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Mulching with plastic sheeting, the use of plastic carriers in seed coatings, and irrigation with wastewater or contaminated surface water have resulted in plastics, and microplastics, becoming ubiquitous in agricultural soils. Once in the environment, plastic surfaces quickly become colonised by microbial biofilm comprised of a diverse microbial community. This so-called ‘plastisphere’ community can also include human pathogens, particularly if the plastic has been exposed to faecal contamination (e.g., from wastewater or organic manures and livestock faeces). The plastisphere is hypothesised to facilitate the survival and dissemination of pathogens, and therefore plastics in agricultural systems could play a significant role in transferring human pathogens to crops, particularly as microplastics adhering to ready to eat crops are difficult to remove by washing. In this paper we critically discuss the pathways for human pathogens associated with microplastics to interact with crop leaves and roots, and the potential for the transfer, adherence, and uptake of human pathogens from the plastisphere to plants. Globally, the concentration of plastics in agricultural soils are increasing, therefore, quantifying the potential for the plastisphere to transfer human pathogens into the food chain needs to be treated as a priority.

Recombination is a common feature of many positive-strand RNA viruses, playing an important role in virus evolution. However, to date, there is limited understanding of the mechanisms behind the process. Utilising in vitro assays, we have previously shown that the template-switching event of recombination is a random and ubiquitous process that often leads to recombinant viruses with imprecise genomes containing sequence duplications. Subsequently, a process termed resolution, that has yet to be mechanistically studied, removes these duplicated sequences resulting in a virus population of wild type length genomes. Using defined imprecise recombinant viruses together with Oxford Nanopore and Illumina high throughput next generation sequencing technologies we have investigated the process of resolution. We show that genome resolution involves subsequent rounds of template-switching recombination with viral fitness resulting in the survival of a small subset of recombinant genomes. This alters our previously held understanding that recombination and resolution are independent steps of the process, and instead demonstrates that viruses undergo frequent and continuous recombination events over a prolonged period until the fittest viruses, predominantly those with wild type length genomes, dominate the population.

Estimating population abundances, densities, and interspecific interactions are common goals in wildlife management. Camera traps have been used to estimate the abundance and density of a single species, and are useful for carnivores that occur at low densities. Spatial capture–recapture (SCR) models can be used to estimate abundance and density from a camera trap array when all, some, or no individuals in the population can be uniquely identified. These SCR models also estimate locations of individual activity centers, the spatial patterning of which could provide important information about interspecific interactions. We used SCR models to estimate abundances, densities, and activity centers of each of 3 carnivore species (i.e., dingo [Canis familiaris], red fox [Vulpes vulpes], and feral cat) using photographs from 1 camera trap array in southeastern Australia during September to November 2015. Some dingoes and feral cats were uniquely identifiable and therefore, we used a spatial mark–resight model for these species. We could not uniquely identify fox individuals, however, so we used a spatial unmarked (SUN) model for this species. Our estimated dingo density was 0.06/km². The fox (0.25/km²) and feral cat (0.16/km²) densities are within the ranges previously reported for these species in Australia. We obtained a relatively imprecise fox density estimate because we did not have detections of uniquely identifiable individuals; hence, the SUN model should be used as a last resort. We next modeled spatial dependence among the estimated activity centers for the 3 species using a spatial pair correlation function for a marked point process. Consistent with our expectations, the activity centers of dingoes and foxes were strongly negatively associated at distances of <1,000 m. Foxes and feral cats were also negatively associated at distances of <1,500 m. Surprisingly, dingoes and feral cats were positively associated at distances of >500 m, with no association evident at distances of <500 m. Our study extends the inferences that can be made from using a camera trap array and SCR methods to include spatial patterning and interspecific interactions, and provides new insights into the carnivore community of dingoes, foxes, and feral cats in southeastern Australia.

