Biological management of the invasive Vachellia nilotica ssp. indica (Benth.) Kyal. & Boatwr. (Fabales: Mimosoideae) in tropical Australia: stress-inducing potential of Anomalococcus indicus Ramakrishna (Insecta: Hemiptera: Coccoidea: Lecanodiaspididae), an agent of promise

Vachellia nilotica ssp. indica (hereafter, V. n. indica) is an important tree weed in Australia. Its dense populations induce undesirable changes in the vast areas of northern Australia. Because chemical and mechanical management options appear unviable for various reasons, biological management of this tree is considered a better option. Among the many trialled arthropods in Australian context, Anomalococcus indicus, a lecanodiaspid native to India, has been identified as a potent-candidate, since in India, its native terrain, it is the most widespread and occurs throughout the year. Severe infestations of A. indicus cause defoliation, wilting and death of branches, and occasionally the tree. Populations of A. indicus have been brought into Australia and are being tested for its host specificity under quarantine conditions. This article reports the physiological damage and stress it inflicts in the shoots of V. n. indica. Younger-nymphal instars of A. indicus feed on cortical-parenchyma cells of young stems, whereas the older instars and adults feed from the phloem of old stems. Two conspicuous responses of V. n. indica arising in response to the feeding action of A. indicus are changes in the cell-wall dynamics and irregular cell divisions. The feeding action of A. indicus elicits a sequence of reactions in the stem tissues of V. n. indica such as differentiation of thick-walled elements in the outer cortical parenchyma, differential thickening of cells with supernumerary layers of either suberin or lignin, proliferations of parenchyma and phloem, wall thickening and obliteration of inner lumen of phloem cells, and the sieve plates plugged with callosic deposits. The responses are the culminations of interaction between the virulence factor (one or more of the salivary proteins?) from A. indicus and the resistance factor in V. n. indica. We have analysed structural changes in the context of their functions, by comparing the feeding action of A. indicus with that of other hemipteroids. From the level of stress it induces, this study confirms that A. indicus has the potential to be an effective biological management of V. n. indica in Australia. © 2014 © 2014 Taylor & Francis and Aboricultural Association.

Australian bat lyssavirus infection in two horses

In May 2013, the first cases of Australian bat lyssavirus infections in domestic animals were identified in Australia. Two horses (filly-H1 and gelding-H2) were infected with the Yellow-bellied sheathtail bat (YBST) variant of Australian bat lyssavirus (ABLV). The horses presented with neurological signs, pyrexia and progressing ataxia. Intra-cytoplasmic inclusion bodies (Negri bodies) were detected in some Purkinje neurons in haematoxylin and eosin (H&E) stained sections from the brain of one of the two infected horses (H2) by histological examination. A morphological diagnosis of sub-acute moderate non-suppurative, predominantly angiocentric, meningo-encephalomyelitis of viral aetiology was made. The presumptive diagnosis of ABLV infection was confirmed by the positive testing of the affected brain tissue from (H2) in a range of laboratory tests including fluorescent antibody test (FAT) and real-time PCR targeting the nucleocapsid (N) gene. Retrospective testing of the oral swab from (H1) in the real-time PCR also returned a positive result. The FAT and immunohistochemistry (IHC) revealed an abundance of ABLV antigen throughout the examined brain sections. ABLV was isolated from the brain (H2) and oral swab/saliva (H1) in the neuroblastoma cell line (MNA). Alignment of the genome sequence revealed a 97.7% identity with the YBST ABLV strain.

Polarized immune responses modulated by layered double hydroxides nanoparticle conjugated with CpG

Modulation of the immune response is an important step in the induction of protective humoral and cellular immunity against pathogens. In this study, we investigated the possibility of using a nanomaterial conjugated with the toll-like receptor (TLR) ligand CpG to modulate the immune response towards the preferred polarity. MgAl-layered double hydroxide (LDH) nanomaterial has a very similar chemical composition to Alum, an FDA approved adjuvant for human vaccination. We used a model antigen, ovalbumin (OVA) to demonstrate that MgAl-LDH had comparable adjuvant activity to Alum, but much weaker inflammation. Conjugation of TLR9 ligand CpG to LDH nanoparticles significantly enhanced the antibody response and promoted a switch from Th2 toward Th1 response, demonstrated by a change in the IgG2a:IgG1 ratio. Moreover, immunization of mice with CpG-OVA-conjugated LDH before challenge with OVA-expressing B16/F10 tumor cells retarded tumor growth. Together, these data indicate that LDH nanomaterial can be used as an immune adjuvant to promote Th1 or Th2 dominant immune responses suitable for vaccination purposes.

