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Following the arrival of rabbit haemorrhagic disease virus 2 (RHDV2) in Australia, average rabbit population abundances were reduced by 60% between 2014 and 2018 based on monitoring data acquired from 18 sites across Australia. During this period, as the seropositivity to RHDV2 increased, concurrent decreases were observed in the seroprevalence of both the previously circulating RHDV1 and RCVA, a benign endemic rabbit calicivirus. However, the detection of substantial RHDV1 seropositivity in juvenile rabbits suggested that infections were continuing to occur, ruling out the rapid extinction of this variant. Here we investigate whether the co-circulation of two pathogenic RHDV variants was sustained after 2018 and whether the initially observed impact on rabbit abundance was still maintained. We monitored rabbit abundance and seropositivity to RHDV2, RHDV1 and RCVA at six of the initial eighteen sites until the summer of 2022. We observed sustained suppression of rabbit abundance at five of the six sites, with the average population reduction across all six sites being 64%. Across all sites, average RHDV2 seroprevalence remained high, reaching 60–70% in adult rabbits and 30–40% in juvenile rabbits. In contrast, average RHDV1 seroprevalence declined to <3% in adult rabbits and 5–6% in juvenile rabbits. Although seropositivity continued to be detected in a low number of juvenile rabbits, it is unlikely that RHDV1 strains now play a major role in the regulation of rabbit abundance. In contrast, RCVA seropositivity appears to be reaching an equilibrium with that of RHDV2, with RCVA seroprevalence in the preceding quarter having a strong negative effect on RHDV2 seroprevalence and vice versa, suggesting ongoing co-circulation of these variants. These findings highlight the complex interactions between different calicivirus variants in free-living rabbit populations and demonstrate the changes in interactions over the course of the RHDV2 epizootic as it has moved towards endemicity. While it is encouraging from an Australian perspective to see sustained suppression of rabbit populations in the eight years following the arrival of RHDV2, it is likely that rabbit populations will eventually recover, as has been observed with previous rabbit pathogens.

Integrated pest management (IPM) is widely recommended for managing invasive pests to maximise effectiveness. However, the extent to which IPM occurs is typically unknown. We used the rabbit in Australia as a case study to investigate whether land managers apply IPM. Using 7415 control records voluntarily submitted to RabbitScan by land managers, we found evidence of IPM in only 39%. The extent to which integration occurred depended on the control method applied [warren ripping, poison baiting, release of rabbit haemorrhagic disease virus (RHDV), warren fumigation]. Two of four major rabbit control methods were frequently integrated with at least one other method; 73% of warren ripping and 55% of poison baiting records were associated with another different control method. In contrast, only 15% of RHDV releases and 35% of warren fumigations were integrated with another different control method. This confirmed reports that land managers view RHDV release as a ‘silver bullet’ and release it to avoid applying more expensive but more effective control methods. We identified control sequences that were neither biologically appropriate nor cost-efficient. In some situations, where there is a risk to other animals, or land managers lack suitable equipment, it may be difficult to apply IPM in an optimal sequence or at the optimal time. However, a greater level of control and at a proportionally lower cost could result if IPM principals were more rigorously applied, for instance, by focusing on strategic application of control methods in sequences known to be highly effective and cost-efficient.

Australia has multiple lagoviruses with differing pathogenicity. The circulation of these viruses was traditionally determined through opportunistic sampling events. In the lead up to the nationwide release of RHDVa-K5 (GI.1aP-GI.1a) in 2017, an existing citizen science program, RabbitScan, was augmented to allow members of the public to submit samples collected from dead leporids for lagovirus testing. This study describes the information obtained from the increased number of leporid samples received between 2015 and 2022 and focuses on the recent epidemiological interactions and evolutionary trajectory of circulating lagoviruses in Australia between October 2020 and December 2022. A total of 2771 samples were tested from January 2015 to December 2022, of which 1643 were lagovirus-positive. Notable changes in the distribution of lagovirus variants were observed, predominantly in Western Australia, where RHDV2-4c (GI.4cP-GI.2) was detected again in 2021 after initially being reported to be present in 2018. Interestingly, we found evidence that the deliberately released RHDVa-K5 was able to establish and circulate in wild rabbit populations in WA. Overall, the incorporation of citizen science approaches proved to be a cost-efficient method to increase the sampling area and enable an in-depth analysis of lagovirus distribution, genetic diversity, and interactions. The maintenance of such programs is essential to enable continued investigations of the critical parameters affecting the biocontrol of feral rabbit populations in Australia, as well as to enable the detection of any potential future incursions.

