in 69.1 per cent of HE cases with a maximum of 1,452,500 OPG and Eimeria zuernii was found to be overwhelmingly dominant. A significant increase in faecal coliform count was observed in HE cases compared with clinically normal animals. Among the animals shedding >10,000 OPG, 42.9 per cent showed a remarkable increase in Clostridium perfringens abundance (>104 CFU/g) in the faeces. In the cases with C. perfringens detected, its abundance was positively correlated with Eimeria OPG and high C. perfringens abundance was always accompanied by high Eimeria OPG. E. zuernii is likely to play a crucial role in massive multiplication of C. perfringens in HE in cattle.

Use of Direct LAMP Screening of Broiler Fecal Samples for Campylobacter jejuni and Campylobacter coli in the Positive Flock Identification Strategy

Rapid identification of Campylobacter-positive flocks before slaughter, following freezing and heat treatment for the Campylobacter-positive carcasses at the slaughterhouses is an effective control strategy against foodborne campylobacteriosis. We evaluated a loop-mediated isothermal amplification (LAMP) assay for the direct screening of naturally contaminated chicken cloacal swabs for C. jejuni/C. coli to compare this assay with conventional quantitative culture methods. In a comparison study of 165 broilers, the LAMP assay showed 82.8% (48/58 by conventional culture) sensitivity, 100% (107/107) specificity, 100% (48/48) positive predictive value (PPV), and 91.5% (107/117) negative predictive value (NPV). In a comparison of 55 flocks, LAMP showed 90.5% (19/21) sensitivity, 100% (34/34) specificity, 100% (19/19) PPV, and 94.4% (34/36) NPV. In the cumulative total of 28 farm-level comparisons, LAMP showed 100% (12/12) sensitivity, 100% (16/16) specificity, 100% (12/12) PPV, and 100% (16/16) NPV. The LAMP assay required less than 90 min from the arrival of the fecal samples to final results in the laboratory. This suggests that the LAMP assay will facilitate the identification of C. jejuni/C. coli-positive broiler flocks at the farm level or in slaughterhouses before slaughtering, which would make it an effective tool in preventing the spread of Campylobacter contamination.

Epidemiological Study and Control Trial of Taeniid Cestode Infection in Farm Dogs in Qinghai Province, China

ABSTRACT An epidemiological study and control trial were conducted to assess taeniid infection in farm dogs in Qinghai Province, China. To improve egg detection by fecal examination, a deworming step with praziquantel was incorporated into the sampling methodology. As a result, a marked increase in the number of egg-positive samples was observed in samples collected at 24 hr after deworming. Then, the fecal examination and barcoding of egg DNA were performed to assess the prevalence of taeniid species in dogs from Xinghai, Haiyan, Gangcha and Chengduo counties. Analysis of 277 dog feces revealed that taeniid cestodes, including Taenia spp. and Echinococcus granulosus, were highly prevalent in Xinghai (34.4%), but eggs were not found in Haiyan where a control trial on canine echinococcosis had been conducted 20 years previously. A control trial involving the administration of 5–10 mg/kg praziquantel to 90 farm dogs at 45-day intervals was conducted in Xinghai. The prevalence of taeniid cestodes in the dogs was reduced to 9.6% and 4.9% after one and two years, respectively, indicating that some dogs were not administered praziquantel properly. A questionnaire survey of farmers in Xinghai and Haiyan revealed that most farmers in Xinghai were not familiar with echinococcosis or the transmission route of the disease, while most farmers in Haiyan had a more thorough understanding of the disease. The findings implied that a program for educating local farmers would be important for efficiently controlling canine taeniid infection in the region.

Population structure of Escherichia coli O26 : H11 with recent and repeated stx2 acquisition in multiple lineages

A key virulence factor of enterohaemorrhagic Escherichia coli (EHEC) is the bacteriophage-encoded Shiga toxin (Stx). Stxs are classified into two types, Stx1 and Stx2, and Stx2-producing strains are thought to cause more severe infections than strains producing only Stx1. Although O26 : H11 is the second most prevalent EHEC following O157 : H7, the majority of O26 : H11 strains produce Stx1 alone. However, Stx2-producing O26 strains have increasingly been detected worldwide. Through a large-scale genome analysis, we present a global phylogenetic overview and evolutionary timescale for E. coli O26 : H11. The origin of O26 has been estimated to be 415 years ago. Sequence type 21C1 (ST21C1), one of the two sublineages of ST21, the most predominant O26 : H11 lineage worldwide, emerged 213 years ago from one of the three ST29 sublineages (ST29C2). The other ST21 lineage (ST21C2) emerged 95 years ago from ST21C1. Increases in population size occurred in the late 20th century for all of the O26 lineages, but most remarkably for ST21C2. Analysis of the distribution of stx2-positive strains revealed the recent and repeated acquisition of the stx2 gene in multiple lineages of O26, both in ST21 and ST29. Other major EHEC virulence genes, such as type III secretion system effector genes and plasmid-encoded virulence genes, were well conserved in ST21 compared to ST29. In addition, more antimicrobial-resistance genes have accumulated in the ST21C1 lineage. Although current attention is focused on several highly virulent ST29 clones that have acquired the stx2 gene, there is also a considerable risk that the ST21 lineage could yield highly virulent clones.