A comparison of welfare outcomes for weaner and mature Bos indicus bulls surgically or tension band castrated with or without analgesia: 2. Responses related to stress, health and productivity

Tension banding castration of cattle is gaining favour because it is relatively simple to perform and is promoted by retailers of the banders as a humane castration method. Two experiments were conducted, under tropical conditions using Bos indicus bulls comparing tension banding (Band) and surgical (Surgical) castration of weaner (7–10 months old) and mature (22–25 months old) bulls with and without pain management (NSAID (ketoprofen) or saline injected intramuscularly immediately prior to castration). Welfare outcomes were assessed using a range of measures; this paper reports on some physiological, morbidity and productivity-related responses to augment the behavioural responses reported in an accompanying paper. Blood samples were taken on the day of castration (day 0) at the time of restraint (0 min) and 30 min (weaners) or 40 min (mature bulls), 2 h, and 7 h; and days 1, 2, 3, 7, 14, 21 and 28 post-castration. Plasmas from day 0 were assayed for cortisol, creatine kinase, total protein and packed cell volume. Plasmas from the other samples were assayed for cortisol and haptoglobin (plus the 0 min sample). Liveweights were recorded approximately weekly to 6 weeks and at 2 and 3 months post-castration. Castration sites were checked at these same times to 2 months post-castration to score the extent of healing and presence of sepsis. Cortisol concentrations (mean ± s.e. nmol/L) were significantly (P < 0.05) higher in the Band (67 ± 4.5) compared with Surgical weaners (42 ± 4.5) at 2 h post-castration, but at 24 h post-castration were greater in the Surgical (43 ± 3.2) compared with the Band weaners (30 ± 3.2). The main effect of ketoprofen was on the cortisol concentrations of the mature Surgical bulls; concentrations were significantly reduced at 40 min (47 ± 7.2 vs. 71 ± 7.2 nmol/L for saline) and 2 h post-castration (24 ± 7.2, vs. 87 ± 7.2 nmol/L for saline). Ketoprofen, however, had no effect on the Band mature bulls, with their cortisol concentrations averaging 54 ± 5.1 nmol/L at 40 min and 92 ± 5.1 nmol/L at 2 h. Cortisol concentrations were also significantly elevated in the Band (83 ± 3.0 nmol/L) compared with Surgical mature bulls (57 ± 3.0 nmol/L) at weeks 2–4 post-castration. The timing of this elevation coincided with significantly elevated haptoglobin concentrations (mg/mL) in the Band bulls (2.97 ± 0.102 for mature bulls and 1.71 ± 0.025 for weaners, vs. 2.10 ± 0.102 and 1.45 ± 0.025 respectively for the Surgical treatment) and evidence of slow wound healing and sepsis in both the weaner (0.81 ± 0.089 not healed at week 4 for Band, 0.13 ± 0.078 for Surgical) and mature bulls (0.81 ± 0.090 at week 4 for Band, 0.38 ± 0.104 for Surgical). Overall, liveweight gains of both age groups were not affected by castration method. The findings of acute pain, chronic inflammation and possibly chronic pain in the mature bulls at least, together with poor wound healing in the Band bulls support behavioural findings reported in the accompanying paper and demonstrate that tension banding produces inferior welfare outcomes for weaner and mature bulls compared with surgical castration.

A comparison of welfare outcomes for weaner and mature Bos indicus bulls surgically or tension band castrated with or without analgesia: 1. Behavioural responses

