Bovine mastitis is an inflammation of the mammary gland caused by a multitude of pathogens with devastating consequences for the dairy industry. Global annual losses are estimated to be around €30 bn and are caused by significant milk losses, poor milk quality, culling of chronically infected animals, and occasional deaths. Moreover, mastitis management routinely implies the administration of antibiotics to treat and prevent the disease which poses serious risks regarding the emergence of antibiotic resistance. Conventional diagnostic methods based on somatic cell counts (SCC) and plate-culture techniques are accurate in identifying the disease, the respective infectious agents and antibiotic resistant phenotypes. However, pressure exists to develop less lengthy approaches, capable of providing on-site information concerning the infection, and in this way, guide, and hasten the most adequate treatment. Biosensors are analytical tools that convert the presence of biological compounds into an electric signal. Benefitting from high signal-to-noise ratios and fast response times, when properly tuned, they can detect the presence of specific cells and cell markers with high sensitivity. In combination with microfluidics, they provide the means for development of automated and portable diagnostic devices. Still, while biosensors are growing at a fast pace in human diagnostics, applications for the veterinary market, and specifically, for the diagnosis of mastitis remain limited. This review highlights current approaches for mastitis diagnosis and describes the latest outcomes in biosensors and lab-on-chip devices with the potential to become real alternatives to standard practices. Focus is given to those technologies that, in a near future, will enable for an on-farm diagnosis of mastitis.
The accurate diagnosis of bacterial infections is of critical importance for effective treatment decisions. Due to the multietiologic nature of most infectious diseases, multiplex assays are essential for diagnostics. However, multiplexability in nucleic acid amplification-based methods commonly resorts to multiple primers and/or multiple reaction chambers, which increases analysis cost and complexity. Herein, a polymerase chain reaction (PCR) offer method based on a universal pair of primers and an array of specific oligonucleotide probes was developed through the analysis of the bacterial 16S ribosomal RNA gene. The detection system consisted of DNA hybridization over an array of magnetoresistive sensors in a microfabricated biochip coupled to an electronic reader. Immobilized probes interrogated single-stranded biotinylated amplicons and were obtained using asymmetric PCR. Moreover, they were magnetically labelled with streptavidin-coated superparamagnetic nanoparticles. The benchmarking of the system was demonstrated to detect five major bovine mastitis-causing pathogens: Escherichia coli, Klebsiella sp., Staphylococcus aureus, Streptococcus uberis, and Streptococcus agalactiae. All selected probes proved to specifically detect their respective amplicon without significant cross reactivity. A calibration curve was performed for S. agalactiae, which demonstrates demonstrating a limit of detection below 30 fg/µL. Thus, a sensitive and specific multiplex detection assay was established, demonstrating its potential as a bioanalytical device for point-of-care applications.
The Egyptian weasel (Mustela subpalmata) is a small mustelid with a distribution restricted to the lower Nile Valley and the Nile Delta. Traditionally considered a subspecies of the least weasel (M. nivalis), it is currently recognized as a separate species based on morphology. Here we present the first genetic assessment of the taxonomic status of the Egyptian weasel by comparing mitochondrial DNA (Cytochrome b gene and control region) sequences to those of least weasels from the western Palearctic, with a focus on the Mediterranean region. Our results provide no evidence to support the view that the Egyptian weasel is genetically distinct from the least weasel, as we found that, for both Cytochrome b and control region, haplotypes were shared between the two taxa. Specifically, the Cytochrome b and control region haplotypes detected in the Egyptian weasel were also present in M. nivalis from Turkey and Malta, two populations genetically analysed here for the first time. Our results suggest that the Egyptian weasel is distinct from the least weasel populations currently living in the Maghreb, which were inferred to be the result of an earlier colonization of North Africa, but the genetic data alone do not allow us to determine whether the Egyptian weasel is native or introduced. Nevertheless, the observed genetic patterns, together with the weasel fossil record in Israel and the unique commensal lifestyle of the Egyptian weasel, are consistent with the hypothesis that the Egyptian population is a relict of past range expansion from the Levant into Egypt. We suggest that the large size and characteristic sexual dimorphism of the Egyptian weasel are likely to represent ecotypic variation, but genomic studies are required to clarify the extent of its functional genetic divergence.
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