Effects of Nutrition and Time of Weaning on Dry Season Liveweight Loss of Breeder Cows

In the seasonally dry tropics of northern Australia, breeder cows may lose up to 30% liveweight during the dry season when pasture is of low nutritive value. This is a major cause of low reproductive rates and high mortality. Weaning early in the dry season is effective to reduce this liveweight loss of the breeder (Holroyd et al. 1988). An experiment examined the dry season liveweight loss of breeders for a range of weaning times and levels of nutrition. From April to October through the dry season, 209 Bos indicus x Shorthorn cross cows 4-6 years of age grazed speargrass pastures in north Queensland. The cows had been joined with bulls from late January until April. Twenty-nine breeders had not suckled a calf during the previous wet season (DRY cows). In addition 180 cows lactating in April were weaned in late April, mid July or early September. The cows were allocated by stratified randomisation based on lactational status, stage of pregnancy and body condition to 15 x 40 ha paddocks. Five paddocks with low fertility soils provided LOW nutrition, while 10 paddocks with medium fertility soils and no supplementation or with supplementation provided MEDIUM and HIGH nutrition, respectively. The supplement consisted of molasses containing 14% urea offered ad libitum. Liveweight was measured at intervals and conceptus-free liveweight (CF-LW) calculated. Data were analyses by AOV within groups of paddocks. Animal production for a consuming world : proceedings of 9th Congress of the Asian-Australasian Association of Animal Production Societies [AAAP] and 23rd Biennial Conference of the Australian Society of Animal Production [ASAP] and 17th Annual Symposium of the University of Sydney, Dairy Research Foundation, [DRF]. 2-7 July 2000, Sydney, Australia.

Genetic variability of Tomato spotted wilt virus in Australia and validation of real time RT-PCR for its detection in single and bulked leaf samples

The potential for large-scale use of a sensitive real time reverse transcription polymerase chain reaction (RT-PCR) assay was evaluated for the detection of Tomato spotted wilt virus (TSWV) in single and bulked leaf samples by comparing its sensitivity with that of DAS-ELISA. Using total RNA extracted with RNeasy® or leaf soak methods, real time RT-PCR detected TSWV in all infected samples collected from 16 horticultural crop species (including flowers, herbs and vegetables), two arable crop species, and four weed species by both assays. In samples in which DAS-ELISA had previously detected TSWV, real time RT-PCR was effective at detecting it in leaf tissues of all 22 plant species tested at a wide range of concentrations. Bulk samples required more robust and extensive extraction methods with real time RT-PCR, but it generally detected one infected sample in 1000 uninfected ones. By contrast, ELISA was less sensitive when used to test bulked samples, once detecting up to 1 infected in 800 samples with pepper but never detecting more than 1 infected in 200 samples in tomato and lettuce. It was also less reliable than real time RT-PCR when used to test samples from parts of the leaf where the virus concentration was low. The genetic variability among Australian isolates of TSWV was small. Direct sequencing of a 587 bp region of the nucleoprotein gene (S RNA) of 29 isolates from diverse crops and geographical locations yielded a maximum of only 4.3% nucleotide sequence difference. Phylogenetic analysis revealed no obvious groupings of isolates according to geographic origin or host species. TSWV isolates, that break TSWV resistance genes in tomato or pepper did not differ significantly in the N gene region studied, indicating that a different region of the virus genome is responsible for this trait.

Detection of equine herpesvirus type 1 using a real-time polymerase chain reaction.

Equid herpesvirus 1 (EHV1) is a major disease of equids worldwide causing considerable losses to the horse industry. A variety of techniques, including PCR have been used to diagnose EHV1. Some of these PCRs were used in combination with other techniques such as restriction enzyme analysis (REA) or hybridisation, making them cumbersome for routine diagnostic testing and increasing the chances of cross-contamination. Furthermore, they involve the use of suspected carcinogens such as ethidium bromide and ultraviolet light. In this paper, we describe a real-time PCR, which uses minor groove-binding probe (MGB) technology for the diagnosis of EHV1. This technique does not require post-PCR manipulations thereby reducing the risk of cross-contamination. Most importantly, the technique is specific; it was able to differentiate EHV1 from the closely related member of the Alphaherpesvirinae, equid herpesvirus 4 (EHV4). It was not reactive with common opportunistic pathogens such as Escherichia coli, Klebsiella oxytoca, Pseudomonas aeruginosa and Enterobacter agglomerans often involved in abortion. Similarly, it did not react with equine pathogens such as Streptococcus equi, Streptococcus equisimilis, Streptococcus zooepidemicus, Taylorella equigenitalis and Rhodococcus equi, which also cause abortion. The results obtained with this technique agreed with results from published PCR methods. The assay was sensitive enough to detect EHV1 sequences in paraffin-embedded tissues and clinical samples. When compared to virus isolation, the test was more sensitive. This test will be useful for the routine diagnosis of EHV1 based on its specificity, sensitivity, ease of performance and rapidity.