Tension-band castration of cattle is gaining favour because it is relatively simple to perform and is promoted by retailers of the devices as a humane castration method. Furthermore, retailers encourage delaying castration to exploit the superior growth rates of bulls compared with steers. Two experiments were conducted, under tropical conditions, comparing tension banding and surgical castration of weaner (7–10 months old) and mature (22–25 months old) Bos indicus bulls with and without pain management (ketoprofen or saline injected intramuscularly immediately prior to castration). Welfare outcomes were assessed using a wide range of measures; this paper reports on the behavioural responses of the bulls and an accompanying paper reports on other measures. Behavioural data were collected at intervals by direct observation and continuously via data loggers on the hind leg of the bulls to 4 weeks post-castration. Tension-banded bulls performed less movement in the crush/chute than the surgically castrated bulls during the procedures (weaner: 2.63 vs. 5.69, P < 0.001; mature: 1.00 vs. 5.94; P < 0.001 for tension-band and surgical castration, respectively), indicating that tension banding was less painful then surgical castration during conduct. To 1.5 h post-castration, tension-banded bulls performed significantly (all P < 0.05) more active behavioural responses indicative of pain compared with surgical castrates, e.g., percentage time walking forwards (weaner: 15.0% vs. 8.1%; mature: 22.3% vs. 15.1%), walking backwards (weaner: 4.3% vs. 1.4%; mature: 2.4% vs. 0.5%), numbers of tail movements (weaner: 21.9 vs. 1.4; mature: 51.5 vs. 39.4) and leg movements (weaner: 12.9 vs. 0.9; mature: 8.5 vs. 1.5), respectively. In contrast, surgically castrated bulls performed more immobile behaviours compared with tension-banded bulls (e.g., standing in mature bulls was 56.6% vs. 34.4%, respectively, P = 0.002). Ketoprofen administration appeared effective in moderating pain-related behaviours in the mature bulls from 1.5 to 3 h, e.g., reducing abnormal standing (0.0% vs. 7.7%, P = 0.009) and increasing feeding (12.7% vs. 0.0%, P = 0.048) in NSAID- and saline-treated bulls, respectively. There were few behavioural differences subsequent to 24 h post-castration, but some limited evidence of chronic pain (3–4 weeks post-castration) with both methods. Interpretation, however, was difficult from behaviours alone. Thus, tension banding is less painful than surgical castration during conduct of the procedures and pain-related behavioural responses differ with castration method (active restlessness in response to tension banding and minimisation of movement in response to surgical castration). Ketoprofen administered immediately prior to castration was somewhat effective in reducing pain, particularly in the mature bulls.

Measuring Physiological Stress in Australian Flying-Fox Populations

Flying-foxes (pteropid bats) are the natural host of Hendra virus, a recently emerged zoonotic virus responsible for mortality or morbidity in horses and humans in Australia since 1994. Previous studies have suggested physiological and ecological risk factors for infection in flying-foxes, including physiological stress. However, little work has been done measuring and interpreting stress hormones in flying-foxes. Over a 12-month period, we collected pooled urine samples from underneath roosting flying-foxes, and urine and blood samples from captured individuals. Urine and plasma samples were assayed for cortisol using a commercially available enzyme immunoassay. We demonstrated a typical post-capture stress response in flying-foxes, established urine specific gravity as an attractive alternative to creatinine to correct urine concentration, and established population-level urinary cortisol ranges (and geometric means) for the four Australian species: Pteropus alecto 0.5–305.1 ng/mL (20.1 ng/mL); Pteropus conspicillatus 0.3–370.9 ng/mL (18.9 ng/mL); Pteropus poliocephalus 0.3–311.3 ng/mL (10.1 ng/mL); Pteropus scapulatus 5.2–205.4 ng/mL (40.7 ng/mL). Geometric means differed significantly except for P. alecto and P. conspicillatus. Our approach is methodologically robust, and has application both as a research or clinical tool for flying-foxes, and for other free-living colonial wildlife species

Remembering the Peel Island Lazaret through re-imagining the Teerk Roo Ra forest

This paper discusses my current research which aims to re-member the site of the Peel Island Lazaret through re-imagining the Teerk Roo Ra forest as a series of animated artworks. Teerk Roo Ra National Park (formally known as Peel Island) is a small island in Moreton Bay, Queensland and is visible on the ferry journey from Cleveland to Stradbroke Island. The island has an intriguing history, and is the site of a former Lazaret and quarantine station. The Lazaret treated patients diagnosed with Hansen’s disease (or Leprosy), and operated between 1907 and 1959. In this paper I will discuss conceptions of the non-indigenous historical context of the Peel Island Lazaret and the notion of the liminal state (Turner,1967). Through this discussion conceptions of place from Australian cultural theorist Ross Gibson are also examined. The concept of two overlapping realms is then explored through the clues and shared stories about the people who inhabited the site. There is then an explanation of my own approach to re-member this place through re-imagining the forest that witnessed the events of the Lazaret. I then draw on theories of the uncanny from German Psychiatrist Ernst Jentsch, Austrian Neurologist Sigmund Freud and South African animation theorist Meg Rickards to argue that my experience of the forest of Teerk Roo Ra was an uncanny experience where two worlds or states of mind existed simultaneously and overlapped, causing a viscerally unsettling uncanny experience. Through an analysis of Czech Surrealist Animator Jan Švankmajer’s cinematic narrative Down to the cellar (1982), my creative work Structure #24(2011), and Australian Artist Patricia Piccinini’s cinematic artwork The Gathering (2007), I discuss the situation of the inanimate and the animate co-existing simultaneously. Using this approach I propose an understanding of the uncanny as an intellectual uncertainty as outlined by Jentsch (1906). I also develop the notion of the familiar being concealed and becoming unfamiliar through mimicry (Freud, 1919). These discussions form an introduction to my creative work Nocturne #5(2014) which re-members the forests of Teerk Roo Ra as an uncanny place primarily expressed through animation.

