Ruminococcus bromii, identification and isolation as a dominant community member in the rumen of cattle fed a barley diet

Aims: To identify dominant bacteria in grain (barley)-fed cattle for isolation and future use to increase the efficiency of starch utilization in these cattle. Methods and Results: Total DNA was extracted from samples of the rumen contents from eight steers fed a barley diet for 9 and 14 days. Bacterial profiles were obtained using denaturing gradient gel electrophoresis (DGGE) of the PCR-amplified V2/V3 region of the 16S rRNA genes from total bacterial DNA. Apparently dominant bands were excised and cloned, and the clone insert sequence was determined. One of the most common and dominant bacteria present was identified as Ruminococcus bromii. This species was subsequently isolated using traditional culture-based techniques and its dominance in the grain-fed cattle was confirmed using a real-time Taq nuclease assay (TNA) designed for this purpose. In some animals, the population of R. bromii reached densities above 1010R. bromii cell equivalents per ml or approximately 10% of the total bacterial population. Conclusions: Ruminococcus bromii is a dominant bacterial population in the rumen of cattle fed a barley-based diet. Significance and Impact of the Study: Ruminococcus bromii YE282 may be useful as a probiotic inoculant to increase the efficiency of starch utilization in barley-fed cattle. The combination of DGGE and real-time TNA has been an effective process for identifying and targeting for isolation, dominant bacteria in a complex ecosystem.

Integrating Rapid Phenotyping and Speed Breeding to Improve Stay-Green and Root Adaptation of Wheat in Changing, Water-Limited, Australian Environments

Temperatures have increased and in-crop rainfall decreased over recent decades in many parts of the Australian wheat cropping region. With these trends set to continue or intensify, improving crop adaptation in the face of climate change is particularly urgent in this, already drought-prone, cropping region. Importantly, improved performance under water-limitation must be achieved while retaining yield potential during more favourable seasons. A multi-trait-based approach to improve wheat yield and yield stability in the face of water-limitation and heat has been instigated in northern Australia using novel phenotyping techniques and a nested association mapping (NAM) approach. An innovative laboratory technique allows rapid root trait screening of hundreds of lines. Using soil grown seedlings, the method offers significant advantages over many other lab-based techniques. Another recently developed method allows novel stay-green traits to be quantified objectively for hundreds of genotypes in standard field trial plots. Field trials in multiple locations and seasons allow evaluation of targeted trait values and identification of superior germplasm. Traits, including yield and yield components are measured for hundreds of NAM lines in rain fed environments under various levels of water-limitation. To rapidly generate lines of interest, the University of Queensland “speed breeding” method is being employed, allowing up to 7 plant generations per annum. A NAM population of over 1000 wheat recombinant inbred lines has been progressed to the F5 generation within 18 months. Genotyping the NAM lines with the genome-wide DArTseq molecular marker system provides up to 40,000 markers. They are now being used for association mapping to validate QTL previously identified in bi-parental populations and to identify novel QTL for stay-green and root traits. We believe that combining the latest techniques in physiology, phenotyping, genetics and breeding will increase genetic progress toward improved adaptation to water-limited environments.

Using simulation techniques to investigate methods to determine resistance of helminths to anthelmintic treatment

The widespread and increasing resistance of internal parasites to anthelmintic control is a serious problem for the Australian sheep and wool industry. As part of control programmes, laboratories use the Faecal Egg Count Reduction Test (FECRT) to determine resistance to anthelmintics. It is important to have confidence in the measure of resistance, not only for the producer planning a drenching programme but also for companies investigating the efficacy of their products. The determination of resistance and corresponding confidence limits as given in anthelmintic efficacy guidelines of the Standing Committee on Agriculture (SCA) is based on a number of assumptions. This study evaluated the appropriateness of these assumptions for typical data and compared the effectiveness of the standard FECRT procedure with the effectiveness of alternative procedures. Several sets of historical experimental data from sheep and goats were analysed to determine that a negative binomial distribution was a more appropriate distribution to describe pre-treatment helminth egg counts in faeces than a normal distribution. Simulated egg counts for control animals were generated stochastically from negative binomial distributions and those for treated animals from negative binomial and binomial distributions. Three methods for determining resistance when percent reduction is based on arithmetic means were applied. The first was that advocated in the SCA guidelines, the second similar to the first but basing the variance estimates on negative binomial distributions, and the third using Wadley’s method with the distribution of the response variate assumed negative binomial and a logit link transformation. These were also compared with a fourth method recommended by the International Co-operation on Harmonisation of Technical Requirements for Registration of Veterinary Medicinal Products (VICH) programme, in which percent reduction is based on the geometric means. A wide selection of parameters was investigated and for each set 1000 simulations run. Percent reduction and confidence limits were then calculated for the methods, together with the number of times in each set of 1000 simulations the theoretical percent reduction fell within the estimated confidence limits and the number of times resistance would have been said to occur. These simulations provide the basis for setting conditions under which the methods could be recommended. The authors show that given the distribution of helminth egg counts found in Queensland flocks, the method based on arithmetic not geometric means should be used and suggest that resistance be redefined as occurring when the upper level of percent reduction is less than 95%. At least ten animals per group are required in most circumstances, though even 20 may be insufficient where effectiveness of the product is close to the cut off point for defining resistance.