Multiplex real-time PCR for the detection and differentiation of equid herpesvirus 1 (EHV-1) and equid herpesvirus 4 (EHV-4).

A multiplex real-time PCR was designed to detect and differentiate equid herpesvirus 1 (EHV-1) and equid herpesvirus 4 (EHV-4). The PCR targets the glycoprotein B gene of EHV-1 and EHV-4. Primers and probes were specific to each equine herpesvirus type and can be used in monoplex or multiplex PCRs, allowing the differentiation of these two closely related members of the Alphaherpesvirinae. The two probes were minor-groove binding probes (MGB?) labelled with 6-carboxy-fluorescein (FAM?) and VIC® for detection of EHV-1 and EHV-4, respectively. Ten EHV-1 isolates, six EHV-1 positive clinical samples, one EHV-1 reference strain (EHV-1.438/77), three EHV-4 positive clinical samples, two EHV-4 isolates and one EHV-4 reference strain (EHV-4 405/76) were included in this study. EHV-1 isolates, clinical samples and the reference strain reacted in the EHV-1 real-time PCR but not in the EHV-4 real-time PCR and similarly EHV-4 clinical samples, isolates and the reference strain were positive in the EHV-4 real-time PCR but not in the EHV-1 real-time PCR. Other herpesviruses, such as EHV-2, EHV-3 and EHV-5 were all negative when tested using the multiplex real-time PCR. When bacterial pathogens and opportunistic pathogens were tested in the multiplex real-time PCR they did not react with either system. The multiplex PCR was shown to be sensitive and specific and is a useful tool for detection and differentiation of EHV-1 and EHV-4 in a single reaction. A comprehensive equine herpesvirus disease investigation procedure used in our laboratory is also outlined. This procedure describes the combination of alphaherpesvirus multiplex real-time PCR along with existing gel-based PCRs described by other authors.

Responses in carcass lean pH at 24 hours post-mortem in lines of Large White pigs selected for growth rate on a fixed ration over a set time

A restricted maximum likelihood analysis applied to an animal model showed no significant differences (P > 0.05) in pH value of the longissimus dorsi measured at 24 h post-mortem (pH24) between high and low lines of Large White pigs selected over 4 years for post-weaning growth rate on restricted feeding. Genetic and phenotypic correlations between pH24 and production and carcass traits were estimated using all performance testing records combined with the pH24 measurements (5.05-7.02) on slaughtered animals. The estimate of heritability for pH24 was moderate (0.29 ± 0.18). Genetic correlations between pH24 and production or carcass composition traits, except for ultrasonic backfat (UBF), were not significantly different from zero. UBF had a moderate, positive genetic correlation with pH24 (0.24 ± 0.33). These estimates of genetic correlations affirmed that selection for increased growth rate on restricted feeding is likely to result in limited changes in pH24 and pork quality since the selection does not put a high emphasis on reduced fatness.

Time-dependent instantaneous mortality rates from multiple tagging experiments with exact times of release and recovery

Instantaneous natural mortality rates and a nonparametric hunting mortality function are estimated from a multiple-year tagging experiment with arbitrary, time-dependent fishing or hunting mortality. Our theory allows animals to be tagged over a range of times in each year, and to take time to mix into the population. Animals are recovered by hunting or fishing, and death events from natural causes occur but are not observed. We combine a long-standing approach based on yearly totals, described by Brownie et al. (1985, Statistical Inference from Band Recovery Data: A Handbook, Second edition, United States Fish and Wildlife Service, Washington, Resource Publication, 156), with an exact-time-of-recovery approach originated by Hearn, Sandland and Hampton (1987, Journal du Conseil International pour l'Exploration de la Mer, 43, 107-117), who modeled times at liberty without regard to time of tagging. Our model allows for exact times of release and recovery, incomplete reporting of recoveries, and potential tag shedding. We apply our methods to data on the heavily exploited southern bluefin tuna (Thunnus maccoyii).

