Trypanosomes (genus Trypanosoma) are parasites of humans, and wild and domestic mammals, in which they cause several economically and socially important diseases, including sleeping sickness in Africa and Chagas disease in the Americas. Despite the development of numerous molecular diagnostics and increasing awareness of the importance of these neglected parasites, there is currently no universal genetic barcoding marker available for trypanosomes. In this review we provide an overview of the methods used for trypanosome detection and identification, discuss the potential application of different barcoding techniques and examine the requirements of the ‘ideal’ trypanosome genetic barcode. In addition, we explore potential alternative genetic markers for barcoding Trypanosoma species, including an analysis of phylogenetically informative nucleotide changes along the length of the 18S rRNA gene.

Thrips palmi and Frankliniella occidentalis (order Thysanoptera) are thrips species that represent major plant pests. They are polyphagous insects capable of adversely affecting crop production. As such, in the European Union, these thrips species should be regulated as quarantine organisms. T. palmi and F. occidentalis can cause considerable damage to susceptible plants by feeding on them and transmitting several viruses responsible for serious plant diseases. Successful pest control strategies are based on an early, fast, and reliable diagnosis, which precedes the selection of appropriate steps to limit the effects of harmful organisms. We herein describe a novel diagnostic approach that enables the sensitive and species-specific detection (and differentiation) of these pests in a duplex polymerase chain reaction assay, which was adapted for both standard and real-time quantitative assays. Our method is based on the amplification of a 5.8S-internal transcribed spacer 2 ribosomal DNA fragment that is conserved between T. palmi and F. occidentalis.

The close relationship between gastrointestinal (GI) microbiota and its host has an impact on the health status of an animal that reaches beyond the GI tract. A balanced microbiome stimulates the immune system, aids in the competitive exclusion of transient pathogens and provides nutritional benefits to the host. With recent rapid advances in high-throughput sequencing technology, molecular approaches have become the routinely used tools for ecological studies of the feline microbiome, and have revealed a highly diverse and complex intestinal ecosystem in the feline GI tract. The major bacterial groups are similar to those found in other mammals, with Firmicutes, Bacteroidetes, Actinobacteria and Proteobacteria constituting more than 99% of intestinal microbiota. Several nutritional studies have demonstrated that the feline microbiota can be modulated by the amount of soluble fibers (i.e., prebiotics) and macronutrients (i.e., protein content) in the diet. Initial clinical studies have suggested the presence of a dysbiosis in feline inflammatory bowel disease (IBD). Recently, metagenomic approaches have attempted to characterize the microbial gene pool. However, more studies are needed to describe the phylogenetic and functional changes in the intestinal microbiome in disease states and in response to environmental and dietary modulations. This paper reviews recent studies cataloging the microbial phylotypes in the GI tract of cats.

Increasingly, the microbiological scientific community is relying on molecular biology to define the complexity of the gut flora and to distinguish one organism from the next. This is particularly pertinent in the field of probiotics, and probiotic therapy, where identifying probiotics from the commensal flora is often warranted. Current techniques, including genetic fingerprinting, gene sequencing, oligonucleotide probes and specific primer selection, discriminate closely related bacteria with varying degrees of success. Additional molecular methods being employed to determine the constituents of complex microbiota in this area of research are community analysis, denaturing gradient gel electrophoresis (DGGE)/temperature gradient gel electrophoresis (TGGE), fluorescent in situ hybridisation (FISH) and probe grids. Certain approaches enable specific aetiological agents to be monitored, whereas others allow the effects of dietary intervention on bacterial populations to be studied. Other approaches demonstrate diversity, but may not always enable quantification of the population. At the heart of current molecular methods is sequence information gathered from culturable organisms. However, the diversity and novelty identified when applying these methods to the gut microflora demonstrates how little is known about this ecosystem. Of greater concern is the inherent bias associated with some molecular methods. As we understand more of the complexity and dynamics of this diverse microbiota we will be in a position to develop more robust molecular-based technologies to examine it. In addition to identification of the microbiota and discrimination of probiotic strains from commensal organisms, the future of molecular biology in the field of probiotics and the gut flora will, no doubt, stretch to investigations of functionality and activity of the microflora, and/or specific fractions. The quest will be to demonstrate the roles of probiotic strains in vivo and not simply their presence or absence.

Molecular methods have provided renewed impetus for the analysis of the composition of the intestinal microflora in health and disease. The polymerase chain reaction coupled with denaturing gradient gel electrophoresis provides a method whereby the bacterial communities in large numbers of samples can be compared efficiently and effectively. Altered bacterial populations associated with disease states can then be targeted for further investigation. In the long-term, an ‘abnormal microflora’ might be rectified by the use of probiotics or prebiotics.