The effectiveness of parallel gamma-interferon testing in New Zealand's bovine tuberculosis eradication programme.

In bovine tuberculosis (bTB) eradication programmes, especially where prevalence is low, sensitivity of testing in infected herds must be maximised to reduce the possibility of recrudescence of prior infection and the risk to other herds via animal movement. The gamma-interferon (γ-IFN) assay applied in parallel with intradermal tuberculin testing has been shown to increase test sensitivity. The aim of this work was to substantiate this effect in the field. A retrospective observational study was conducted on 239 New Zealand cattle breeding and dairy herds with bTB infection between 1 July 2011 and 1 September 2015 to evaluate the outcomes of new policy introduced in 2011. The investigation defined the number and proportion of reactors (animals testing positive and slaughtered) found with lesions of bTB in intradermal caudal fold testing (CFT) and parallel γ-IFN testing, at the breakdown test or first whole herd test after breakdown, WHT(1), and at the final or projected final whole herd test, WHT(F). Parallel γ-IFN testing was used in 26.8% of the 239 herds at WHT(1), and 430 animals in 49 herds were deemed reactors. One hundred and sixty (37.2%) of these reactors from 32 herds were found to have bTB lesions, despite having been negative to caudal fold testing. These 160 infected animals accounted for 29.6% of all infection found at WHT(1). At WHT(F), parallel γ-IFN testing was conducted on 93 herds and detected a total of 122 reactors in 49 herds, in addition to those found by CFT. Twenty-one of these reactors, from 13 herds, had bTB lesions at slaughter, accounting for 67.7% of all reactors found with bTB at WHT(F). Eleven of these 13 herds would have had their movement restrictions revoked based on a negative herd CFT alone, and could potentially have caused outward transmission of bTB to other herds, as well as experiencing recrudescent breakdowns. We conclude that γ-IFN testing in infected herds, in parallel with intradermal tuberculin testing, is a valuable tool in a bTB eradication programme, as it enables higher test sensitivity at both herd and animal level. The use of the γ-IFN test over a risk cohort early in a breakdown assists in removal of early infection and some cases of anergy to intradermal tuberculin testing. Parallel γ-IFN with compulsory slaughter of reactors should be considered in breeding and dairy herds in conjunction with tuberculin testing before movement control is revoked, and will assist in achieving TB freedom on a herd level and nationally.

Epidemiology and control of Mycobacterium bovis infection in brushtail possums (Trichosurus vulpecula), the primary wildlife host of bovine tuberculosis in New Zealand.

The introduced Australian brushtail possum (Trichosurus vulpecula) is a maintenance host for bovine tuberculosis (TB) in New Zealand and plays a central role in the TB problem in this country. The TB-possum problem emerged in the late 1960s, and intensive lethal control of possums is now used to reduce densities to low levels over 8 million ha of the country. This review summarises what is currently known about the pathogenesis and epidemiology of TB in possums, and how the disease responds to possum control. TB in possums is a highly lethal disease, with most possums likely to die within 6 months of becoming infected. The mechanisms of transmission between possums remain unclear, but appear to require some form of close contact or proximity. At large geographic scales, TB prevalence in possum populations is usually low (1-5%), but local prevalence can sometimes reach 60%. Intensive, systematic and uniform population control has been highly effective in breaking the TB cycle in possum populations, and where that control has been sustained for many years the prevalence of TB is now zero or near zero. Although some uncertainties remain, local eradication of TB from possums appears to be straightforward, given that TB managers now have the ability to reduce possum numbers to near zero levels and to maintain them at those levels for extended periods where required. We conclude that, although far from complete, the current understanding of TB-possum epidemiology, and the current management strategies and tactics, are sufficient to achieve local, regional, and even national disease eradication from possums in New Zealand.

Epidemiology, diagnostics, and management of tuberculosis in domestic cattle and deer in New Zealand in the face of a wildlife reservoir.

The control of tuberculosis (TB) in cattle and farmed deer in New Zealand has been greatly influenced by the existence of a wildlife reservoir of Mycobacterium bovis infection, principally the Australian brushtail possum (Trichosurus vulpecula). The reduction in possum numbers in areas with endemic M. bovis infection through vigorous vector control operations has been a major contributor to the marked reduction in the number of infected cattle and farmed deer herds in the past two decades. Management of TB in cattle and farmed deer in New Zealand has involved a combination of vector control, regionalisation of diagnostic testing of cattle and deer herds, abattoir surveillance and movement control from vector risk areas. Accurate diagnosis of infected cattle and deer has been a crucial component in the control programme. As the control programme has evolved, test requirements have changed and new tests have been introduced or test interpretations modified. Subspecific strain typing of M. bovis isolates has proved to be a valuable component in the epidemiological investigation of herd breakdowns to identify whether the source of infection was domestic livestock or wildlife. New initiatives will include the use of improved models for analysing diagnostic test data and characterising disease outbreaks leading to faster elimination of infection from herds. The introduction of the National Animal Identification Tracing programme will allow better risk profiling of individual herds and more reliable tracing of animal movements. TB in cattle and farmed deer in New Zealand can only be controlled by eliminating the disease in both domestic livestock and the wildlife reservoir.