Non-specific conducting polymer-based array capable of monitoring odour emissions from a biofiltration system in a piggery building

A commercial non-specific gas sensor array system was evaluated in terms of its capability to monitor the odour abatement performance of a biofiltration system developed for treating emissions from a commercial piggery building. The biofiltration system was a modular system comprising an inlet ducting system, humidifier and closed-bed biofilter. It also included a gravimetric moisture monitoring and water application system for precise control of moisture content of an organic woodchip medium. Principal component analysis (PCA) of the sensor array measurements indicated that the biofilter outlet air was significantly different to both inlet air of the system and post-humidifier air. Data pre-processing techniques including normalising and outlier handling were applied to improve the odour discrimination performance of the non-specific gas sensor array. To develop an odour quantification model using the sensor array responses of the non-specific sensor array, PCA regression, artificial neural network (ANN) and partial least squares (PLS) modelling techniques were applied. The correlation coefficient (r(2)) values of the PCA, ANN, and PLS models were 0.44, 0.62 and 0.79, respectively.

Soil fertility in sweetpotato-based cropping systems in the highlands of Papua New Guinea

The main outputs anticipated include enchanced knowledge of key water-nutrient dynamics in relation to key soil management techniques and a suite of improved and practical soil management options in sweet potatoes.

Laboratory-rearing Techniques for Tephritid Fruit Flies in the South Pacific

Laboratory colonies of 15 economically important species of multi-host fruit flies (Diptera:Tephritidae) have been established in eight South Pacific island countries for the purpose of undertaking biological studies, particularly host status testing and research on quarantine treatments. Laboratory rearing techniques are based on the development of artificial diets for larvae consisting predominately of the pulp of locally available fruits including pawpaw, breadfruit and banana. The pawpaw diet is the standard diet and is used in seven countries for rearing 11 species. Diet ingredients are standard proportions of fruit pulp, hydrolysed protein and a bacterial and fungal inhibitor. The diet is particularly suitable for post-harvest treatment studies when larvae of known age are required. Another major development in the laboratory rearing system is the use of pure strains of Enterobacteriaceae bacterial cultures as important adult-feeding supplements. These bacterial cultures are dissected out of the crop of wild females, isolated by sub-culturing, and identified before supply to adults on peptone yeast extract agar plates. Most species are egged using thin, plastic receptacles perforated with 1 mm oviposition holes, with fruit juice or larval diet smeared internally as an oviposition stimulant. Laboratory rearing techniques have been standardised for all of the Pacific countries. Quality control monitoring is based on acceptable ranges in per cent egg hatch, pupal weight and pupal mortality. Colonies are rejuvenated every 6 to 12 months by crossing wild males with laboratory-reared females and vice versa. The standard rearing techniques, equipment and ingredients used in collecting, establishment, maintenance and quality control of these fruit fly species are detailed in this paper.

Archaea in the foregut of macropod marsupials: PCR and amplicon sequence-based observations

Aims To investigate, using culture-independent techniques, the presence and diversity of methanogenic archaea in the foregut of kangaroos. Methods and Results DNA was extracted from forestomach contents of 42 kangaroos (three species), three sheep and three cattle. Four qualitative and quantitative PCR assays targeting the archaeal domain (16S rRNA gene) or the functional methanogenesis gene, mcrA, were used to determine the presence and population density of archaea in kangaroos and whether they were likely to be methanogens. All ruminal samples were positive for archaea, produced PCR product of expected size, contained high numbers of archaea and high numbers of cells with mcrA genes. Kangaroos were much more diverse and contradictory. Fourteen kangaroos had detectable archaea with numbers 10- to 1000-fold fewer than sheep and cattle. Many kangaroos that did not possess archaea were positive for the mcrA gene and had detectable numbers of cells with this gene and vice versa. DNA sequence analysis of kangaroos' archaeal 16S rRNA gene clones show that many methanogens were related to Methanosphaera stadmanae. Other sequences were related to non-methanogenic archaea (Thermoplasma sp.), and a number of kangaroos had mcrA gene sequences related to methane oxidising archaea (ANME). Conclusions Discrepancies between qualitative and quantitative PCR assays for archaea and the mcrA gene suggest that the archaeal communities are very diverse and it is possible that novel species exist. Significance and Impact of the Study Archaea (in general) were below detectable limits in many kangaroos, especially Red kangaroos; when present they are in lower numbers than in ruminants, and the archaea are not necessarily methanogenic. The determination of why this is the case in the kangaroo foregut could assist in reducing emissions from other ecosystems in the future.

Modelling the manager: Representing rule-based management in farming systems simulation models

We trace the evolution of the representation of management in cropping and grazing systems models, from fixed annual schedules of identical actions in single paddocks toward flexible scripts of rules. Attempts to define higher-level organizing concepts in management policies, and to analyse them to identify optimal plans, have focussed on questions relating to grazing management owing to its inherent complexity. “Rule templates” assist the re-use of complex management scripts by bundling commonly-used collections of rules with an interface through which key parameters can be input by a simulation builder. Standard issues relating to parameter estimation and uncertainty apply to management sub-models and need to be addressed. Techniques for embodying farmers' expectations and plans for the future within modelling analyses need to be further developed, especially better linking planning- and rule-based approaches to farm management and analysing the ways that managers can learn.