Real-time PCR as a surveillance tool for the detection of Trichinella infection in muscle samples from wildlife

Trichinella nematodes are the causative agent of trichinellosis, a meat-borne zoonosis acquired by consuming undercooked, infected meat. Although most human infections are sourced from the domestic environment, the majority of Trichinella parasites circulate in the natural environment in carnivorous and scavenging wildlife. Surveillance using reliable and accurate diagnostic tools to detect Trichinella parasites in wildlife hosts is necessary to evaluate the prevalence and risk of transmission from wildlife to humans. Real-time PCR assays have previously been developed for the detection of European Trichinella species in commercial pork and wild fox muscle samples. We have expanded on the use of real-time PCR in Trichinella detection by developing an improved extraction method and SYBR green assay that detects all known Trichinella species in muscle samples from a greater variety of wildlife. We simulated low-level Trichinella infections in wild pig, fox, saltwater crocodile, wild cat and a native Australian marsupial using Trichinella pseudospiralis or Trichinella papuae ethanol-fixed larvae. Trichinella-specific primers targeted a conserved region of the small subunit of the ribosomal RNA and were tested for specificity against host and other parasite genomic DNAs. The analytical sensitivity of the assay was at least 100 fg using pure genomic T. pseudospiralis DNA serially diluted in water. The diagnostic sensitivity of the assay was evaluated by spiking log of each host muscle with T. pseudospiralis or T. papuae larvae at representative infections of 1.0, 0.5 and 0.1 larvae per gram, and shown to detect larvae at the lowest infection rate. A field sample evaluation on naturally infected muscle samples of wild pigs and Tasmanian devils showed complete agreement with the EU reference artificial digestion method (k-value = 1.00). Positive amplification of mouse tissue experimentally infected with T. spiralis indicated the assay could also be used on encapsulated species in situ. This real-time PCR assay offers an alternative highly specific and sensitive diagnostic method for use in Trichinella wildlife surveillance and could be adapted to wildlife hosts of any region. (C) 2012 Elsevier B.V. All rights reserved.

Red Imported Fire Ants: a threat to eastern Australian wildlife?

The discovery of the Red Imported Fire ants (Solenopsis invicta) in Brisbane on 22 February 2001 sent shock waves through urban and rural communities alike. This article is an attempt to address the often repeated question ‘What will become of Australia's unique fauna if they spread along Australia's eastern seaboard?

Toxoplasmosis in Indo-Pacific humpbacked dolphins (Sousa chinensis) from Queensland

Objective: To describe the clinical signs, gross pathology, serology, bacteriology, histopathology, electron microscopy and immunohistochemistry findings associated with toxoplasmosis in four Indo-Pacific humpbacked dolphins (Sousa chinensis) that stranded in Queensland in 2000 and 2001. Design: Clinical assessment, gross necropsy, and laboratory examinations. Procedure: Necropsies were performed on four S chinensis to determine cause of death. Laboratory tests including serology, bacteriology, histopathology and transmission electron microscopy were done on the four dolphins. lmmunohistochemistry was done on the brain, heart, liver, lung, spleen and adrenal gland from various dolphins to detect Toxoplasma gondii antigens. Results: Necropsies showed all of four S chinensis that stranded in Queensland in 2000 and 2001 had evidence of predatory shark attack and three were extremely emaciated. Histopathological examinations showed all four dolphins had toxoplasmosis with tissue cysts resembling T gondii in the brain. Tachyzoite stages of T gondii were detected in the lungs, heart, liver, spleen and adrenal gland, variously of all four dolphins. Electron microscopy studies and immunohistochemistry confirmed the tissues cysts were those of Tgondii. All four dolphins also had intercurrent disease including pneumonia, three had peritonitis and one had pancreatitis. Conclusion: Four S chinensis necropsied in Queensland in 2000 and 2001 were found to be infected with toxoplasmosis. It is uncertain how these dolphins became infected and further studies are needed to determine how S chinensis acquire toxoplasmosis. All four dolphins stranded after periods of heavy rainfall, and coastal freshwater runoff may be a risk factor for T gondii infection in S chinensis. This disease should be of concern to wildlife managers since S chinensis is a rare species and its numbers appear to be declining.