Detection of bovine herpesvirus type 4 antibodies and bovine lymphotropic herpesvirus in New Zealand dairy cows.

AIM: To detect the presence of bovine herpesvirus (BoHV) type 4 in New Zealand dairy cows with clinical metritis. METHODS: Serum samples taken from 92 dairy cows with clinical metritis, each from a different farm, were tested for the presence of antibodies against BoHV-4 using a commercially available, indirect ELISA. Peripheral blood mononuclear cells (PBMC) were collected from 10 BoHV-4 seropositive cows, and PBMC were examined by a pan-herpesvirus nested PCR to detect herpesvirus. PCR products were sequenced directly and a proportion of the PCR products were cloned and sequenced to identify the virus present. RESULTS: Antibodies to BoHV-4 were detected in 23/92 (25%) serum samples. The pan-herpesvirus PCR was positive in 8/10 PBMC samples. Cloning and sequencing identified that all of the eight PCR-positive PBMC contained bovine lymphotropic herpesvirus (BLHV); no BoHV-4 DNA was detected. CONCLUSIONS: This study reports the finding of the presence of apparent antibodies to BoHV-4, and BLHV DNA in New Zealand dairy cows affected by metritis. CLINICAL RELEVANCE: Bovine herpesvirus type 4 and BLHV are reported to have the potential to cause reproduction failure in cows. This is the first report of apparent BoHV-4 antibodies, and BLHV in New Zealand. The importance and epidemiology of these viruses in cattle in New Zealand requires further investigation.

Oral administration of Lactobacillus brevis KB290 to mice alleviates clinical symptoms following influenza virus infection.

UNLABELLED: Lactobacillus brevis KB290 (KB290), isolated from a traditional Japanese pickle 'Suguki', has been reported to have immunomodulatory effects. We investigated whether oral administration of KB290 has protective effects against influenza virus (IFV) infection in mice. After 14 days of administration of lyophilized KB290 suspended in phosphate-buffered saline by oral gavage, BALB/c mice were intranasally infected with 2 × MLD50 (50% mouse lethal dose) of IFV A/PR/8/34 (H1N1). Prophylactically administered KB290 significantly alleviated the loss of body weight and the deterioration in observational physical conditions induced by the infection. In addition, 7 days after infection, the levels of IFV-specific immunoglobulin (Ig)A in bronchoalveolar lavage fluid were significantly increased in mice fed KB290 compared with controls. Moreover, there was a significant elevation of serum interferon (IFN)-α in KB290 group mice, even at three and 7 days after infection, despite the administration of KB290 being stopped before IFV infection. Our results demonstrated that oral administration of KB290 before infection could alleviate IFV-induced clinical symptoms. Alleviation of clinical symptoms by KB290 consumption may have been induced by long-lasting enhancement of IFN-α production and the augmentation of IFV-specific IgA production. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrated that oral administration of Lactobacillus brevis KB290 (KB290), a probiotic strain derived from a Japanese traditional pickle, could protect against influenza virus (IFV) infection in mice. Our results demonstrated that continual intake of KB290 for 14 days prior to IFV infection alleviated clinical symptoms such as loss of body weight and deterioration in observational physical conditions induced by the infection. The beneficial effects of KB290 consumption may have been elicited by the long-lasting enhancement of interferon-α production and the augmentation of IFV-specific immunoglobulin A production.

Overview of vaccination trials for control of tuberculosis in cattle, wildlife and humans.

Vaccination is a key strategy for control of tuberculosis (TB), and considerable progress has been made in the past 5 years to develop improved vaccines for humans and animals, differentiate vaccinated animals from those infected with Mycobacterium bovis and deliver vaccines to wildlife. Studies have moved from testing vaccines in small animal models to clinical trials in humans and from experimental challenge studies in cattle and wildlife to evaluation of vaccines in the field. Candidate vaccines undergoing testing in humans include live mycobacterial vaccines to replace bacille Calmette Guérin (BCG), subunit vaccines (virus vector or protein) to boost BCG and therapeutic vaccines used as an adjunct to chemotherapy. In cattle, a number of diagnostic tests have been developed and successfully tested for differentiating infected from vaccinated animals, which will facilitate the use of BCG vaccine in cattle. Encouraging results have been obtained from recent field trials in cattle using BCG vaccine to protect against natural exposure to M. bovis. To date, no subunit TB vaccines have induced improved protection compared with that for BCG, but prime-boost combinations of BCG with DNA, protein or virus-vectored vaccines have induced better protection than BCG vaccine alone. Development of an oral bait BCG formulation has demonstrated the practicality of delivering TB vaccines to wildlife. Oral BCG preparations have induced protection against experimental challenge of M. bovis in possums, badgers, wild boar and white-tailed deer and against natural exposure to M. bovis in possums. Recent progress in TB vaccine development has provided much impetus for their future use.

Progress in the development of vaccines against rumen methanogens.

