Detection of a putative hemolysin operon, hhdBA, of Haemophilus parasuis from pigs with Glasser disease

The aim of the current study was to investigate whether polymerase chain reaction amplification of 16S ribosomal (r)RNA and a putative hemolysin gene operon, hhdBA, can be used to monitor live pigs for the presence of Haemophilus parasuis and predict the virulence of the strains present. Nasal cavity swabs were taken from 30 live, healthy, 1- to 8-week-old pigs on a weekly cycle from a commercial Thai nursery pig herd. A total of 27 of these pigs (90%) tested positive for H. parasuis as early as week 1 of age. None of the H. parasuis-positive samples from healthy pigs was positive for the hhdBA genes. At the same pig nursery, swab samples from nasal cavity, tonsil, trachea, and lung, and exudate samples from pleural/peritoneal cavity were taken from 30 dead pigs displaying typical pathological lesions consistent with Glasser disease. Twenty-two of 140 samples (15.7%) taken from 30 diseased pigs yielded a positive result for H. parasuis. Samples from the exudate (27%) yielded the most positive results, followed by lung, tracheal swab, tonsil, and nasal swab, respectively. Out of 22 positive samples, 12 samples (54.5%) harbored hhdA and/or hhdB genes. Detection rates of hhdA were higher than hhdB. None of the H. parasuis-positive samples taken from nasal cavity of diseased pigs tested positive for hhdBA genes. More work is required to determine if the detection of hhdBA genes is useful for identifying the virulence potential of H. parasuis field isolates.

Electrotransformation of Haemophilus parasuis with in vitro modified DNA based on a novel shuttle vector

The objective of the present study was to establish a valid transformation method of Haemophilus parasuis, the causative agent of Glasser's disease in pigs, using a novel H. parasuis-Escherichia coli shuttle vector. A 4.2 kb endogenous plasmid pYC93 was extracted from an H. parasuis field isolate and completely sequenced. Analysis of pYC93 revealed a region approximately 800 bp showing high homology with the defined replication origin oriV of pLS88, a native plasmid identified in Haemophilus ducreyi. Based on the origin region of pYC93, E. coli cloning vector pBluescript SK(+) and the Tn903 derived kanamycin cassette, a shuttle vector pSHK4 was constructed by overlapping PCR strategy. When electroporation of the 15 H. parasuis serovar reference strains and one clinical isolate SH0165 with pSHK4 was performed, only one of these strains yielded transformants with an efficiency of 8.5 x 10(2) CFUhlg of DNA. Transformation efficiency was notably increased (1.3 x 10(5) CFU/mu g of DNA) with vector DNA reisolated from the homologous transformants. This demonstrated that restriction-modification systems were involved in the barrier to transformation of H. parasuis. By utilizing an in vitro DNA modification method with cell-free extracts of the host H. parasuis strains, 15 out of 16 strains were transformable. The novel shuttle vector pSHK4 and the established electrotransformation method constitute useful tools for the genetic manipulation of H. parasuis to gain a better understanding of the pathogen. (C) 2011 Elsevier B.V. All rights reserved.

An exploratory study to assess the activity of the acarine growth inhibitor, fluazuron, against Sarcoptes scabei infestation in pigs

Background: The most common treatments for scabies in human and veterinary settings are topical 5% permethrin or systemic treatment with ivermectin. However, these treatments have very little activity against arthropod eggs, and therefore repeated treatment is frequently required. In-vitro, biochemical and molecular studies have demonstrated that human mites are becoming increasingly resistant to both acaricides. To identify alternate acaricides, we undertook a pilot study of the in vivo activity of the benzoylphenyl urea inhibitor of chitin synthesis, fluazuron, in pigs with sarcoptic mange. Findings: Pigs (n = 5) were infested with S. scabei var suis, and randomised to treatment at the start of peak infestation with fluazuron at a dose of 10 mg/kg/day per os for 7 days (n = 3) or no treatment (n = 2). Clinical scores, skin scrapings for mite counts and blood sampling for pharmacokinetic analysis were undertaken. Fluazuron was well absorbed in treated pigs with measureable blood levels up to 4 weeks post treatment. No adverse effects were observed. Modest acaricidal activity of the compound was observed, with a reduction in severity of skin lesions in treated pigs, as well as a reduction in number of scabies mite's early life stages. Conclusions: The moderate efficacy of fluazuron against scabies mites indicates a lead to the development of alternate treatments for scabies, such as combination therapies that maybe applicable for human use in the future.