Biological control of prickly acacia: Current research and future prospects

Prickly acacia, Vachellia nilotica subsp. indica (syn. Acacia nilotica subsp. indica) (Fabaceae), a major weed in the natural grasslands of western Queensland, has been a target of biological control since the 1980s with limited success to date. Surveys in India, based on genetic and climate matching, identified five insects and two rust pathogens as potential agents. Host-specificity tests were conducted for the insects in India and under quarantine conditions in Australia, and for the rust pathogens under quarantine conditions at CABI in the UK. In no-choice tests, the brown leaf-webber, Phycita sp. A, (Lepidoptera: Pyralidae) completed development on 17 non-target plant species. Though the moth showed a clear preference for prickly acacia in oviposition choice trials screening of additional test-plant species was terminated in view of the potential non-target risk. The scale insect Anomalococcus indicus (Hemiptera: Lecanodiaspididae) developed into mature gravid females on 13 out of 58 non-target plant species tested. In the majority of cases very few female scales matured but development was comparable to that on prickly acacia on four of the non-target species. In multiple choice tests, the scale insect showed a significant preference for the target weed over non-target species tested. In a paired-choice trial under field conditions in India, crawler establishment occurred only on prickly acacia and not on the non-target species tested. Further choice trials are to be conducted under natural field conditions in India. A colony of the green leaf-webber Phycita sp. B has been established in quarantine facilities in Australia and host-specificity testing has commenced. The gall-rust Ravenelia acaciae-arabicae and the leaf-rust Ravenelia evansii (Puccineales: Raveneliaceae) both infected and produced viable urediniospores on Vachellia sutherlandii (Fabaceae), a non-target Australian native plant species. Hence, no further testing with the two rust species was pursued. Inoculation trials using the gall mite Aceria liopeltus (Acari: Eriophyidae) from V. nilotica subsp. kraussiana in South Africa resulted in no gall induction on V. nilotica subsp. indica. Future research will focus on the leaf-weevil Dereodus denticollis (Coleoptera: Curculionidae) and the leaf-beetle Pachnephorus sp. (Coleoptera: Chrysomelidae) under quarantine conditions in Australia. Native range surveys for additional potential biological control agents will also be pursued in northern and western Africa.

Biological control of bellyache bush (Jatropha gossypiifolia) in Australia: South America as a possible source of natural enemies

Bellyache bush (Jatropha gossypiifolia, Euphorbiaceae), a deciduous shrub introduced as an ornamental from tropical America, is a major and expanding weed of rangelands and riparian zones in northern Australia. Biological control is the most economically viable and long-term management solution for this weed. Surveys for potential biological control agents for J gossypiifolia in Mexico,Central America and the Caribbean resulted in release of the seed-feeding jewel bug Agonosoma trilineatum (Hemiptera: Scutelleridae), which failed to establish, and prioritisation of a leaf-rust Phakopsora arthuriana (Puccineales: Phakopsoraceae) for host-specificity testing, which is ongoing. With poor prospects for new agents from Mexico and Central America and the Caribbean, the search for candidate agents on J gossypiifolia shifted to localities south of the equator. Surveys were conducted on the purple-leaf form of J gossypiifolia, Jatropha excisa, Jatropha clavuligera and Jatropha curcas in Peru, Bolivia and Paraguay in 2012 and 2013. A total of 11 insect species, one mite species and the leaf-rust (P. arthuriana) were observed. These include a yet to be described leafmining moth (Stomphastis sp.) (Lepidoptera: Gracillaridae), a shoot and leaf-galling midge Prodiplosis longifila, and leaf-feeding midge Prodiplosis sp. near longifila (both Diptera:Cecidomyiidae) and an unidentified leaf-feeding moth larva (Lepidoptera: Notodontidae). The leafminer is widespread and damaging and has a field host range restricted to the genus Jatropha in Peru and Bolivia, holds the greatest promise as a biological control agent in Australia. Phakopsora arthuriana was recorded for the first time ever from Bolivia and Peru. Further exploration will be conducted in Peru and Bolivia during the wet season to confirm the field host range of collected agents,and to look for more new agents. Promising agents with field host-range restricted to Jatropha spp. will be imported into a quarantine facility in Australia for host-specificity testing.

