The in vivo fate of nanoparticles and nanoparticle-loaded microcapsules after oral administration in mice: Evaluation of their potential for colon-specific delivery

Anti-cancer drug loaded-nanoparticles (NPs) or encapsulation of NPs in colon-targeted delivery systems shows potential for increasing the local drug concentration in the colon leading to improved treatment of colorectal cancer. To investigate the potential of the NP-based strategies for colon-specific delivery, two formulations, free Eudragit® NPs and enteric-coated NP-loaded chitosan–hypromellose microcapsules (MCs) were fluorescently-labelled and their tissue distribution in mice after oral administration was monitored by multispectral small animal imaging. The free NPs showed a shorter transit time throughout the mouse digestive tract than the MCs, with extensive excretion of NPs in faeces at 5 h. Conversely, the MCs showed complete NP release in the lower region of the mouse small intestine at 8 h post-administration. Overall, the encapsulation of NPs in MCs resulted in a higher colonic NP intensity from 8 h to 24 h post-administration compared to the free NPs, due to a NP ‘guarding’ effect of MCs during their transit along mouse gastrointestinal tract which decreased NP excretion in faeces. These imaging data revealed that this widely-utilised colon-targeting MC formulation lacked site-precision for releasing its NP load in the colon, but the increased residence time of the NPs in the lower gastrointestinal tract suggests that it is still useful for localised release of chemotherapeutics, compared to NP administration alone. In addition, both formulations resided in the stomach of mice at considerable concentrations over 24 h. Thus, adhesion of NP- or MC-based oral delivery systems to gastric mucosa may be problematic for colon-specific delivery of the cargo to the colon and should be carefully investigated for a full evaluation of particulate delivery systems.

Free Range Hens Use the Range More When the Outdoor Environment Is Enriched.

To evaluate the role of using forage, shade and shelterbelts in attracting birds into the range, three trials were undertaken with free range layers both on a research facility and on commercial farms. Each of the trials on the free range research facility in South Australia used a total of 120 laying hens (Hyline Brown). Birds were housed in an eco-shelter which had 6 internal pens of equal size with a free range area adjoining the shelter. The on-farm trials were undertaken on commercial free range layer farms in the Darling Downs in Southeast Queensland with bird numbers on farms ranging from 2,000-6,800 hens. The first research trial examined the role of shaded areas in the range; the second trial examined the role of forage and the third trial examined the influence of shelterbelts in the range. These treatments were compared to a free range area with no enrichment. Aggressive feather pecking was only observed on a few occasions in all of the trials due to the low bird numbers housed. Enriching the free range environment attracted more birds into the range. Shaded areas were used by 18% of the hens with a tendency (p = 0.07) for more hens to be in the paddock. When forage was provided in paddocks more control birds (55%) were observed in the range in morning than in the afternoon (30%) while for the forage treatments 45% of the birds were in the range both during the morning and afternoon. When shelterbelts were provided there was a significantly (p<0.05) higher % of birds in the range (43% vs. 24%) and greater numbers of birds were observed in areas further away from the poultry house. The results from the on-farm trials mirrored the research trials. Overall 3 times more hens used the shaded areas than the non shaded areas, with slightly more using the shade in the morning than in the afternoon. As the environmental temperature increased the number of birds using the outdoor shade also increased. Overall 17 times more hens used the shelterbelt areas than the control areas, with slightly more using the shelterbelts in the afternoon than in the morning. Approximately 17 times more birds used the forage areas compared to the control area in the corresponding range. There were 8 times more birds using a hay bale enriched area compared to the area with no hay bales. The use of forage sources (including hay bales) were the most successful method on-farm to attract birds into the range followed by shelterbelts and artificial shade. Free range egg farmers are encouraged to provide pasture, shaded areas and shelterbelts to attract birds into the free range.

The Distribution of Henipaviruses in Southeast Asia and Australasia: Is Wallace's Line a Barrier to Nipah Virus?