Absolute quantitation of Marek's disease virus and Herpesvirus of turkeys in chicken lymphocyte, feather tip and dust samples using real-time PCR

The further development of Taqman quantitative real-time PCR (qPCR) assays for the absolute quantitation of Marek's disease virus serotype 1 (MDV1) and Herpesvirus of turkeys (HVT) viruses is described and the sensitivity and reproducibility of each assay reported. Using plasmid DNA copies, the lower limit of detection was determined to be 5 copies for the MDV1 assay and 75 copies for the HVT assay. Both assays were found to be highly reproducible for Ct values and calculated copy numbers with mean intra- and inter-assay coefficients of variation being less than 5% for Ct and 20% for calculated copy number. The genome copy number of MDV1 and HVT viruses was quantified in PBL and feather tips from experimentally infected chickens, and field poultry dust samples. Parallelism was demonstrated between the plasmid-based standard curves, and standard curves derived from infected spleen material containing both viral and host DNA, allowing the latter to be used for absolute quantification. These methods should prove useful for the reliable differentiation and absolute quantitation of MDV1 and HVT viruses in a wide range of samples.

Testing the simulation capability of APSIM-ORYZA under different levels of nitrogen fertiliser and transplanting time regimes in Korea

APSIM-ORYZA is a new functionality developed in the APSIM framework to simulate rice production while addressing management issues such as fertilisation and transplanting, which are particularly important in Korean agriculture. To validate the model for Korean rice varieties and field conditions, the measured yields and flowering times from three field experiments conducted by the Gyeonggi Agricultural Research and Extension Services (GARES) in Korea were compared against the simulated outputs for different management practices and rice varieties. Simulated yields of early-, mid- and mid-to-late-maturing varieties of rice grown in a continuous rice cropping system from 1997 to 2004 showed close agreement with the measured data. Similar results were also found for yields simulated under seven levels of nitrogen application. When different transplanting times were modelled, simulated flowering times ranged from within 3 days of the measured values for the early-maturing varieties, to up to 9 days after the measured dates for the mid- and especially mid-to-late-maturing varieties. This was associated with highly variable simulated yields which correlated poorly with the measured data. This suggests the need to accurately calibrate the photoperiod sensitivity parameters of the model for the photoperiod-sensitive rice varieties in Korea.

Estimating crop area using seasonal time series of enhanced vegetation index from MODIS satellite imagery

Cereal grain is one of the main export commodities of Australian agriculture. Over the past decade, crop yield forecasts for wheat and sorghum have shown appreciable utility for industry planning at shire, state, and national scales. There is now an increasing drive from industry for more accurate and cost-effective crop production forecasts. In order to generate production estimates, accurate crop area estimates are needed by the end of the cropping season. Multivariate methods for analysing remotely sensed Enhanced Vegetation Index (EVI) from 16-day Moderate Resolution Imaging Spectroradiometer (MODIS) satellite imagery within the cropping period (i.e. April-November) were investigated to estimate crop area for wheat, barley, chickpea, and total winter cropped area for a case study region in NE Australia. Each pixel classification method was trained on ground truth data collected from the study region. Three approaches to pixel classification were examined: (i) cluster analysis of trajectories of EVI values from consecutive multi-date imagery during the crop growth period; (ii) harmonic analysis of the time series (HANTS) of the EVI values; and (iii) principal component analysis (PCA) of the time series of EVI values. Images classified using these three approaches were compared with each other, and with a classification based on the single MODIS image taken at peak EVI. Imagery for the 2003 and 2004 seasons was used to assess the ability of the methods to determine wheat, barley, chickpea, and total cropped area estimates. The accuracy at pixel scale was determined by the percent correct classification metric by contrasting all pixel scale samples with independent pixel observations. At a shire level, aggregated total crop area estimates were compared with surveyed estimates. All multi-temporal methods showed significant overall capability to estimate total winter crop area. There was high accuracy at pixel scale (>98% correct classification) for identifying overall winter cropping. However, discrimination among crops was less accurate. Although the use of single-date EVI data produced high accuracy for estimates of wheat area at shire scale, the result contradicted the poor pixel-scale accuracy associated with this approach, due to fortuitous compensating errors. Further studies are needed to extrapolate the multi-temporal approaches to other geographical areas and to improve the lead time for deriving cropped-area estimates before harvest.