Vaccination against rumen methanogens offers a practical approach to reduce methane emissions in livestock, particularly ruminants grazing on pasture. Although successful vaccination strategies have been reported for reducing the activity of the rumen-dwelling organism Streptococcus bovis in sheep and S. bovis and Lactobacillus spp. in cattle, earlier approaches using vaccines based on whole methanogen cells to reduce methane production in sheep have produced less promising results. An anti-methanogen vaccine will need to have broad specificity against methanogens commonly found in the rumen and induce antibody in saliva resulting in delivery of sufficiently high levels of antibodies to the rumen to reduce methanogen activity. Our approach has focussed on identifying surface and membrane-associated proteins that are conserved across a range of rumen methanogens. The identification of potential vaccine antigens has been assisted by recent advances in the knowledge of rumen methanogen genomes. Methanogen surface proteins have been shown to be immunogenic in ruminants and vaccination of sheep with these proteins induced specific antibody responses in saliva and rumen contents. Current studies are directed towards identifying key candidate antigens and investigating the level and types of salivary antibodies produced in sheep and cattle vaccinated with methanogen proteins, stability of antibodies in the rumen and their impact on rumen microbial populations. In addition, there is a need to identify adjuvants that stimulate high levels of salivary antibody and are suitable for formulating with protein antigens to produce a low-cost and effective vaccine.

Development and evaluation of an enzyme-linked immunosorbent assay for use in the detection of bovine tuberculosis in cattle.

As a consequence of continued spillover of Mycobacterium bovis into cattle from wildlife reservoirs and increased globalization of cattle trade with associated transmission risks, new approaches such as vaccination and novel testing algorithms are seriously being considered by regulatory agencies for the control of bovine tuberculosis. Serologic tests offer opportunities for identification of M. bovis-infected animals not afforded by current diagnostic techniques. The present study describes assay development and field assessment of a new commercial enzyme-linked immunosorbent assay (ELISA) that detects antibody to M. bovis antigens MPB83 and MPB70 in infected cattle. Pertinent findings include the following: specific antibody responses were detected at ∼90 to 100 days after experimental M. bovis challenge, minimal cross-reactive responses were elicited by infection/sensitization with nontuberculous Mycobacterium spp., and the apparent sensitivity and specificity of the ELISA with naturally infected cattle were 63% and 98%, respectively, with sensitivity improving as disease severity increased. The ELISA also detected infected animals missed by the routine tuberculin skin test, and antibody was detectable in bulk tank milk samples from M. bovis-infected dairy herds. A high-throughput ELISA could be adapted as a movement, border, or slaughter surveillance test, as well as a supplemental test to tuberculin skin testing.

Newly attenuated Mycobacterium bovis mutants as vaccines against bovine tuberculosis, particularly for possums.

Bovine tuberculosis costs New Zealand more than $80 million per year, mostly because extensive areas of the country are occupied by brushtail possums infected with Mycobacterium bovis. AgResearch has a major programme to produce new live tuberculosis vaccines that can be delivered to possums. Primary work involved development of molecular biological methods to enable genetic manipulation of M. bovis, including the production of random and specific mutants. Many avirulent mutants of M. bovis have been produced and their vaccine efficacy has been compared to BCG in guinea pigs. Selected mutants that perform at least as well as BCG are retested in guinea pigs using an extended vaccination protocol in which animals are pre-sensitized to environmental mycobacteria to mimic natural exposure. Ten candidate vaccines that have induced good protection in guinea pigs have been subsequently tested as vaccines in possums. While the protective efficacy of an M. bovis mutant inoculated into guinea pigs reliably indicated that some protection would be induced in possums, the most protective mutant in guinea pigs was different from that in possums. This illustrates the importance of testing in the target species as part of new vaccine development. An important outcome of this work was the identification of an operon in M. bovis whose inactivation produced an avirulent M. bovis vaccine candidate that was better than BCG in protecting possums from experimental tuberculosis. Allelic exchange methods are now being used to produce vaccine strains with multiple specific mutations to improve safety and immunological characteristics.

Bovine tuberculosis: a review of current and emerging diagnostic techniques in view of their relevance for disease control and eradication.

Existing strategies for long-term bovine tuberculosis (bTB) control/eradication campaigns are being reconsidered in many countries because of the development of new testing technologies, increased global trade, continued struggle with wildlife reservoirs of bTB, redistribution of international trading partners/agreements, and emerging financial and animal welfare constraints on herd depopulation. Changes under consideration or newly implemented include additional control measures to limit risks with imported animals, enhanced programs to mitigate wildlife reservoir risks, re-evaluation of options to manage bTB-affected herds/regions, modernization of regulatory framework(s) to re-focus control efforts, and consideration of emerging testing technologies (i.e. improved or new tests) for use in bTB control/eradication programs. Traditional slaughter surveillance and test/removal strategies will likely be augmented by incorporation of new technologies and more targeted control efforts. The present review provides an overview of current and emerging bTB testing strategies/tools and a vision for incorporation of emerging technologies into the current control/eradication programs.