Several conventional and recently available tools are available for an integrated control of European rabbits in Australia. We quantified the impact of the release of rabbit haemorrhagic disease virus K5 (RHDV K5, hereafter K5) and pindone (2‐pivalyl‐1,3‐indandione) baiting at 13 sites within Cudlee Creek fire scar in the Adelaide Hills, South Australia. K5 release was followed by pindone baiting between December 2021 and March 2022; the application of both control methods followed industry best practice. We counted rabbits using spotlights before and after the application of both control methods. Fly samples and livers from dead rabbits were collected to track K5 transmission within and between sites, and to detect the natural circulation of rabbit haemorrhagic disease virus 2 (RHDV2). K5 release had minimal impact on rabbit populations, with treated populations increasing by a mean of 65.5% at 14 days post‐release and 27.9% at 77 days post‐K5 release across all sites, comparable to the changes at control sites. K5 detection in flies up to 77 days post its release, and its detection in rabbit livers, demonstrates that it can survive and transmit in the environment for prolonged periods and that it can lethally infect some rabbits. This limited impact of K5 is consistent with previous studies and may be explained by pre‐existing RHDV/RHDV2 immunity in the target populations or the presence of young rabbits with natural innate RHDV immunity. The detection of K5 in flies from control sites demonstrates that it was vectored beyond its release location. A reduction in rabbit counts post‐pindone baiting was observed at most treatment sites, with a mean population reduction of 36.6% across all sites. Landholders need to carefully and strategically plan their integrated rabbit control programmes. Not all combinations of controls, even if theoretically logical, achieve meaningful outcomes for rabbit management. This study quantified the impact of the release of rabbit haemorrhagic disease virus K5 (RHDV K5, K5 hereafter) and pindone baiting at 13 sites within Cudlee Creek fire scar in the Adelaide Hills region, South Australia. K5 release had minimal impact on rabbit populations, whereas a reduction in rabbit counts post‐pindone baiting was observed at most treatment sites. Landholders need to carefully and strategically plan their integrated rabbit control programmes as not all combinations of controls, even if theoretically logical, achieve meaningful outcomes for rabbit management.

Rabbit haemorrhagic disease virus (RHDV) is established as a landscape-scale biocontrol that assists the management of invasive European rabbits and their impacts in both Australia and New Zealand. In addition to this, it is also available to land managers to augment rabbit control efforts at a local scale. However, current methods of deploying RHDV to rabbits that rely on the consumption of virus-treated baits can be problematic as rabbits are reluctant to consume bait when there is abundant, green, protein-rich feed available. We ran a suite of interrupted time-series experiments to compare the duration of infectivity of two conventional (carrot and oat baits) and two novel (meat bait and soil burrow spray) methods of deploying RHDV to rabbits. All methods effectively killed exposed rabbits. Soil burrow spray and carrot baits resulted in infection and mortality out to 5 days post their deployment in the field, and meat baits caused infection out to 10 days post their deployment. In contrast, oat baits continued to infect and kill exposed rabbits out to 20 days post deployment. Molecular assays demonstrated high viral loads in deployed baits beyond the duration for which they were infectious or lethal to rabbits. Based on our results, we suggest that the drying of meat baits may create a barrier to effective transmission of RHDV by adult flies within 10 days. We therefore hypothesise that fly larvae production and development on infected tissues is critical to prolonged viral transmission from meat baits, and similarly from carcasses of RHDV mortalities, via mechanical fly vectors. Our study demonstrates that meat baits and soil spray could provide additional virus deployment options that remove the need for rabbits to consume baits at times when they are reluctant to do so.

