Prior exposure to non-pathogenic calicivirus RCV-A1 reduces both infection rate and mortality from rabbit haemorrhagic disease in a population of wild rabbits in Australia.

Mortality caused by rabbit haemorrhagic disease virus (RHDV) in wild rabbits is reduced in parts of Australia where the related, non-pathogenic calicivirus RCV-A1 is endemic. Laboratory experiments previously showed that prior infection with RCV-A1 enabled rabbits to better withstand subsequent infection with highly virulent RHDV, and this was assumed to explain higher survival. Here, we analyse serological data from the field suggesting that reduced mortality rates among wild rabbits may also result from rabbits previously infected with RCV-A1 having a reduced likelihood of RHDV infection. We discuss the possible mechanisms underlying this finding and its implications. The methods we describe for analysing field data gave far greater insights into epidemiological processes and virus interactions than gained from reporting basic seroprevalence rates alone.

Rabbit haemorrhagic disease: are Australian rabbits (Oryctolagus cuniculus) evolving resistance to infection with Czech CAPM 351 RHDV?

Rabbit haemorrhagic disease is a major tool for the management of introduced, wild rabbits in Australia. However, new evidence suggests that rabbits may be developing resistance to the disease. Rabbits sourced from wild populations in central and southeastern Australia, and domestic rabbits for comparison, were experimentally challenged with a low 60 ID50 oral dose of commercially available Czech CAPM 351 virus - the original strain released in Australia. Levels of resistance to infection were generally higher than for unselected domestic rabbits and also differed (0-73% infection rates) between wild populations. Resistance was lower in populations from cooler, wetter regions and also low in arid regions with the highest resistance seen within zones of moderate rainfall. These findings suggest the external influences of non-pathogenic calicivirus in cooler, wetter areas and poor recruitment in arid populations may influence the development rate of resistance in Australia.

Rabbit haemorrhagic disease: field epidemiology and the management of wild rabbit populations.

Rabbit haemorrhagic disease (RHD) has become established in wild rabbit populations throughout Western Europe, Australia and New Zealand. The abundance of wild rabbits has been significantly reduced, particularly in drier areas of southern Spain, inland Australia and South Island New Zealand. A detailed knowledge of the epidemiology of RHD is essential for the management of the disease in natural rabbit populations, either to rebuild or to control populations. When RHD first spread among naive wild rabbits, epidemiological studies provided unique information on the rate of spread, the possible role of insect vectors in transmission, and the correlation between the impact of disease on populations and climatic variables. Current research shows a consistent pattern of epidemiology between Europe and Australasia. Typically, the most severe epizootics of RHD occur among young sub-adult rabbits which have lost age-related resilience and maternal antibodies. However, the timing of these outbreaks reflects climatic variables that determine the breeding season of the rabbits and the periods when RHD virus (RDHV) is most likely to persist and spread. Further factors that may complicate epidemiology include the possibility that non-pathogenic RHDV-like viruses are present in natural rabbit populations. Additionally, the question of how the virus persists from year to year remains unresolved; persistence in carrier rabbits is a possibility. Understanding of the epidemiology of RHD is now sufficiently advanced to consider the possibility of manipulating rabbit populations to alter the epidemiological pattern of RHD and thereby maximise or minimise the mortality caused by the disease. Altering the epidemiology of RHD in this manner would assist the management of wild rabbit populations either for conservation or pest control purposes.

Evidence for insect transmission of rabbit haemorrhagic disease virus.

The spread of rabbit haemorrhagic disease (RHD) virus from quarantine on Wardang Island to mainland Australia in 1995 suggested that insects could be potential vectors. Field observations and laboratory experiments were conducted to address aspects of this hypothesis. Firstly, the variation in insect populations on the island during the field trials was examined. There was approximately a 1,000-fold increase in the number of bushflies, Musca vetustissima, shortly before the spread of the virus. Secondly, M. vetustissima were tested in the laboratory as potential vectors of RHD virus, and it was demonstrated that disease could be transmitted between rabbits by flies. Finally, 13 of 16 insect samples, collected from Wardang Island and from several sites on the mainland following the spread of virus off the island, were positive for the presence of RHD virus by a specific polymerase chain reaction (PCR). Only one sample contained sufficient infectious virus to kill a susceptible rabbit. These data, combined with previously published information on fly biology, suggested that flies, particularly bushflies, may be involved in the transmission of RHD virus. Other possible routes of spread were not assessed in this study.

