Simultaneous evaluation of diagnostic marker utility for enzootic bovine leukosis.

BACKGROUND: Enzootic bovine leukosis (EBL) is a disease of cattle caused by bovine leukemia virus (BLV). More than 60% of BLV-infected cattle remain subclinical and are thus referred to as aleukemic (AL) cattle. Approximately 30% of infected cattle show a relatively stable increase in the number of B lymphocytes; these cattle are termed persistent lymphocytosis (PL) cattle. A small percentage of infected cattle develop BLV-induced B cell lymphoma (EBL) and are called EBL cattle. Due to the increase in the number of BLV-infected cattle, the number of EBL cattle has featured a corresponding increase over recent years in Japan. Several diagnostic criteria for EBL (e.g., enlarged superficial lymph nodes, protrusion of the eye, increased peripheral blood lymphocyte, etc.) are used for on-farm diagnosis and antemortem tests at slaughterhouses. Since the slaughter of EBL cattle for human consumption is not allowed, on-farm detection of EBL cattle is important for reducing the economic loss incurred by farms. Therefore, establishing new diagnostic markers to improve the efficiency and accuracy of the antemortem detection of EBL cattle is a critical, unmet need. To simultaneously evaluate the utility of candidate markers, this study measured the values of each marker using the blood samples of 687 cattle with various clinical statuses of BLV infection (EBL, PL, AL and non-infected cattle). RESULTS: Sensitivity (Se) and specificity (Sp) were highest for the serum thymidine kinase (TK) followed by the serum lactate dehydrogenase (LDH) isozyme 2. The number of peripheral blood lymphocytes and proviral load in peripheral blood had the lowest Se and Sp. The values of all markers other than TK were influenced by the sex of the tested cattle. CONCLUSIONS: Although tLDH and its isozymes (LDHs) may be influenced by the sex of the tested cattle, the high accuracy of TK and LDH2 as well as accessibility and simplicity of the protocol used to measure these enzymes recommend the utility of TK and LDHs for EBL cattle detection. Using these markers for screening followed by the application of existing diagnostic criteria may improve the efficiency and accuracy of EBL cattle detection on farms, thereby contributing to the reduction of economic losses in farms.

Fraction of bovine leukemia virus-infected dairy cattle developing enzootic bovine leukosis.

Enzootic bovine leucosis (EBL) is a transmissible disease caused by the bovine leukemia virus that is prevalent in cattle herds in many countries. Only a small fraction of infected animals develops clinical symptoms, such as malignant lymphosarcoma, after a long incubation period. In the present study, we aimed to determine the fraction of EBL-infected dairy cattle that develop lymphosarcoma and the length of the incubation period before clinical symptoms emerge. These parameters were determined by a mathematical modeling approach based on the maximum-likelihood estimation method, using the results of a nationwide serological survey of prevalence in cattle and passive surveillance records. The best-fit distribution to estimate the disease incubation period was determined to be the Weibull distribution, with a median and average incubation period of 7.0 years. The fraction of infected animals developing clinical disease was estimated to be 1.4% with a 95% confidence interval of 1.2-1.6%. The parameters estimated here contribute to an examination of efficient control strategies making quantitative evaluation available.

The role of neighboring infected cattle in bovine leukemia virus transmission risk.

A cohort study was conducted to evaluate the risk of bovine leukemia virus (BLV) transmission to uninfected cattle by adjacent infected cattle in 6 dairy farms. Animals were initially tested in 2010-2011 using a commercial ELISA kit. Uninfected cattle were repeatedly tested every 4 to 6 months until fall of 2012. The Cox proportional hazard model with frailty showed that uninfected cattle neighboring to infected cattle (n=53) had a significant higher risk of seroconversion than those without any infected neighbors (n=81) (hazard ratio: 12.4, P=0.001), implying that neighboring infected cattle were a significant risk factor for BLV transmission. This finding provides scientific support for animal health authorities and farmers to segregate infected cattle on farms to prevent spread of BLV.

Nationwide survey of bovine leukemia virus infection among dairy and beef breeding cattle in Japan from 2009-2011.

A nationwide survey of bovine leukemia virus (BLV) infection was conducted among dairy and beef breeding cattle in Japan from 2009-2011 using an enzyme-linked immunosorbent assay. Of a total of 20,835 cattle tested, 35.2% were seropositive for BLV and the animal type-level seroprevalences in dairy and beef breeding cattle were 40.9 and 28.7%, respectively. By the time animals were 1 year old, 21.0% of dairy and 13.7% of beef breeding cattle were considered infected. Our findings indicate that BLV is widespread among dairy and beef breeding cattle in Japan with the BLV seroprevalences approximately 10- and 4-fold higher, respectively, than previously reported for 1980-1982 in Japan.

The recent prevalence of bovine leukemia virus (BLV) infection among Japanese cattle.