1. Predicting the current and potential distributions of established invasive species is critical for evaluating management options, but methods for differentiating these distributions have received little attention. In particular, there is uncertainty among invasive species managers about the value of information from incidental sightings compared to data from designed field surveys. This study compares the two approaches, and develops a unifying framework, using the case of invasive sambar deer Cervus unicolor in Victoria, Australia. 2. We first used 391 incidental sightings of sambar deer and 12 biophysical variables to construct a presence-only habitat suitability model using Maxent. We then used that model to stratify field sampling, with proportionately greater sampling of cells with high predicted habitat suitability. Field sampling, consisting of faecal pellet surveys, sign surveys and camera trapping, was conducted in 80 4-km² grid cells. A Bayesian state-space occupancy model was used to predict probability of suitable habitat from the field data. 3. The Maxent and occupancy models predicted similar spatial distributions of habitat suitability for sambar deer in Victoria and there was a strong positive correlation between the rankings of cells by the two approaches. The congruence of the two models suggests that any spatial and detection biases in the presence-only data were relatively unimportant in our study. 4. We predicted the extent of suitable habitat from the occupancy model using a threshold that gave a false negative error rate of 0·05. The current distribution was the suitable habitat within a kernel that had a 99·5% chance of including the presence locations pooled from incidental sightings and field surveys: the potential distribution was suitable habitat outside that kernel. Several discrete areas of potential distribution were identified as priorities for surveillance monitoring with the aim of detecting and managing incursions of sambar deer. 5. Synthesis and applications. Our framework enables managers to robustly estimate the current and potential distributions of established invasive species using either presence-only and/or presence-absence data. Managers can then focus control and/or containment actions within the current distribution and establish surveillance monitoring to detect incursions within the potential distribution.

Environmental and agricultural pollination services by honey bees, Apis mellifera, and honey production are compromised by high levels of annual colony losses globally. The majority are associated with disease caused by deformed wing virus (DWV), a positive-strand RNA virus, exacerbated by the ectoparasitic mite Varroa destructor. To improve honey bee health, a better understanding of virus transmission and pathogenesis is needed which requires the development of tools to study virus replication, transmission, and localisation. We report the use of reverse genetic (RG) systems for the predominant genetically distinct variants of DWV to address these questions. All RG-recovered viruses replicate within 24 h post-inoculation of pupae and could recapitulate the characteristic symptoms of DWV disease upon eclosion. Larvae were significantly less susceptible but could be infected orally and subsequently developed disease. Using genetically tagged RG DWV and an in vitro Varroa feeding system, we demonstrate virus replication in the mite by accumulation of tagged negative-strand viral replication intermediates. We additionally apply a modified DWV genome expressing a fluorescent reporter protein for direct in vivo observation of virus distribution in injected pupae or fed larvae. Using this, we demonstrate extensive sites of virus replication in a range of pupal tissues and organs and in the nascent wing buds in larvae fed high levels of virus, indicative of a direct association between virus replication and pathogenesis. These studies provide insights into virus replication kinetics, tropism, transmission, and pathogenesis, and produce new tools to help develop the understanding needed to control DWV-mediated colony losses.

There is much interest in understanding how anthropogenic food resources subsidise carnivore populations. Carcasses of hunter-shot ungulates are a potentially substantial food source for mammalian carnivores. The sambar deer (Rusa unicolor) is a large (≥ 150 kg) exotic ungulate that can be hunted throughout the year in south-eastern Australia, and hunters are not required to remove or bury carcasses. We investigated how wild dogs/dingoes and their hybrids (Canis lupus familiaris/dingo), red foxes (Vulpes vulpes) and feral cats (Felis catus) utilised sambar deer carcasses during the peak hunting seasons (i.e. winter and spring). We placed carcasses at 1-km intervals along each of six transects that extended 4-km into forest from farm boundaries. Visits to carcasses were monitored using camera traps, and the rate of change in edible biomass estimated at ∼ 14-day intervals. Wild dogs and foxes fed on 70% and 60% of 30 carcasses, respectively, but feral cats seldom (10%) fed on carcasses. Spatial and temporal patterns of visits to carcasses were consistent with the hypothesis that foxes avoid wild dogs. Wild dog activity peaked at carcasses 2 and 3 km from farms, a likely legacy of wild dog control, whereas fox activity peaked at carcasses 0 and 4 km from farms. Wild dog activity peaked at dawn and dusk, whereas nearly all fox activity occurred after dusk and before dawn. Neither wild dogs nor foxes remained at carcasses for long periods and the amount of feeding activity by either species was a less important predictor of the loss of edible biomass than season. Reasons for the low impacts of wild dogs and foxes on sambar deer carcass biomass include the spatially and temporally unpredictable distribution of carcasses in the landscape, the rapid rate of edible biomass decomposition in warm periods, low wild dog densities and the availability of alternative food resources.