Nipah virus (NiV) (Genus Henipavirus) is a recently emerged zoonotic virus that causes severe disease in humans and has been found in bats of the genus Pteropus. Whilst NiV has not been detected in Australia, evidence for NiV-infection has been found in pteropid bats in some of Australia's closest neighbours. The aim of this study was to determine the occurrence of henipaviruses in fruit bat (Family Pteropodidae) populations to the north of Australia. In particular we tested the hypothesis that Nipah virus is restricted to west of Wallace's Line. Fruit bats from Australia, Papua New Guinea, East Timor and Indonesia were tested for the presence of antibodies to Hendra virus (HeV) and Nipah virus, and tested for the presence of HeV, NiV or henipavirus RNA by PCR. Evidence was found for the presence of Nipah virus in both Pteropus vampyrus and Rousettus amplexicaudatus populations from East Timor. Serology and PCR also suggested the presence of a henipavirus that was neither HeV nor NiV in Pteropus alecto and Acerodon celebensis. The results demonstrate the presence of NiV in the fruit bat populations on the eastern side of Wallace's Line and within 500 km of Australia. They indicate the presence of non-NiV, non-HeV henipaviruses in fruit bat populations of Sulawesi and Sumba and possibly in Papua New Guinea. It appears that NiV is present where P. vampyrus occurs, such as in the fruit bat populations of Timor, but where this bat species is absent other henipaviruses may be present, as on Sulawesi and Sumba. Evidence was obtained for the presence henipaviruses in the non-Pteropid species R. amplexicaudatus and in A. celebensis. The findings of this work fill some gaps in knowledge in geographical and species distribution of henipaviruses in Australasia which will contribute to planning of risk management and surveillance activities.

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.

Agricultural intensification, priming for persistence and the emergence of Nipah virus: a lethal bat-borne zoonosis

Emerging zoonoses threaten global health, yet the processes by which they emerge are complex and poorly understood. Nipah virus (NiV) is an important threat owing to its broad host and geographical range, high case fatality, potential for human-to-human transmission and lack of effective prevention or therapies. Here, we investigate the origin of the first identified outbreak of NiV encephalitis in Malaysia and Singapore. We analyse data on livestock production from the index site (a commercial pig farm in Malaysia) prior to and during the outbreak, on Malaysian agricultural production, and from surveys of NiV's wildlife reservoir (flying foxes). Our analyses suggest that repeated introduction of NiV from wildlife changed infection dynamics in pigs. Initial viral introduction produced an explosive epizootic that drove itself to extinction but primed the population for enzootic persistence upon reintroduction of the virus. The resultant within-farm persistence permitted regional spread and increased the number of human infections. This study refutes an earlier hypothesis that anomalous El Nino Southern Oscillation-related climatic conditions drove emergence and suggests that priming for persistence drove the emergence of a novel zoonotic pathogen. Thus, we provide empirical evidence for a causative mechanism previously proposed as a precursor to widespread infection with H5N1 avian influenza and other emerging pathogens.

Systemic toxoplasmosis in captive flying-foxes

Systemic toxoplasmosis caused by Toxoplasma gondii was diagnosed in two juvenile, captive flying-foxes (Pteropus conspicillatus and P. scapulatus), which died following respiratory distress. One animal displayed clinical signs suggestive of neurological disease. This is the first report of this disease in megachiropteran bats and adds to the list of differential diagnoses for both systemic and neurological disease in these animals. The role of captivity in the exposure and development of the disease is discussed.

Foraging Behaviour and Landscape Utilisation by the Endangered Golden-Crowned Flying Fox (Acerodon jubatus), The Philippines

Species of Old World fruit-bats (family Pteropodidae) have been identified as the natural hosts of a number of novel and highly pathogenic viruses threatening livestock and human health. We used GPS data loggers to record the nocturnal foraging movements of Acerodon jubatus, the Golden-crowned flying fox in the Philippines to better understand the landscape utilisation of this iconic species, with the dual objectives of pre-empting disease emergence and supporting conservation management. Data loggers were deployed on eight of 54 A. jubatus (two males and six females) captured near Subic Bay on the Philippine island of Luzon between 22 November and 2 December 2010. Bodyweight ranged from 730 g to 1002 g, translating to a weight burden of 3–4% of bodyweight. Six of the eight loggers yielded useful data over 2–10 days, showing variability in the nature and range of individual bat movements. The majority of foraging locations were in closed forest and most were remote from evident human activity. Forty-six discrete foraging locations and five previously unrecorded roost locations were identified. Our findings indicate that foraging is not a random event, with the majority of bats exhibiting repetitious foraging movements night-to-night, that apparently intact forest provides the primary foraging resource, and that known roost locations substantially underestimate the true number (and location) of roosts. Our initial findings support policy and decision-making across perspectives including landscape management, species conservation, and potentially disease emergence.