A seroepidemiological survey of bovine leukemia virus (BLV) infection was conducted in Japan in 2007 using an enzyme-linked immunosorbent assay (ELISA) and an agar gel immunodiffusion (AGID) test. A total of 5420 cattle (dairy, 3966; breeding beef, 797; fattening beef, 657) from 209 farms in seven prefectures in Japan were tested. The overall prevalence of BLV infection was 28.6%. The prevalence of BLV infection in dairy cattle (34.7%) was higher than for both fattening beef cattle (7.9%) and breeding beef cattle (16.3%). Age-specific prevalence showed that BLV prevalence increased with age in all types of cattle and was notably different between dairy and beef cattle under 1 year of age. Among 207 farms, 141 herds (68.1%) had one or more positive animals. The proportion of these positive farms was significantly higher among dairy farms (79.1%) than among beef breeding farms (39.5%) and beef fattening farms (51.9%) (P<0.001). Dairy farms (40.5%) also showed a significantly higher within-herd prevalence than beef breeding (27.4%) and fattening (14.9%) farms (P=0.001). This study indicated that BLV is more widely spread in dairy cattle than in beef breeding cattle in Japan. Given the prevalence of BLV infection in dairy and beef cattle was 8- and 1.7-fold higher, respectively, than rates previously found in 1980-1982, BLV appears to be spreading particularly among the dairy cattle population during the last two decades. Further investigation is required to determine the risk factors necessary to control BLV infection that take into account the different farming practices that exist between dairy and beef sectors.

Estimation of the within-herd transmission parameter of bovine leukemia virus.

In this study, we estimated the transmission parameter of bovine leukemia virus (BLV) infection within herds by following up the serological status of cattle in beef and dairy herds. Two consecutive serological tests using ELISA were conducted for cattle at four beef breeding farms and nine dairy farms with an interval of 5 months. Assuming that sero-converted animals were newly infected, transmission parameters were estimated using a hierarchical Bayesian Poisson model. All tested herds had at least one positive animal at the first testing, but sero-prevalence considerably varied between herds. It is estimated that 0.62 animals were infected by one infected animal introduced into a fully susceptible population for both beef breeding and dairy farms during the 5-month interval. The 95% credible interval of the transmission parameter for the combined herds was 0.37-0.89 over this period. Asymptomatically infected animals with BLV can become the source of lifelong infection in herds. These estimated transmission parameters indicated that the early removal of infected animals with periodical tests could reduce the dissemination of BLV infection within herds.

Risk factors associated with within-herd transmission of bovine leukemia virus on dairy farms in Japan.

BACKGROUND: Although several attempts have been made to control enzootic bovine leukosis (EBL) at the local level, a nationwide control program has not been implemented in Japan, except for passive surveillance. Effective control of EBL requires that the transmission routes of bovine leukemia virus (BLV) infection should be identified and intercepted based on scientific evidence. In this cross-sectional study, we examined the risk factors associated with within-herd transmission of BLV on infected dairy farms in Japan. Blood samples taken from 30 randomly selected adult cows at each of 139 dairy farms were tested by enzyme-linked immunosorbent assay (ELISA). Information on herd management was collected using a structured questionnaire. RESULTS: Infected farms were defined as those with more than one ELISA-positive animal and accounted for 110 (79.1%) of the 139 farms in the study. Completed questionnaires obtained from 90 of these 110 farms were used for statistical analysis. Seroprevalence, which was defined as the proportions of animals that tested positive out of all animals tested on the farm, was 17.1%, 48.1%, and 68.5% for the 25th, 50th, and 75th percentiles, respectively. A mixed logistic regression analysis implicated a loose housing system, dehorning, and a large number of horseflies in summer as risk factors (coefficient = 0.71, 1.11, and 0.82; p = 0.03, < 0.01, and 0.01, respectively) and feeding of colostrum to newborn calves from their dams as a protective factor (coefficient = -1.11, p = 0.03) against within-farm transmission of BLV on infected farms. CONCLUSION: Control of EBL in infected dairy farms in Japan will be improved by focusing particularly on these risk and protective factors.

Assessment of the impact of cattle testing strategies on human exposure to BSE agents in Japan.

In Japan, cattle screening tests for BSE are conducted at slaughterhouses for surveillance purposes and as a meat safety measure, but the public health impacts of such testing and the subsequent removal of positive animals from the food chain have not been quantitatively assessed. We evaluated the influence of removing specified risk materials and the alternation of age limits for testing cattle at the slaughterhouse on human exposure to the BSE agent in Japan by constructing a probabilistic risk model. A stochastic model using Monte Carlo simulation was constructed in order to estimate the BSE infectivity destined for the food chain from a single BSE-infected animal at slaughter. The impact of different testing strategies and risk material removal were then compared. Murine intra-cerebral ID50 (m.i.c. ID50) units were used as units for BSE infectivity. Sensitivity analysis was conducted for key input variables by changing values within plausible ranges. The expected fraction of BSE-infected cattle presented for slaughter that would be detected by screening tests was 20%, even if all slaughtered cattle were tested. The removal of risk materials reduced the median value estimate of infectivity destined for human consumption by 95%. Cattle screening tests reduced the infectivity further, but reduction efficacy did not differ among the various testing strategies. Sensitivity analysis indicated that the characteristics of BSE infectivity accumulation during the incubation period, extension of the incubation period, and lowering the detection limit of screening tests had no significant impact on relative infectivity reduction, which remained stable irrespective of testing strategy or changes in these parameters. This study suggests that the impact of changing the age limit for testing cattle on beef safety is small, provided that the removal of risk materials is conducted properly.