Low-density koala (Phascolarctos cinereus) populations in the mulgalands of south-west Queensland. II. Distribution and diet

This study used faecal pellets to investigate the broadscale distribution and diet of koalas in the mulgalands biogeographic region of south-west Queensland. Koala distribution was determined by conducting faecal pellet searches within a 30-cm radius of the base of eucalypts on 149 belt transects, located using a multi-scaled stratified sampling design. Cuticular analysis of pellets collected ffom 22 of these sites was conducted to identify the dietary composition of koalas within the region. Our data suggest that koala distribution is concentrated in the northern and more easterly regions of the study area, and appears to be strongly linked with annual rainfall. Over 50% of our koala records were obtained from non-riverine communities, indicating that koalas in the study area are not primarily restricted to riverine communities, as bas frequently been suggested. Cuticular analysis indicates that more than 90% of koala diet within the region consists of five eucalypt species. Our data highlights the importance of residual Tertiary landforms to koala conservation in the region.

Low-density koala (Phascolarctos cinereus) populations in the mulgalands of south-west Queensland. IV. Abundance and conservation status

Urban encroachment on dense, coastal koala populations has ensured that their management has received increasing government and public attention. The recently developed National Koala Conservation Strategy calls for maintenance of viable populations in the wild. Yet the success of this, and other, conservation initiatives is hampered by lack of reliable and generally accepted national and regional population estimates. In this paper we address this problem in a potentially large, but poorly studied, regional population in the State that is likely to have the largest wild populations. We draw on findings from previous reports in this series and apply the faecal standing-crop method (FSCM) to derive a regional estimate of more than 59 000 individuals. Validation trials in riverine communities showed that estimates of animal density obtained from the FSCM and direct observation were in close agreement. Bootstrapping and Monte Carlo simulations were used to obtain variance estimates for our population estimates in different vegetation associations across the region. The most favoured habitat was riverine vegetation, which covered only 0.9% of the region but supported 45% of the koalas. We also estimated that between 1969 and 1995 -30% of the native vegetation associations that are considered as potential koala habitat were cleared, leading to a decline of perhaps 10% in koala numbers. Management of this large regional population has significant implications for the national conservation of the species: the continued viability of this population is critically dependent on the retention and management of riverine and residual vegetation communities, and future vegetation-management guidelines should be cognisant of the potential impacts of clearing even small areas of critical habitat. We also highlight eight management implications.

Biology of Plant Rhabdoviruses

The Rhabdoviridae, whose members collectively infect invertebrates, animals, and plants, form a large family that has important consequences for human health, agriculture, and wildlife ecology. Plant rhabdoviruses can be separated into the genera Cytorhabdovirus and Nucleorhabdovirus, based on their sites of replication and morphogenesis. This review presents a general overviewof classical and contemporary findings about rhabdovirus ecology, pathology, vector relations, and taxonomy. The genome organization and structure of several recently sequenced nucleorhabdoviruses and cytorhabdoviruses is integrated with new cell biology findings to provide a model for the replication of the two genera. A prospectus outlines the exciting opportunities for future research that will contribute to a more detailed understanding of the biology, biochemistry, replication and host interactions of the plant rhabdoviruses.

Neuro-angiostrongylosis in wild Black and Grey-headed flying foxes (Pteropus spp)

Objective: To identify nematodes seen in histological sections of brains of flying foxes (fruit bats) and describe the associated clinical disease and pathology. Proceedures: Gross and histological examination of brains from 86 free-living flying foxes with neurological disease was done as part of an ongoing surveillance program for Australian bat lyssavirus. Worms were recovered, or if seen in histological sections, extracted by maceration of half the brain and identified by microscopic examination. Histological archives were also reviewed. Results: There was histological evidence of angiostrongylosis in 16 of 86 recently submitted flying foxes with neurological disease and in one archival case from 1992. In 10 flying foxes, worms were definitively identified as Angiostrongylus cantonensis fifth-stage larvae. A worm fragment and third stage larvae were identified as Angiostrongylus sp, presumably A cantonensis, in a further three cases. The clinical picture was dominated by paresis, particularly of the hindlimbs, and depression, with flying foxes surviving up to 22 days in the care of wildlife volunteers. Brains containing fifthstage larvae showed a moderate to severe eosinophilic and granulomatous meningoencephalitis (n = 14), whereas there was virtually no inflammation of the brains of bats which died when infected with only smaller, third-stage larvae (n = 3). There was no histological evidence of pulmonary involvement. Conclusion: This is the first report of the recovery and identification of A cantonensis from free-living Australian wildlife. While angiostrongylosis is a common cause of paresis in flying foxes, the initial clinical course cannot be differentiated from Australian bat lyssavirus infection, and wildlife carers should be urged not to attempt to rehabilitate flying foxes with neurological disease.