The application of one health approaches to henipavirus research

Henipaviruses cause fatal infection in humans and domestic animals. Transmission from fruit bats, the wildlife reservoirs of henipaviruses, is putatively driven (at least in part) by anthropogenic changes that alter host ecology. Human and domestic animal fatalities occur regularly in Asia and Australia, but recent findings suggest henipaviruses are present in bats across the Old World tropics. We review the application of the One Health approach to henipavirus research in three locations: Australia, Malaysia and Bangladesh. We propose that by recognising and addressing the complex interaction among human, domestic animal and wildlife systems, research within the One Health paradigm will be more successful in mitigating future human and domestic animal deaths from henipavirus infection than alternative single-discipline approaches. © Springer-Verlag Berlin Heidelberg 2013.

Nipah Virus in the Fruit Bat Pteropus vampyrus in Sumatera, Indonesia

Nipah virus causes periodic livestock and human disease with high case fatality rate, and consequent major economic, social and psychological impacts. Fruit bats of the genus Pteropus are the natural reservoir. In this study, we used real time PCR to screen the saliva and urine of P. vampyrus from North Sumatera for Nipah virus genome. A conventional reverse transcriptase (RT-PCR) assay was used on provisionally positive samples to corroborate findings. This is the first report of Nipah virus detection in P. vampyrus in Sumatera, Indonesia.

In vivo delivery of bovine viral diahorrea virus, E2 protein using hollow mesoporous silica nanoparticles

Our work focuses on the application of mesoporous silica nanoparticles as a combined delivery vehicle and adjuvant for vaccine applications. Here we present results using the viral protein, E2, from bovine viral diarrhoea virus (BVDV). BVDV infection occurs in the target species of cattle and sheep herds worldwide and is therefore of economic importance. E2 is a major immunogenic determinant of BVDV and is an ideal candidate for the development of a subunit based nanovaccine using mesoporous silica nanoparticles. Hollow type mesoporous silica nanoparticles with surface amino functionalisation (termed HMSA) were characterised and assessed for adsorption and desorption of E2. A codon-optimised version of the E2 protein (termed Opti-E2) was produced in Escherichia coli. HMSA (120 nm) had an adsorption capacity of 80 [small mu ]g Opti-E2 per mg HMSA and once bound E2 did not dissociate from the HMSA. Immunisation studies in mice with a 20 [small mu ]g dose of E2 adsorbed to 250 [small mu ]g HMSA was compared to immunisation with Opti-E2 (50 [small mu ]g) together with the traditional adjuvant Quillaja saponaria Molina tree saponins (QuilA, 10 [small mu ]g). The humoral responses with the Opti-E2/HMSA nanovaccine although slightly lower than those obtained for the Opti-E2 + QuilA group demonstrated that HMSA particles are an effective adjuvant that stimulated E2-specific antibody responses. Importantly the cell-mediated immune responses were consistently high in all mice immunised with Opti-E2/HMSA nanovaccine formulation. Therefore we have shown the Opti-E2/HMSA nanoformulation acts as an excellent adjuvant that gives both T-helper 1 and T-helper 2 mediated responses in a small animal model. This study has provided proof-of-concept towards the development of an E2 subunit nanoparticle based vaccine.

Health assessment of the Christmas Island flying fox (Pteropus melanotus natalis)

During July/August 2010, 28 Christmas Island flying foxes (Pteropus melanotus natalis) were captured and anesthetized for examination, sample collection, and release to determine the potential role of disease in recent population declines. Measurements and samples were taken for morphologic, hematologic, biochemical, and parasitologic analysis. These are the first blood reference ranges reported for this species. These data are being used to inform investigations into conservation status and population management strategies for the Christmas Island flying fox.