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.

Impacts and management of wild pigs Sus scrofa in Australia

Globally, wild or feral pigs Sus scrofa are a widespread and important pest. Mitigation of their impacts requires a sound understanding of those impacts and the benefits and limitations of different management approaches. Here, we review published and unpublished studies to provide a synopsis of contemporary understanding of wild pig impacts and management in Australia, and to identify important shortcomings. Wild pigs can have important impacts on biodiversity values, ecosystem functioning and agricultural production. However, many of these impacts remain poorly described, and therefore, difficult to manage effectively. Many impacts are highly variable, and innovative experimental and analytical approaches may be necessary to elucidate them. Most contemporary management programmes use lethal techniques to attempt to reduce pig densities, but it is often unclear how effective they are at reducing damage. We conclude that greater integration of experimental approaches into wild pig management programmes is necessary to improve our understanding of wild pig impacts, and our ability to manage those impacts effectively and efficiently.

Evaluation of ELISA coupled with Western blot as a surveillance tool for Trichinella infection in wild boar (Sus scrofa)

Trichinella surveillance in wildlife relies on muscle digestion of large samples which are logistically difficult to store and transport in remote and tropical regions as well as labour-intensive to process. Serological methods such as enzyme-linked immunosorbent assays (ELISAs) offer rapid, cost-effective alternatives for surveillance but should be paired with additional tests because of the high false-positive rates encountered in wildlife. We investigated the utility of ELISAs coupled with Western blot (WB) in providing evidence of Trichinella exposure or infection in wild boar. Serum samples were collected from 673 wild boar from a high- and low-risk region for Trichinella introduction within mainland Australia, which is considered Trichinella-free. Sera were examined using both an 'in-house' and a commercially available indirect-ELISA that used excretory secretory (E/S) antigens. Cut-off values for positive results were determined using sera from the low-risk population. All wild boar from the high-risk region (352) and 139/321 (43.3%) of the wild boar from the low-risk region were tested by artificial digestion. Testing by Western blot using E/S antigens, and a Trichinella-specific real-time PCR was also carried out on all ELISA-positive samples. The two ELISAs correctly classified all positive controls as well as one naturally infected wild boar from Gabba Island in the Torres Strait. In both the high- and low-risk populations, the ELISA results showed substantial agreement (k-value = 0.66) that increased to very good (k-value = 0.82) when WB-positive only samples were compared. The results of testing sera collected from the Australian mainland showed the Trichinella seroprevalence was 3.5% (95% C.I. 0.0-8.0) and 2.3% (95% C.I. 0.0-5.6) using the in-house and commercial ELISA coupled with WB respectively. These estimates were significantly higher (P < 0.05) than the artificial digestion estimate of 0.0% (95% C.I. 0.0-1.1). Real-time PCR testing of muscle from seropositive animals did not detect Trichinella DNA in any mainland animals, but did reveal the presence of a second larvae-positive wild boar on Gabba Island, supporting its utility as an alternative, highly sensitive method in muscle examination. The serology results suggest Australian wildlife may have been exposed to Trichinella parasites. However, because of the possibility of non-specific reactions with other parasitic infections, more work using well-defined cohorts of positive and negative samples is required. Even if the specificity of the ELISAs is proven to be low, their ability to correctly classify the small number of true positive sera in this study indicates utility in screening wild boar populations for reactive sera which can be followed up with additional testing. (C) 2013 Elsevier B.V. All rights reserved.