Australian Bat Lyssavirus: Observations of Natural and Experimental Infection in Bats

This conference abstract gives data and conclusions arising from targeted surveillance of wild bats for naturally occuring Australian bat lyssavirus (ABLV) infection and other central nervous system diseases. It also provides data and conclusions arising from experimental infection of 10 Greyheaded flying foxes (Pteropus poliocephalus).

Emerging henipaviruses and flying foxes - Conservation and management perspectives

Wildlife populations are affected by a series of emerging diseases, some of which pose a significant threat to their conservation. They can also be reservoirs of pathogens that threaten domestic animal and human health. In this paper, we review the ecology of two viruses that have caused significant disease in domestic animals and humans and are carried by wild fruit bats in Asia and Australia. The first, Hendra virus, has caused disease in horses and/or humans in Australia every five years since it first emerged in 1994. Nipah virus has caused a major outbreak of disease in pigs and humans in Malaysia in the late 1990s and has also caused human mortalities in Bangladesh annually since 2001. Increased knowledge of fruit bat population dynamics and disease ecology will help improve our understanding of processes driving the emergence of diseases from bats. For this, a transdisciplinary approach is required to develop appropriate host management strategies that both maximise the conservation of bat populations as well as minimise the risk of disease outbreaks in domestic animals and humans.

Henipaviruses: Emerging Paramyxoviruses Associated with Fruit Bats

Two related, novel, zoonotic paramyxoviruses have been described recently. Hendra virus was first reported in horses and thence humans in Australia in 1994; Nipah virus was first reported in pigs and thence humans in Malaysia in 1998. Human cases of Nipah virus infection, apparently unassociated with infection in livestock, have been reported in Bangladesh since 2001. Species of fruit bats (genus Pteropus ) have been identified as natural hosts of both agents. Anthropogenic changes (habitat loss, hunting) that have impacted the population dynamics of Pteropus species across much of their range are hypothesised to have facilitated emergence. Current strategies for the management of henipaviruses are directed at minimising contact with the natural hosts, monitoring identified intermediate hosts, improving biosecurity on farms, and better disease recognition and diagnosis. Investigation of the emergence and ecology of henipaviruses warrants a broad, cross-disciplinary ecosystem health approach that recognises the critical linkages between human activity, ecological change, and livestock and human health.

Non-target impacts of poison baiting for predator control in Australia

1. Mammalian predators are controlled by poison baiting in many parts of the world, often to alleviate their impacts on agriculture or the environment. Although predator control can have substantial benefits, the poisons used may also be potentially harmful to other wildlife. 2. Impacts on non-target species must be minimized, but can be difficult to predict or quantify. Species and individuals vary in their sensitivity to toxins and their propensity to consume poison baits, while populations vary in their resilience. Wildlife populations can accrue benefits from predator control, which outweigh the occasional deaths of non-target animals. We review recent advances in Australia, providing a framework for assessing non-target effects of poisoning operations and for developing techniques to minimize such effects. We also emphasize that weak or circumstantial evidence of non-target effects can be misleading. 3. Weak evidence that poison baiting presents a potential risk to non-target species comes from measuring the sensitivity of species to the toxin in the laboratory. More convincing evidence may be obtained by quantifying susceptibility in the field. This requires detailed information on the propensity of animals to locate and consume poison baits, as well as the likelihood of mortality if baits are consumed. Still stronger evidence may be obtained if predator baiting causes non-target mortality in the field (with toxin detected by post-mortem examination). Conclusive proof of a negative impact on populations of non-target species can be obtained only if any observed non-target mortality is followed by sustained reductions in population density. 4. Such proof is difficult to obtain and the possibility of a population-level impact cannot be reliably confirmed or dismissed without rigorous trials. In the absence of conclusive evidence, wildlife managers should adopt a precautionary approach which seeks to minimize potential risk to non-target individuals, while clarifying population-level effects through continued research.