Top-predator control-induced trophic cascades: an alternative hypothesis to the conclusion of Colman et al.

Recently argued that observed positive relationships between dingoes and small mammals were a result of top-down processes whereby lethal dingo control reduced dingoes and increased mesopredators and herbivores, which then suppressed small mammals. Here, I show that the prerequisite negative effects of dingo control on dingoes were not shown, and that the same positive relationships observed may simply represent well-known bottom-up processes whereby more generalist predators are found in places with more of their preferred prey. Identification of top-predator controlinduced trophic cascades first requires demonstration of some actual effect of control on predators, typically possible only through manipulative experiments with the ability to identify cause and effect.

Demographic and functional responses of wild dogs to poison baiting

Lethal control of wild dogs - that is Dingo (Canis lupus dingo) and Dingo/Dog (Canis lupus familiaris) hybrids - to reduce livestock predation in Australian rangelands is claimed to cause continental-scale impacts on biodiversity. Although top predator populations may recover numerically after baiting, they are predicted to be functionally different and incapable of fulfilling critical ecological roles. This study reports the impact of baiting programmes on wild dog abundance, age structures and the prey of wild dogs during large-scale manipulative experiments. Wild dog relative abundance almost always decreased after baiting, but reductions were variable and short-lived unless the prior baiting programme was particularly effective or there were follow-up baiting programmes within a few months. However, age structures of wild dogs in baited and nil-treatment areas were demonstrably different, and prey populations did diverge relative to nil-treatment areas. Re-analysed observations of wild dogs preying on kangaroos from a separate study show that successful chases that result in attacks of kangaroos by wild dogs occurred when mean wild dog ages were higher and mean group size was larger. It is likely that the impact of lethal control on wild dog numbers, group sizes and age structures compromise their ability to handle large difficult-to-catch prey. Under certain circumstances, these changes sometimes lead to increased calf loss (Bos indicus/B. taurus genotypes) and kangaroo numbers. Rangeland beef producers could consider controlling wild dogs in high-risk periods when predation is more likely and avoid baiting at other times.

Ecological dynamics of emerging bat virus spillover

Viruses that originate in bats may be the most notorious emerging zoonoses that spill over from wildlife into domestic animals and humans. Understanding how these infections filter through ecological systems to cause disease in humans is of profound importance to public health. Transmission of viruses from bats to humans requires a hierarchy of enabling conditions that connect the distribution of reservoir hosts, viral infection within these hosts, and exposure and susceptibility of recipient hosts. For many emerging bat viruses, spillover also requires viral shedding from bats, and survival of the virus in the environment. Focusing on Hendra virus, but also addressing Nipah virus, Ebola virus, Marburg virus and coronaviruses, we delineate this cross-species spillover dynamic from the within-host processes that drive virus excretion to land-use changes that increase interaction among species. We describe how land-use changes may affect co-occurrence and contact between bats and recipient hosts. Two hypotheses may explain temporal and spatial pulses of virus shedding in bat populations: episodic shedding from persistently infected bats or transient epidemics that occur as virus is transmitted among bat populations. Management of livestock also may affect the probability of exposure and disease. Interventions to decrease the probability of virus spillover can be implemented at multiple levels from targeting the reservoir host to managing recipient host exposure and susceptibility.

Haematology and Plasma Biochemistry of Wild Black Flying-Foxes, (Pteropus alecto) in Queensland, Australia

This paper establishes reference ranges for hematologic and plasma biochemistry values in wild Black flying-foxes (Pteropus alecto) captured in South East Queensland, Australia. Values were found to be consistent with those of other Pteropus species. Four hundred and forty-seven animals were sampled over 12 months and significant differences were found between age, sex, reproductive and body condition cohorts in the sample population. Mean values for each cohort fell within the determined normal adult reference range, with the exception of elevated levels of alkaline phosphatase in juvenile animals. Hematologic and biochemistry parameters of injured animals showed little or no deviation from the normal reference values for minor injuries, while two animals with more severe injury or abscessation showed leucocytosis, anaemia, thrombocytosis, hyperglobulinemia and hypoalbuminemia.