In this study, three different diagnostic tests for parvovirus were compared with vaccination status and parvovirus genotype in suspected canine parvovirus cases. Faecal samples from vaccinated (N17) and unvaccinated or unknown vaccination status (N41) dogs that had clinical signs of parvovirus infection were tested using three different assays of antigen tests, conventional and quantitative PCR tests. The genotype of each sample was determined by sequencing. In addition to the suspected parvovirus samples, 21 faecal samples from apparently healthy dogs were tested in three diagnostic tests to evaluate the sensitivity and specificity of the tests. The antigen test was positive in 41.2% of vaccinated dogs and 73.2% of unvaccinated diseased dogs. Conventional PCR and qPCR were positive for canine parvovirus (CPV) in 82.4% of vaccinated dogs and 92.7% of unvaccinated dogs. CPV type-2c (CPV-2c) was detected in 82.75% of dogs (12 vaccinated and 36 unvaccinated dogs), CPV-2b was detected in 5.17% dogs (one vaccinated and two unvaccinated) and CPV-2a in 1.72% vaccinated dog. Mean Ct values in qPCR for vaccinated dogs were higher than the unvaccinated dogs (p = 0.049), suggesting that vaccinated dogs shed less virus, even in clinical forms of CPV. CPV-2c was the dominant subtype infecting dogs in both vaccinated and unvaccinated cases. Faecal antigen testing failed to identify a substantial proportion of CPV-2c infected dogs, likely due to low sensitivity. The faecal samples from apparently healthy dogs (n = 21) showed negative results in all three tests. Negative CPV faecal antigen results should be viewed with caution until they are confirmed by molecular methods.

To mitigate the negative impacts of invasive rabbits in Australia, land managers are permitted to release the biocontrol virus, rabbit hemorrhagic disease virus (RHDV), to reduce rabbit numbers. However, it is strongly recommended that RHDV is not released when young rabbits are present in the population as infection in this cohort is sublethal and induces life‐long virus immunity. The recruitment of these rabbits into the breeding population may make the population harder to control in future, potentially leading to increasing rather than decreased population size. To investigate whether the recommended release guidelines are followed, we obtained data on the supply and release of RHDV by land managers. We then used generalized additive models to investigate Australia‐wide and state‐specific annual and long‐term temporal trends in the supply and release of RHDV. Half of all RHDV supply (47%) and three quarters of reported releases (74%) Australia‐wide occurred during the anticipated major rabbit breeding seasons and when the risk of immunizing young rabbits is greatest. We found evidence of both RHDV supply and release during the anticipated major rabbit breeding seasons in almost all states for which data existed. RHDV supply increased with below average annual rainfall. This may indicate a tendency for land managers to notice, and want to control, rabbits and their impacts more following drier years when both rabbits and their impacts are potentially more damaging. Our study raises concerns regarding the inappropriate release of RHDV by land managers and whether its supply should be restricted to ensure ongoing and effective management of invasive rabbits. More broadly, our study serves as a warning to other conservation and pest management activities reliant on land managers or citizens following implementation guidelines. In some cases, good intentions may have adverse outcomes. Rabbit hemorrhagic disease virus release timing.

In the present study, carboxymethylchitosan (CMCS) was prepared from chitosan, crosslinked with glutaraldehyde and evaluated in vitro as a potential carrier for site specific drug delivery of lercanidipine hydrochloride (LERH). LERH was incorporated at the time of crosslinking of CMCS. The chitosan was evaluated for its degree of deacetylation ( DD ) and average molecular weight, which were found to be 84·6% and 3·5 × 10 4 Da, respectively. The degree of substitution on prepared CMCS was found to be 0·68. All hydrogel formulations showed more than 86% and 77% yield and drug loading, respectively. The swelling behaviour of prepared hydrogels were checked in different pH values, 1·2, 6·8 and 7·4, indicated pH responsive swelling characteristic with very less swelling at pH 1·2 and quick swelling at pH 6·8 followed by linear swelling at pH 7·4 with slight increase. In vitro release profile was carried out at the same conditions as in swelling and drug release was found to be dependent on swelling of hydrogels and showed biphasic release pattern with non-fickian diffusion kinetics at higher pH. The carboxymethylation of chitosan, entrapment of drug and its interaction in prepared hydrogels were checked by FTIR, 1 H-NMR, DSC and p -XRD studies, which confirmed formation of CMCS from chitosan and absence of any significant chemical change in LERH after being entrapped in crosslinked hydrogel formulations. The surface morphology of formulation S 6 was checked before and after dissolution, revealed open channel like pores formation after dissolution.