Simulated Clinical Skills for Veterinary Students Supplement Limited Animal and Clinical Resources in Developing Countries.

As part of an OIE Veterinary Education Twinning Project linking The University of Queensland, Australia and Nong Lam University, Vietnam, the limited access to animal and clinical resources was identified as an impediment to high quality veterinary education at Nong Lam University. However, student focused, simulated learning spaces, which have been widely adopted in veterinary training, are a cost-effective opportunity to provide initial clinical skills to students in countries where resourcing is constrained. In clinical skills training facilities, students use models and simulators to practice their clinical skills to develop the confidence, competence and muscle memory to enter the clinical phase of their training. While high-fidelity veterinary simulators and models are expensive, effective models for foundational clinical skills development can be built in-house for students to practice their skills authentically. This article outlines the cost effective establishment of a veterinary clinical skills training facility at Nong Lam University.

Identification of bacteria and fungi sampled from the conjunctival surface of normal horses in South-East Queensland, Australia.

OBJECTIVE: To identify bacteria and fungi found on the conjunctival surface of normal horse eyes; to investigate potential risk factors for these microflora; and to determine their susceptibility to common topical ophthalmic antimicrobials. ANIMALS STUDIED: A total of 95 client-owned horses were studied. PROCEDURES: Horses within sub-tropical Australia (South-East Queensland) were sampled once between April 2012 and March 2013. A conjunctival swab was taken from each eye and cultured for aerobic bacteria and fungi. Organisms were identified by colony morphology and phenotype. Antimicrobial disk diffusion susceptibility testing for commonly used antimicrobials was performed. RESULTS: Positive bacterial cultures were returned from 187/190 (98.4%) eyes from 94/95 (98.9%) horses. The most common species included Staphylococcus spp. (25.2% of total bacterial isolates), Bacillus cereus (17.4%), Bacillus spp. (14.1%), and Corynebacterium spp. (8.9%). Most bacterial isolates were susceptible to neomycin and fluoroquinolones. Positive fungal cultures were returned from 111/190 (58.4%) eyes from 73 (76.8%) horses. The most common species identified included: Penicillium spp. (16.7% of fungal isolates), Aspergillus spp. (15.4%), and Scopulariopsis spp. (10.3%). Most (≥90%) molds were susceptible to ketoconazole, voriconazole, itraconazole, and miconazole. Yeasts were most susceptible to ketoconazole. There was no significant effect of breed, age, sex, purpose, or housing of the horse or climatic conditions on bacterial or fungal culture status. CONCLUSIONS: Bacteria and fungi were commonly isolated from the eyes of healthy horses. The antibiotic and antifungal susceptibilities identified can be used as a guide for empirical therapy after cytology in the treatment of corneal ulceration in horses.

The polymerase chain reaction (PCR): general methods.

The polymerase chain reaction (PCR) converts very low quantities of DNA into very high quantities and is the foundation of many specialized techniques of molecular biology. PCR utilizes components of the cellular machinery of mitotic cell division in vitro which respond predictably to user inputs. This chapter introduces the principles of PCR and discusses practical considerations from target sequence definition through to optimization and application.

Genome walking.

Genome walking is a method for determining the DNA sequence of unknown genomic regions flanking a region of known DNA sequence. The Genome walking has the potential to capture 6-7 kb of sequence in a single round. Ideal for identifying gene promoter regions where only the coding region. Genome walking also has significant utility for capturing homologous genes in new species when there are areas in the target gene with strong sequence conservation to the characterized species. The increasing use of next-generation sequencing technologies will see the principles of genome walking adapted to in silico methods. However, for smaller projects, PCR-based genome walking will remain an efficient method of characterizing unknown flanking sequence.

High-throughput sequencing and mutagenesis to accelerate the domestication of Microlaena stipoides as a new food crop.

Global food demand, climatic variability and reduced land availability are driving the need for domestication of new crop species. The accelerated domestication of a rice-like Australian dryland polyploid grass, Microlaena stipoides (Poaceae), was targeted using chemical mutagenesis in conjunction with high throughput sequencing of genes for key domestication traits. While M. stipoides has previously been identified as having potential as a new grain crop for human consumption, only a limited understanding of its genetic diversity and breeding system was available to aid the domestication process. Next generation sequencing of deeply-pooled target amplicons estimated allelic diversity of a selected base population at 14.3 SNP/Mb and identified novel, putatively mutation-induced polymorphisms at about 2.4 mutations/Mb. A 97% lethal dose (LD97) of ethyl methanesulfonate treatment was applied without inducing sterility in this polyploid species. Forward and reverse genetic screens identified beneficial alleles for the domestication trait, seed-shattering. Unique phenotypes observed in the M2 population suggest the potential for rapid accumulation of beneficial traits without recourse to a traditional cross-breeding strategy. This approach may be applicable to other wild species, unlocking their potential as new food, fibre and fuel crops.