Bentonite can decrease ammonia volatilisation losses from poultry litter: laboratory studies

Ammonia volatilisation from manure materials within poultry sheds can adversely affect production, and also represents a loss of fertiliser value from the spent litter. This study sought to compare the ability of alum and bentonite to decrease volatilisation losses of ammonia from spent poultry litter. An in-vessel volatilisation trial with air flushing, ammonia collection, and ammonia analysis was conducted over 64 days to evaluate the mitigation potential of these two materials. Water-saturated spent litter was incubated at 25°C in untreated condition (control) or with three treatments: an industry-accepted rate of alum [4% Al2(SO4)3·18H2O by dry mass of litter dry mass; ALUM], air-dry bentonite (127% by dry mass; BENT), or water-saturated bentonite (once again at 127% by dry mass; SATBENT). A high proportion of the nitrogen contained in the untreated spent litter was volatilised (62%). Bentonite additions were superior to alum additions at retaining spent litter ammonia (nitrogen losses: 15%, SATBENT; 34%, BENT; 54%, ALUM). Where production considerations favour comparable high rates of bentonite addition (e.g. where the litter is to be re-formulated as a fertiliser), this clay has potential to decrease ammonia volatilisation either in-shed or in spent litter stockpiles or formulated products, without the associated detrimental effect of alum on phosphorus availability.

Large-chamber methane and nitrous oxide measurements are comparable to the backward Lagrangian stochastic method

Measurement of individual emission sources (e.g., animals or pen manure) within intensive livestock enterprises is necessary to test emission calculation protocols and to identify targets for decreased emissions. In this study, a vented, fabric-covered large chamber (4.5 × 4.5 m, 1.5 m high; encompassing greater spatial variability than a smaller chamber) in combination with on-line analysis (nitrous oxide [N2O] and methane [CH4] via Fourier Transform Infrared Spectroscopy; 1 analysis min-1) was tested as a means to isolate and measure emissions from beef feedlot pen manure sources. An exponential model relating chamber concentrations to ambient gas concentrations, air exchange (e.g., due to poor sealing with the surface; model linear when ≈ 0 m3 s-1), and chamber dimensions allowed data to be fitted with high confidence. Alternating manure source emission measurements using the large-chamber and the backward Lagrangian stochastic (bLS) technique (5-mo period; bLS validated via tracer gas release, recovery 94-104%) produced comparable N2O and CH4 emission values (no significant difference at P < 0.05). Greater precision of individual measurements was achieved via the large chamber than for the bLS (mean ± standard error of variance components: bLS half-hour measurements, 99.5 ± 325 mg CH4 s-1 and 9.26 ± 20.6 mg N2O s-1; large-chamber measurements, 99.6 ± 64.2 mg CH4 s-1 and 8.18 ± 0.3 mg N2O s-1). The large-chamber design is suitable for measurement of emissions from manure on pen surfaces, isolating these emissions from surrounding emission sources, including enteric emissions. © © American Society of Agronomy, Crop Science Society of America, and Soil Science Society of America.

Use of a proposed antimicrobial susceptibility testing method for Haemophilus parasuis

The aim of this study was to examine the antimicrobial susceptibility of 97 Haemophilus parasuis cultured from Australian pigs. As there is no existing standard antimicrobial susceptibility technique available for H. parasuis, methods utilising the supplemented media, BA/SN for disc diffusion and test medium broth (TMB) for a microdilution technique, were initially evaluated with the reference strains recommended by the Clinical and Laboratory Standards Institute. The results of the media evaluation suggested that BA/SN and TMB can be used as suitable media for susceptibility testing of H. parasuis. The proposed microdilution technique was then used with 97 H. parasuis isolates and nine antimicrobial agents. The study found that Australian isolates showed elevated minimum inhibitory concentrations (MICs) for ampicillin (1%), penicillin (2%), erythromycin (7%), tulathromycin (9%), tilmicosin (22%), tetracycline (31%) and trimethoprim-sulfamethoxazole (40%). This study has described potential antimicrobial susceptibility methods for H. parasuis and has detected a low percentage of Australian H. parasuis isolates with elevated antimicrobial MICs.