Do Human Extraintestinal Escherichia coli Infections Resistant to Expanded-Spectrum Cephalosporins Originate From Food-Producing Animals? A Systematic Review

To find out whether food-producing animals (FPAs) are a source of extraintestinal expanded-spectrum cephalosporin-resistant Escherichia coli (ESCR-EC) infections in humans, Medline, Embase, and the Cochrane Database of Systematic Reviews were systematically reviewed. Thirty-four original, peer-reviewed publications were identified for inclusion. Six molecular epidemiology studies supported the transfer of resistance via whole bacterium transmission (WBT), which was best characterized among poultry in the Netherlands. Thirteen molecular epidemiology studies supported transmission of resistance via mobile genetic elements, which demonstrated greater diversity of geography and host FPA. Seventeen molecular epidemiology studies did not support WBT and two did not support mobile genetic element-mediated transmission. Four observational epidemiology studies were consistent with zoonotic transmission. Overall, there is evidence that a proportion of human extraintestinal ESCR-EC infections originate from FPAs. Poultry, in particular, is probably a source, but the quantitative and geographical extent of the problem is unclear and requires further investigation.

Development of a Rapid Multiplex PCR Assay To Genotype Pasteurella multocida Strains by Use of the Lipopolysaccharide Outer Core Biosynthesis Locus

Pasteurella multocida is a Gram-negative bacterial pathogen that is the causative agent of a wide range of diseases in many animal species, including humans. A widely used method for differentiation of P. multocida strains involves the Heddleston serotyping scheme. This scheme was developed in the early 1970s and classifies P. multocida strains into 16 somatic or lipopolysaccharide (LPS) serovars using an agar gel diffusion precipitin test. However, this gel diffusion assay is problematic, with difficulties reported in accuracy, reproducibility, and the sourcing of quality serovar-specific antisera. Using our knowledge of the genetics of LPS biosynthesis in P. multocida, we have developed a multiplex PCR (mPCR) that is able to differentiate strains based on the genetic organization of the LPS outer core biosynthesis loci. The accuracy of the LPS-mPCR was compared with classical Heddleston serotyping using LPS compositional data as the "gold standard." The LPS-mPCR correctly typed 57 of 58 isolates; Heddleston serotyping was able to correctly and unambiguously type only 20 of the 58 isolates. We conclude that our LPS-mPCR is a highly accurate LPS genotyping method that should replace the Heddleston serotyping scheme for the classification of P. multocida strains.

Antimicrobial susceptibility of Histophilus somni isolated from clinically affected cattle in Australia

This study investigated antimicrobial resistance traits, clonal relationships and epidemiology of Histophilus somni isolated from clinically affected cattle in Queensland and New South Wales, Australia. Isolates (n = 53) were subjected to antimicrobial susceptibility testing against six antimicrobial agents (ceftiofur, enrofloxacin, florfenicol, tetracycline, tilmicosin and tulathromycin) using disc diffusion and minimum inhibitory concentration (MIC) assays. Clonal relationships were assessed using repetitive sequence PCR and descriptive epidemiological analysis was performed. The H. somni isolates appeared to be geographically clonal, with 27/53 (47%) isolates grouping in one cluster from one Australian state. On the basis of disc diffusion, 34/53 (64%) isolates were susceptible to all antimicrobial agents tested; there was intermediate susceptibility to tulathromycin in 12 isolates, tilmicosin in seven isolates and resistance to tilmicosin in one isolate. Using MIC, all but one isolate was susceptible to all antimicrobial agents tested; the non-susceptible isolate was resistant to tetracycline, but this MIC result could not be compared to disc diffusion, since there are no interpretative guidelines for disc diffusion for H. somni against tetracycline. In this study, there was little evidence of antimicrobial resistance in H. somni isolates from Australian cattle. Disc diffusion susceptibility testing results were comparable to MIC results for most antimicrobial agents tested; however, results for isolates with intermediate susceptibility or resistance to tilmicosin and tulathromycin on disc diffusion should be interpreted with caution in the absence of MIC results.