Analysis of adaptive ribosomal gene diversity in wild plant populations from contrasting climatic environments.

Plant populations may contain variation that reflects adaptation to local environmental conditions. Clues to adaptive evolution of plants may be found in the genomes of species growing in diverse environments or across steep environmental gradients, and under stress. We have examined populations of wild relatives of barley and rice across diverse environmental gradients. Greater diversity, in a nuclear biotic stress defense gene and in chloroplast genes, was found in the more stressed, hotter and dryer environments. This may reflect the greater heterogeneity of these environments. Adaptation of plants to different abiotic stresses (temperatures and levels of water availability) may also require significant adaptation to the different biotic (pest and disease) pressures in these environments.

Genome diversity in wild grasses under environmental stress.

Patterns of diversity distribution in the Isa defense locus in wild-barley populations suggest adaptive selection at this locus. The extent to which environmental selection may act at additional nuclear-encoded defense loci and within the whole chloroplast genome has now been examined by analyses in two grass species. Analysis of genetic diversity in wild barley (Hordeum spontaneum) defense genes revealed much greater variation in biotic stress-related genes than abiotic stress-related genes. Genetic diversity at the Isa defense locus in wild populations of weeping ricegrass [Microlaena stipoides (Labill.) R. Br.], a very distant wild-rice relative, was more diverse in samples from relatively hotter and drier environments, a phenomenon that reflects observations in wild barley populations. Whole-chloroplast genome sequences of bulked weeping ricegrass individuals sourced from contrasting environments showed higher levels of diversity in the drier environment in both coding and noncoding portions of the genome. Increased genetic diversity may be important in allowing plant populations to adapt to greater environmental variation in warmer and drier climatic conditions.

Characterizing homologues of crop domestication genes in poorly described wild relatives by high-throughput sequencing of whole genomes.

Wild crop relatives represent a source of novel alleles for crop genetic improvement. Screening biodiversity for useful or diverse gene homologues has often been based upon the amplification of targeted genes using available sequence information to design primers that amplify the target gene region across species. The crucial requirement of this approach is the presence of sequences with sufficient conservation across species to allow for the design of universal primers. This approach is often not successful with diverse organisms or highly variable genes. Massively parallel sequencing (MPS) can quickly produce large amounts of sequence data and provides a viable option for characterizing homologues of known genes in poorly described genomes. MPS of genomic DNA was used to obtain species-specific sequence information for 18 rice genes related to domestication characteristics in a wild relative of rice, Microlaena stipoides. Species-specific primers were available for 16 genes compared with 12 genes using the universal primer method. The use of species-specific primers had the potential to cover 92% of the sequence of these genes, while traditional universal primers could only be designed to cover 80%. A total of 24 species-specific primer pairs were used to amplify gene homologues, and 11 primer pairs were successful in capturing six gene homologues. The 23 million, 36-base pair (bp) paired end reads, equated to an average of 2X genome coverage, facilitated the successful amplification and sequencing of six target gene homologues, illustrating an important approach to the discovery of useful genes in wild crop relatives.

Domestication to crop improvement: genetic resources for Sorghum and Saccharum (Andropogoneae).

BACKGROUND: Both sorghum (Sorghum bicolor) and sugarcane (Saccharum officinarum) are members of the Andropogoneae tribe in the Poaceae and are each other's closest relatives amongst cultivated plants. Both are relatively recent domesticates and comparatively little of the genetic potential of these taxa and their wild relatives has been captured by breeding programmes to date. This review assesses the genetic gains made by plant breeders since domestication and the progress in the characterization of genetic resources and their utilization in crop improvement for these two related species. GENETIC RESOURCES: The genome of sorghum has recently been sequenced providing a great boost to our knowledge of the evolution of grass genomes and the wealth of diversity within S. bicolor taxa. Molecular analysis of the Sorghum genus has identified close relatives of S. bicolor with novel traits, endosperm structure and composition that may be used to expand the cultivated gene pool. Mutant populations (including TILLING populations) provide a useful addition to genetic resources for this species. Sugarcane is a complex polyploid with a large and variable number of copies of each gene. The wild relatives of sugarcane represent a reservoir of genetic diversity for use in sugarcane improvement. Techniques for quantitative molecular analysis of gene or allele copy number in this genetically complex crop have been developed. SNP discovery and mapping in sugarcane has been advanced by the development of high-throughput techniques for ecoTILLING in sugarcane. Genetic linkage maps of the sugarcane genome are being improved for use in breeding selection. The improvement of both sorghum and sugarcane will be accelerated by the incorporation of more diverse germplasm into the domesticated gene pools using molecular tools and the improved knowledge of these genomes.