Use of ELISAs in field studies of rabbit haemorrhagic disease (RHD) in Australia.

ELISA techniques developed for the veterinary diagnosis of Rabbit Haemorrhagic Disease (RHD) in domestic rabbits were used for studying the epidemiology of RHD in Australian wild rabbits. The combination of ELISA techniques that distinguished IgA, IgG and IgM antibody responses and a longitudinal data set, mainly based on capture-mark-recapture of rabbits, provided a reliable basis for interpreting serology and set the criteria used to classify rabbits' immunological status. Importantly, young with maternal antibodies, immune rabbits and rabbits apparently re-exposed to RHD were readily separated. Three outbreaks of RHD occurred in 1996-7. The timing of RHD outbreaks was mainly driven by recruitment of young rabbits that generally contracted RHD after they lost their maternally derived immunity. Young that lost maternal antibodies in summer were not immediately infected, apparently because transmission of RHDV slows at that time, but contracted RHD in the autumn when conditions were again suitable for disease spread.

Sequence analysis of rabbit haemorrhagic disease virus (RHDV) in Australia: alterations after its release.

Liver samples from rabbits killed by RHDV, collected from five States in Australia in 1996 and 1997 were analysed by RT-PCR. A 398 bp fragment of the capsid protein (VP60) gene was amplified by PCR and directly sequenced. The alignment of the nucleotide and amino acid sequences and their comparison with the original strain of the virus released in Australia indicated genetic changes after two years have been small with 98.2% to 100% identity. The constructed phylogenetic tree suggests slight differences in nucleotide substitutions in various States but there is no clear evidence of clustering of sequences according to their geographic origin. In practical terms, sequencing of viral RNA provides a means of testing the efficacy of further releases and subsequent spread of the virus if such a strategy is employed as a means of enhancing RHD as a biological control of the wild rabbit in Australia.

Notes on the life-history of the rabbit flea Caenopsylla laptevi ibera Beaucournu & Marquez, 1987 (Siphonaptera: Ceratophyllidae) in eastern Spain.

The rabbit flea Caenopsylla laptevi ibera occurs in arid environments of central and eastern Spain. Although the fleas breed during the coolest, most humid part of the year, the larvae survive and grow in sand at only 50-60% relative humidity. At 22 degrees C and 80% relative humidity eggs hatch in six days and the cocoon stage is reached 10-11 days after hatching. Female fleas emerge from pupation at about 17 days after cocoon spinning; males emerge a little later at a mean of 20 days. Adult fleas are mainly found on the host Oryctolagus cuniculus. Measurements of burrow microclimate confirmed that in south-eastern Spain burrow humidity was adequate for the development of C. I. ibera larvae over most of the year. However, breeding may be restricted for at least part of the year, as the larvae of C. I. ibera apparently cannot complete development at 25 degrees C or above. In the laboratory, fleas can enter a prolonged quiescent period while in the cocoon. This is possibly a facultative, pre-pupal diapause and the likely mechanism that accounts for the disappearance of adult fleas from the field by spring and their reappearance each autumn.

Field evidence for mechanical transmission of rabbit haemorrhagic disease virus (RHDV) by flies (Diptera:Calliphoridae) among wild rabbits in Australia.

Field collected flies were screened for the presence of rabbit haemorrhagic disease virus (RHDV) by applying reverse transcriptase PCR (RT-PCR) in which primers specific to the capsid protein of the virus were used. The virus was detected in flies from locations where rabbit haemorrhagic disease (RHD) was reported and also soon after the release of RHDV in a 'clean' area. Oral and/or anal excretions of flies (flyspots) were found to contain viable virus and oral inoculation of rabbits revealed that a single flyspot was able to cause RHD. We conclude that flyspots are a major potential source of the virus for oral or conjunctival transmission of the virus to rabbits.