Varroa destructor is an ectoparasitic mite associated with significant losses of honeybee colonies globally. The mite vectors a range of pathogenic viruses, the most important of which is the Deformed wing virus (DWV). In the absence of Varroa, DWV exists as a low-level, highly diverse virus population. However, when transmitted by Varroa, certain variants become highly elevated, and may become near-clonal and cause symptomatic infections. Mite transmission between colonies can occur when parasitised workers drift from or rob adjacent hives. These activities can result in elevated mite levels, but the resulting change in the DWV population, the primary determinant of winter colony losses, has not been determined. In reciprocal studies, we investigated the influence of the removal of mites, or their acquisition, on the DWV population. When mites were removed from heavily infested colonies, there was a striking and rapid reduction in virus load. Conversely, siting Varroa-naïve colonies in a mite-infested apiary resulted in the acquisition of mites and concomitant changes in the virus population. We observed both near-clonal and highly divergent virus populations regardless of titre, suggesting changes were stochastic and colony-specific. Our findings have implications for the outcome of strategies in areas with total or patchy implementation of Varroa control plans.

ABSTRACT Deformed wing virus (DWV) is the most important viral pathogen of honey bees. It usually causes asymptomatic infections but, when vectored by the ectoparasitic mite Varroa destructor, it is responsible for the majority of overwintering colony losses globally. Although DWV was discovered four decades ago, research has been hampered by the absence of an in vitro cell culture system or the ability to culture pure stocks of the virus. The recent developments of reverse genetic systems for DWV go some way to addressing these limitations. They will allow the investigation of specific questions about strain variation, host tropism and pathogenesis to be answered, and are already being exploited to study tissue tropism and replication in Varroa and non-Apis pollinators. Three areas neatly illustrate the advances possible with reverse genetic approaches: (i) strain variation and recombination, in which reverse genetics has highlighted similarities rather than differences between virus strains; (ii) analysis of replication kinetics in both honey bees and Varroa, in studies that likely explain the near clonality of virus populations often reported; and (iii) pathogen spillover to non-Apis pollinators, using genetically tagged viruses to accurately monitor replication and infection. Deformed wing virus is the most important viral pathogen of honey bees and along with its vector, Varroa destructor, is responsible for large-scale annual colony losses; recent advances in molecular biology have offered new insights into the biology of this honey bee virus, which address the determinants of tropism and pathogenesis, the importance of strain variation and the significance of possible pathogen spillover to other pollinating insects.

Deformed wing virus (DWV) is a persistent pathogen of European honey bees and the major contributor to overwintering colony losses. The prevalence of DWV in honey bees has led to significant concerns about spillover of the virus to other pollinating species. Bumble bees are both a major group of wild and commercially-reared pollinators. Several studies have reported pathogen spillover of DWV from honey bees to bumble bees, but evidence of a sustained viral infection characterized by virus replication and accumulation has yet to be demonstrated. Here we investigate the infectivity and transmission of DWV in bumble bees using the buff-tailed bumble bee Bombus terrestris as a model. We apply a reverse genetics approach combined with controlled laboratory conditions to detect and monitor DWV infection. A novel reverse genetics system for three representative DWV variants, including the two master variants of DWV—type A and B—was used. Our results directly confirm DWV replication in bumble bees but also demonstrate striking resistance to infection by certain transmission routes. Bumble bees may support DWV replication but it is not clear how infection could occur under natural environmental conditions.

The World Health Organization has classified five species of Candida as pathogens of critical concern on its recent ’Fungal Pathogen Priority List’. Candida albicans is the most common species aetiologically associated with human and animal mucosa; however, we know almost nothing about the environmental survival and the transfer pathways of C. albicans in natural environments. Here, we have isolated C. albicans from the faeces of newly arrived migratory geese in two agricultural fields in Scotland (the United Kingdom). All 14 confirmed isolates were characterised in terms of their pathogenicity (in a Galleria larval infection model) and resistance to four classes of antifungal drugs. Multilocus sequence typing (MLST) was used to assess the relatedness of these isolates with C. albicans isolated from the faeces of sheep grazing in the same field. Finally, mesocosms were used to test whether C. albicans in geese faeces could persist and remain viable when incorporated into agricultural soils. All C. albicans isolates were virulent, and exhibited resistance to at least one of the four classes of antifungal drugs against which they were screened. Maximum likelihood phylogenetic analysis of MLST sequences revealed that C. albicans isolated from goose and sheep faeces were relatively conserved, although they did cluster independently on separate clades. C. albicans remained viable in common agricultural soils, and after 60 days, the concentrations of all three representative C. albicans isolates had only declined by one log. The unrestricted movement of migratory birds provides a significant opportunity for the widespread dissemination of pathogens. Although C. albicans is not generally considered to be zoonotic, its introduction into new environments and subsequent persistence in agricultural systems has the potential to facilitate transmission between animal species or humans. Therefore, it is critical that more emphasis be put on monitoring animal migration and the potential for cross‐boundary movement of pathogens, particularly with climate change widening the geographic range for pathogen persistence.