Rhipicephalus microplus serine protease inhibitor family: annotation, expression and functional characterisation assessment

Background: Rhipicephalus (Boophilus) microplus evades the host's haemostatic system through a complex protein array secreted into tick saliva. Serine protease inhibitors (serpins) conform an important component of saliva which are represented by a large protease inhibitor family in Ixodidae. These secreted and non-secreted inhibitors modulate diverse and essential proteases involved in different physiological processes. Methods: The identification of R. microplus serpin sequences was performed through a web-based bioinformatics environment called Yabi. The database search was conducted on BmiGi V1, BmiGi V2.1, five SSH libraries, Australian tick transcriptome libraries and RmiTR V1 using bioinformatics methods. Semi quantitative PCR was carried out using different adult tissues and tick development stages. The cDNA of four identified R. microplus serpins were cloned and expressed in Pichia pastoris in order to determine biological targets of these serpins utilising protease inhibition assays. Results: A total of four out of twenty-two serpins identified in our analysis are new R. microplus serpins which were named as RmS-19 to RmS-22. The analyses of DNA and predicted amino acid sequences showed high conservation of the R. microplus serpin sequences. The expression data suggested ubiquitous expression of RmS except for RmS-6 and RmS-14 that were expressed only in nymphs and adult female ovaries, respectively. RmS-19, and -20 were expressed in all tissues samples analysed showing their important role in both parasitic and non-parasitic stages of R. microplus development. RmS-21 was not detected in ovaries and RmS-22 was not identified in ovary and nymph samples but were expressed in the rest of the samples analysed. A total of four expressed recombinant serpins showed protease specific inhibition for Chymotrypsin (RmS-1 and RmS-6), Chymotrypsin / Elastase (RmS-3) and Thrombin (RmS-15). Conclusion: This study constitutes an important contribution and improvement to the knowledge about the physiologic role of R. microplus serpins during the host-tick interaction.

The insecticidal spider toxin SFI1 is a knottin peptide that blocks the pore of insect voltage-gated sodium channels via a large β-hairpin loop

Spider venoms contain a plethora of insecticidal peptides that act on neuronal ion channels and receptors. Because of their high specificity, potency and stability, these peptides have attracted much attention as potential environmentally friendly insecticides. Although many insecticidal spider venom peptides have been isolated, the molecular target, mode of action and structure of only a small minority have been explored. Sf1a, a 46-residue peptide isolated from the venom of the tube-web spider Segesteria florentina, is insecticidal to a wide range of insects, but nontoxic to vertebrates. In order to investigate its structure and mode of action, we developed an efficient bacterial expression system for the production of Sf1a. We determined a high-resolution solution structure of Sf1a using multidimensional 3D/4D NMR spectroscopy. This revealed that Sf1a is a knottin peptide with an unusually large β-hairpin loop that accounts for a third of the peptide length. This loop is delimited by a fourth disulfide bond that is not commonly found in knottin peptides. We showed, through mutagenesis, that this large loop is functionally critical for insecticidal activity. Sf1a was further shown to be a selective inhibitor of insect voltage-gated sodium channels, consistent with its 'depressant' paralytic phenotype in insects. However, in contrast to the majority of spider-derived sodium channel toxins that function as gating modifiers via interaction with one or more of the voltage-sensor domains, Sf1a appears to act as a pore blocker.

A field investigation into a suspected outbreak of pyrrolizidine alkaloid toxicosis in horses in western Queensland

A disease outbreak investigation was conducted in western Queensland to investigate a rare suspected outbreak of pyrrolizidine alkaloid (PA) toxicosis in horses. Thirty five of 132 horses depastured on five properties on the Mitchell grass plains of western Queensland died in the first six months of 2010. Clinical–pathological findings were consistent with PA toxicosis. A local variety of Crotalaria medicaginea was the only hepatotoxic plant found growing on affected properties. Pathology reports and departure and arrival dates of two brood mares provided evidence of a pre wet season exposure period. All five affected properties experienced a very dry spring and early summer preceded by a large summer wet season. The outbreak was characterised as a point epidemic with a sudden peak of deaths in March followed by mortalities steadily declining until the end of June. The estimated morbidity (serum IGG > 50 IU/L) rate was 76%. Average crude mortality was 27% but higher in young horses (67%) and brood mares (44%). Logistic regression analysis showed that young horses and brood mares and those grazing denuded pastures in December were most strongly associated with dying whereas those fed hay and/or grain based supplements were less likely to die. This is the first detailed study of an outbreak of PA toxicosis in central western Queensland and the first to provide evidence that environmental determinants were associated with mortality, that the critical exposure period was towards the end of the dry season, that supplementary feeding is protective and that denuded pastures and the horses physiological protein requirement are risk factors.