Odour, dust and non-methane volatile organic-compound emissions from tunnel-ventilated layer-chicken sheds: a case study of two farms

An observational study was undertaken to measure odour and dust (PM10 and PM2.5) emission rates and identify non-methane volatile organic compounds (NMVOCs) and odorants in the exhaust air from two tunnel-ventilated layer-chicken sheds that were configured with multi-tiered cages and manure belts. The study sites were located in south-eastern Queensland and the West Gippsland region of Victoria, Australia. Samples were collected in summer and winter on sequential days across the manure-belt cleaning cycle. Odour emissions ranged from 58 to 512 ou/s per 1000 birds (0.03-0.27 ou/s.kg) and dust emission rates ranged 0.014-0.184 mg/s per 1000 birds for PM10 and 0.001-0.190 mg/s per 1000 birds for PM2.5. Twenty NMVOCs were identified, including three that were also identified as odorants using thermal desorption-gas chromatography-mass spectrometry/olfactometry analysis. Odour emission rates were observed to vary with the amount of manure accumulation on the manure belts, being lowest 2-4 days after removing manure. Odour emission rates were also observed to vary with diurnal and seasonal changes in ventilation rate. Dust emissions were observed to increase with ventilation rate but not with manure accumulation. Some NMVOCs were identified at both farms and in different seasons whereas others were observed only at one farm or in one season, indicating that odorant composition was influenced by farm-specific practices and season.

Good science for improving policy: greenhouse gas emissions from agricultural manures

Australia’s and New Zealand’s major agricultural manure management emission sources are reported to be, in descending order of magnitude: (1) methane (CH4) from dairy farms in both countries; (2) CH4 from pig farms in Australia; and nitrous oxide (N2O) from (3) beef feedlots and (4) poultry sheds in Australia. We used literature to critically review these inventory estimates. Alarmingly for dairy farm CH4 (1), our review revealed assumptions and omissions that when addressed could dramatically increase this emission estimate. The estimate of CH4 from Australian pig farms (2) appears to be accurate, according to industry data and field measurements. The N2O emission estimates for beef feedlots (3) and poultry sheds (4) are based on northern hemisphere default factors whose appropriateness for Australia is questionable and unverified. Therefore, most of Australasia’s key livestock manure management greenhouse gas (GHG) emission profiles are either questionable or are unsubstantiated by region-specific research. Encouragingly, GHG from dairy shed manure are relatively easy to mitigate because they are a point source which can be managed by several ‘close-to-market’ abatement solutions. Reducing these manure emissions therefore constitutes an opportunity for meaningful action sooner compared with the more difficult-to-implement and long-term strategies that currently dominate agricultural GHG mitigation research. At an international level, our review highlights the critical need to carefully reassess GHG emission profiles, particularly if such assessments have not been made since the compilation of original inventories. Failure to act in this regard presents the very real risk of missing the ‘low hanging fruit’ in the rush towards a meaningful response to climate change

Impacts on nontarget avian species from aerial meat baiting for feral pigs

Bait containing sodium fluoroacetate (1080) is widely used for the routine control of feral pigs in Australia. In Queensland, meat baits are popular in western and northern pastoral areas where they are readily accepted by feral pigs and can be distributed aerially. Field studies have indicated some levels of interference and consumption of baits by nontarget species and, based on toxicity data and the 1080 content of baits, many nontarget species (particularly birds and varanids) are potentially at risk through primary poisoning. While occasional deaths of species have been recorded, it remains unclear whether the level of mortality is sufficient to threaten the viability or ecological function of species. A series of field trials at Culgoa National Park in south-western Queensland was conducted to determine the effect of broadscale aerial baiting (1.7 baits per km2) on the density of nontarget avian species that may consume baits. Counts of susceptible bird species were conducted prior to and following aerial baiting, and on three nearby unbaited properties, in May and November 2011, and May 2012. A sample of baits was monitored with remote cameras in the November 2011 and May 2012 trials. Over the three baiting campaigns, there was no evidence of a population-level decline among the seven avian nontarget species that were monitored. Thirty per cent and 15% of baits monitored by remote cameras in the November 2011 and May 2012 trials were sampled by birds, varanids or other reptiles. These results support the continued use of 1080 meat baits for feral pig management in western Queensland and similar environs.