Is night-time wind direction important to best practice wild dog trapping and baiting?

We discovered a significant bias for wild dog scent station spoor (scats and scratches) to be positioned on the north-easterly side of roads and intersections. Counts of this spoor, 50 metres in each direction of north-south and east-west intersections were made in state forests near Roma in southwest Queensland, Cecil Plains on the Darling Downs and Maryborough on the coast during mating season in April/May 2007. While 51% of 190 and 83% of 120 scent station spoor were located on the north-eastern sector of the intersections at Cecil Plains and Roma respectively, spoor were more evenly distributed across all four sectors at Maryborough (n=47).

Are we focussing wild dog control the wrong time of the year and going about it the wrong way?

Our evaluation of the predation of calves by wild dogs in the 1990s found that the number of calves killed and frequency of years that calf losses occurred, is higher in baited areas compared to adjoining, non-baited areas of similar size. Calf losses were highest with poor seasonal conditions, low prey numbers and where baited areas were re-colonised by wild dogs soon after baiting. We monitored wild dog “activity” before and after 35 baiting programs in southwest, central west and far north Queensland between 1994 and 2006 and found change in activity depends on the timing of the baiting. Baiting programs conducted between October and April show an increase in dog activity post-baiting (average increase of 219.1%, SEM 100.9, n=9, for programs conducted in October and November; an increase of 82.5%, SEM 54.5, n=7 for programs conducted in March and April; and a decrease in activity of 46.5%, SEM 10.2, n=19 for programs conducted between May and September). We monitored the seasonal activity and dispersal of wild dogs fitted with satellite transmitters 2006 to present. We have found that: • Activity of breeding males and females, whilst rearing and nurturing pups, is focussed around the den between July to September and away from areas of human activity. Activity of breeding groups appears to avoid locations of human activity until juveniles become independent (around late November). • While independent and solitary yearlings often have unstable, elliptically-shaped territories in less favourable areas, members of breeding groups have territories that appear seasonally stable and circular located in more favourable habitats. • Extra-territorial forays of solitary yearlings can be huge, in excess of 200 km. The largest forays we have monitored have occurred when the activity of pack members is focussed around rearing pups and juveniles (August to November). • Where wild dogs have dispersed or had significant territorial expansion, it has occurred within days of baiting programs and onto recently baited properties. • The wild dogs we have tracked have followed netting barrier fences for hundreds of kilometres and lived adjacent to or bypassed numerous grids in the barrier. Based on these studies, we conclude that a proportion of the perceived decline in dog activity between May and September, post baiting, is due to a decline in dog activity in areas associated with human activity. The increase in dog activity post-baiting between October and May (and increased calf predation on baited properties) is likely caused by wild dogs dispersing (juveniles and yearlings) or expanding (adults) their territory into baited, now ‘vacant’, areas. We hypothesise that baiting programs should be focussed in summer and autumn commencing late November as soon as juveniles become independent of adults. We also hypothesise that instead of large, annual or semi-annual baiting programs, laying the same number of baits over 4-6 weeks may be more effective. These hypotheses need to be tested through an adaptive management project.

Effect of foliar herbicides on the germination and viability of Siam weed (Chromolaena odorata) seeds located on plants at the time of application.