Melaleuca densispicata in Currawinya National Park, South-West Queensland: Ecology and preliminary implications for management of a rare plant species

Melaleuca densispicata Byrnes is an uncommon species with a limited distribution, comprising disjunct populations in inland southern Queensland and northern New South Wales, Australia. It is a dense, woody shrub, 2–4 m in height, which exhibits a marked 'clumping' growth habit. It has thick, papery bark and displays many white flowers during spring or early summer. Although it has long been known to exist, M. densispicata was only formally described in 1984, and very little is currently known about its ecology or specific management requirements. There are only seven known subpopulations of the species across its range. A major population at the western limit of its distribution occurs on Currawinya National Park (28°52'S, 144°30'E). Here, it is locally abundant and listed as a noteworthy plant species under the Management Plan (Queensland Parks & Wildlife Service 2001). This study aimed to identify patterns in the distribution of M. densispicata in Currawinya National Park, describe its ecological niche and role, and provide management recommendations for the species within the study area. Recent anecdotal observations of recruitment failure in south-western Queensland (Peter McRae, QPWS, October 2004, pers. comm.; Dick O'Connell, local grazier, July 2005 pers. comm.) caused additional emphasis to be placed on the examination of recruitment and recruitment factors.

Frequency and precision of aerial surveys for kangaroo management

The appropriate frequency and precision for surveys of wildlife populations represent a trade-off between survey cost and the risk of making suboptimal management decisions because of poor survey data. The commercial harvest of kangaroos is primarily regulated through annual quotas set as proportions of absolute estimates of population size. Stochastic models were used to explore the effects of varying precision, survey frequency and harvest rate on the risk of quasiextinction for an arid-zone and a more mesic-zone kangaroo population. Quasiextinction probability increases in a sigmoidal fashion as survey frequency is reduced. The risk is greater in more arid regions and is highly sensitive to harvest rate. An appropriate management regime involves regular surveys in the major harvest areas where harvest rate can be set close to the maximum sustained yield. Outside these areas, survey frequency can be reduced in relatively mesic areas and reduced in arid regions when combined with lowered harvest rates. Relative to other factors, quasiextinction risk is only affected by survey precision (standard error/mean × 100) when it is >50%, partly reflecting the safety of the strategy of harvesting a proportion of a population estimate.

The influence of animal mobility on the assumption of uniform distances in aerial line-transect surveys

Line-transect distance sampling is a widely used method for estimating animal density from aerial surveys. Analysis of line-transect distance data usually relies on a requirement that the statistical distribution of distances of animal groups from the transect line is uniform. We show that this requirement is satisfied by the survey design if all other assumptions of distance sampling hold, but it can be violated by consistent survey problems such as responsive movement of the animals towards or away from the observer. We hypothesise that problems with the uniform requirement are unlikely to be encountered for immobile taxa, but might become substantial for species of high mobility. We test evidence for non-uniformity using double-observer distance data from two aerial surveys of five species with a spectrum of mobility capabilities and tendencies. No clear evidence against uniformity was found for crabeater seals or emperor penguins on the pack-ice in East Antarctica, while minor non-uniformity consistent with responsive movement up to 30 m was found for Adelie penguins. Strong evidence of either non-uniformity or a failure of the capture-recapture validating method was found for eastern grey kangaroos and red kangaroos in Queensland.

A comparison of mark-recapture distance-sampling methods applied to aerial surveys of eastern grey kangaroos

Aerial surveys of kangaroos (Macropus spp.) in Queensland are used to make economically important judgements on the levels of viable commercial harvest. Previous analysis methods for aerial kangaroo surveys have used both mark-recapture methodologies and conventional distance-sampling analyses. Conventional distance sampling has the disadvantage that detection is assumed to be perfect on the transect line, while mark-recapture methods are notoriously sensitive to problems with unmodelled heterogeneity in capture probabilities. We introduce three methodologies for combining together mark-recapture and distance-sampling data, aimed at exploiting the strengths of both methodologies and overcoming the weaknesses. Of these methods, two are based on the assumption of full independence between observers in the mark-recapture component, and this appears to introduce more bias in density estimation than it resolves through allowing uncertain trackline detection. Both of these methods give lower density estimates than conventional distance sampling, indicating a clear failure of the independence assumption. The third method, termed point independence, appears to perform very well, giving credible density estimates and good properties in terms of goodness-of-fit and percentage coefficient of variation. Estimated densities of eastern grey kangaroos range from 21 to 36 individuals km-2, with estimated coefficients of variation between 11% and 14% and estimated trackline detection probabilities primarily between 0.7 and 0.9.