Crusted scabies is associated with increased IL-17 secretion by skin T cells

Scabies is an ectoparasitic infestation by the mite Sarcoptes scabiei. Although commonly self-limiting, a fraction of patients develop severely debilitating crusted scabies. The immune mechanisms underlying the development of crusted scabies are unclear, and undertaking longitudinal infection studies in humans is difficult. We utilized a porcine model to compare cellular immune responses in peripheral blood and skin of pigs with different clinical manifestations of scabies (n = 12), and in uninfected controls (n = 6). Although clinical symptoms were not evident until at least 4 weeks post-infestation, the numbers of peripheral IFNγ-secreting CD4+ T cells and γδ T cells increased in infected pigs from week 1 post-infestation. γδ T cells remained increased in the blood at week 15 post-infestation. At week 15, skin cell infiltrates from pigs with crusted scabies had significantly higher CD8+ T cell, γδ T cell and IL-17+ cell numbers than those with ordinary scabies. Peripheral IL-17 levels were not increased, suggesting that localized skin IL-17-secreting T cells may play a critical role in the pathogenesis of crusted scabies development. Given the potential of anti-IL-17 immunotherapy demonstrated for other inflammatory skin diseases, this study may provide a novel therapeutic avenue for patients with recurrent crusted scabies.

Sorghum genotypes differ in high temperature responses for seed set

Significant genotypic differences in tolerance of pollen germination and seed set to high temperatures have been shown in sorghum. However, it is unclear whether differences were associated with variation in either the threshold temperature above which reproductive processes are affected, or in the tolerance to increased temperature above that threshold. The objectives of this study were to (a) dissect known differences in heat tolerance for a range of sorghum genotypes into differences in the threshold temperature and tolerance to increased temperatures, (b) determine whether poor seed set under high temperatures can be compensated by increased seed mass, and (c) identify whether genotypic differences in heat tolerance in a controlled environment facility (CEF) can be reproduced in field conditions. Twenty genotypes were grown in a CEF under four day/night temperatures (31.9/21.0 °C, 32.8/21.0 °C, 36.1/21.0 °C, and 38.0/21.0 °C), and a subset of six genotypes was grown in the field under four different temperature regimes around anthesis. The novelty of the findings in this study related to differences in responsiveness to high temperature—genotypic differences in seed set percentage were found for both the threshold temperature and the tolerance to increased maximum temperature above that threshold. Further, the response of seed set to high temperature in the field study was well correlated to that in the CEF (R2 = 0.69), although the slope was significantly less than unity, indicating that heat stress effects may have been diluted under the variable field conditions. Poor seed set was not compensated by increased seed mass in either CEF or field environments. Grain yield was thus closely related to seed set percentage. This result demonstrates the potential for development of a low-cost field screening method to identify high-temperature tolerant varieties that could deliver sustainable yields under future warmer climates.

Assessing refrigerating and freezing effects on the biological/chemical composition of two livestock manures

Assessing storage impacts on manure properties is relevant to research associated with nutrient-use efficiency and greenhouse gas (GHG) emissions. We examined the impact of cold storage on physicochemical properties, biochemical methane-emitting potential (BMP) and the composition of microbial communities of beef feedlot manure and poultry broiler litter. Manures were analysed within 2 days of collection and after 2 and 8 weeks in refrigerated (4 °C) or frozen (–20 °C) storage. Compared with fresh manure, stored manures had statistically significant (p < 0.05) but comparatively minor (<10%) changes in electrical conductivity, chloride and ammonium concentrations. Refrigeration and freezing did not significantly affect (p > 0.05) BMP in both manure types. We did not detect ammonium- or nitrite-oxidising bacterial taxa (AOB, NOB) using fluorescence in situ hybridisation (FISH). Importantly, the viability of microbes was unchanged by storage. We conclude that storage at –20 °C or 4 °C adequately preserves the investigated traits of the studied manures for research aimed at improving nutrient cycling and reducing GHG emissions.