This paper reports a field study undertaken to determine if the foliar application of herbicides fluroxypyr (150 mL 100 L-1 a.i.) and metsulfuron-methyl (12 g 100 L-1 a.i.) were capable of reducing the germination and viability of Chromolaena odorata (L.) R.M.King & H.Rob. (Siam weed) seeds at three different stages of maturity. After foliar application of fluroxypyr germination of mature seeds was reduced by 88% and intermediate and immature seeds were reduced by 100%, compared to the control. Fluroxypyr also reduced the viability of mature, intermediate and immature seeds by 79, 89 and 67% respectively, compared to the control. Metsulfuron-methyl reduced germination of intermediate and immature seeds by 53 and 99% respectively compared to the control. Viability was also reduced by 74 and 96% respectively, compared to the control. Mature seeds were not affected by metsulfuron-methyl as germination and viability increased by 2% and 1% respectively, as compared to the control. These results show that these herbicides are capable of reducing the amount of viable seed entering the seed bank. However depending on the treatment and stage of seed development a percentage of seeds on the plants will remain viable and contribute to the seed bank. This information is of value to Siam weed eradication teams as plants are most easily located and subsequently treated at the time of flowering. Knowledge of the impact of control methods on seeds at various stages of development will help determine the most suitable chemical control option for a given situation.

Specific detection of the Old World screwworm fly, Chrysomya bezziana, in bulk fly trap catches using real-time PCR.

The Old World screwworm fly (OWS), Chrysomya bezziana Villeneuve (Diptera: Calliphoridae), is a myiasis-causing blowfly of major concern for both animals and humans. Surveillance traps are used in several countries for early detection of incursions and to monitor control strategies. Examination of surveillance trap catches is time-consuming and is complicated by the presence of morphologically similar flies that are difficult to differentiate from Ch. bezziana, especially when the condition of specimens is poor. A molecular-based method to confirm or refute the presence of Ch. bezziana in trap catches would greatly simplify monitoring programmes. A species-specific real-time polymerase chain reaction (PCR) assay was designed to target the ribosomal DNA internal transcribed spacer 1 (rDNA ITS1) of Ch. bezziana. The assay uses both species-specific primers and an OWS-specific Taqman MGB probe. Specificity was confirmed against morphologically similar and related Chrysomya and Cochliomyia species. An optimal extraction protocol was developed to process trap catches of up to 1000 flies and the assay is sensitive enough to detect one Ch. bezziana in a sample of 1000 non-target species. Blind testing of 29 trap catches from Australia and Malaysia detected Ch. bezziana with 100% accuracy. The probability of detecting OWS in a trap catch of 50 000 flies when the OWS population prevalence is low (one in 1000 flies) is 63.6% for one extraction. For three extractions (3000 flies), the probability of detection increases to 95.5%. The real-time PCR assay, used in conjunction with morphology, will greatly increase screening capabilities in surveillance areas where OWS prevalence is low.

Positive results in a real-time PCR for type A influenza associated with the use of an inactivated vaccine.

A real-time reverse transcription polymerase chain reaction (qRT-PCR) test for the matrix gene of type A influenza viruses was used during the 2007 Australian equine influenza (EI) outbreak in order to confirm diagnosis and, later, eradication of the virus. During the EI outbreak, horses being exported required vaccination and individual proof of freedom from EI. At the end of the outbreak, positive results were obtained from four horses destined for export, because of contamination of the samples with the vaccine. This report highlights the need for EI testing and vaccination to occur on separate days and with the collection of swabs for testing to precede vaccination.

Detection and differentiation of bovine herpesvirus 1 and 5 using a multiplex real-time polymerase chain reaction.

A multiplex real-time PCR was developed for the detection and differentiation of two closely related bovine herpesviruses 1 (BoHV-1) and 5 (BoHV-5). The multiplex real-time PCR combines a duplex real-time PCR that targets the DNA polymerase gene of BoHV-1 and BoHV-5 and a real-time PCR targeting mitochondrial DNA, as a house-keeping gene, described previously by Cawthraw et al. (2009). The assay correctly identified 22 BoHV-1 and six BoHV-5 isolates from the Biosecurity Sciences Laboratory virus collection. BoHV-1 and BoHV-5 were also correctly identified when incorporated in spiked semen and brain tissue samples. The detection limits of the duplex assay were 10 copies of BoHV-1 and 45 copies of BoHV-5. The multiplex real-time PCR had reaction efficiencies of 1.04 for BoHV-1 and 1.08 for BoHV-5. Standard curves relating Ct value to template copy number had correlation coefficients of 0.989 for BoHV-1 and 0.978 for BoHV-5. The assay specificity was demonstrated by testing bacterial and viral DNA from pathogens commonly isolated from bovine respiratory and reproductive tracts. The validated multiplex real-time PCR was used to detect and differentiate BoHV-1 and BoHV-5 in bovine clinical samples with known histories.