Longitudinal Survey of Fecal Microbiota in Healthy Dogs Administered a Commercial Probiotic

The aim of this longitudinal microbiome study was to investigate the effects of a commercially available veterinary synbiotic product (Blackmore's® Paw DigestiCare 60™) on the fecal microbiome of healthy dogs using 16S rRNA gene microbial profiling. Fifteen healthy, privately-owned dogs participated in a 2-week trial administration of the product. Fecal samples were collected at different time points, including baseline (prior to treatment), during administration and after discontinuation of product. Large intra- and inter-individual variation was observed throughout the study, but microbiome composition at higher phylogenetic levels, alpha and beta diversity were not significantly altered after 2 weeks of probiotic administration, suggesting an absence of probiotic impact on microbial diversity. Administration of the synbiotic preparation did, however, result in transient increases in probiotic species from Enterococacceae and Streptococacceae families as well as an increase in Fusobacteria; with the fecal microbiota partially reverting to its baseline state 3-weeks after cessation of probiotic administration

Thorough assessment of DNA preservation from fossil bone and sediments excavated from a late Pleistocenee-Holocene cave deposit on Kangaroo Island, South Australia

Fossils and sediments preserved in caves are an excellent source of information for investigating impacts of past environmental changes on biodiversity. Until recently studies have relied on morphology-based palaeontological approaches, but recent advances in molecular analytical methods offer excellent potential for extracting a greater array of biological information from these sites. This study presents a thorough assessment of DNA preservation from late Pleistocene-Holocene vertebrate fossils and sediments from Kelly Hill Cave Kangaroo Island, South Australia. Using a combination of extraction techniques and sequencing technologies, ancient DNA was characterised from over 70 bones and 20 sediment samples from 15 stratigraphic layers ranging in age from >20 ka to ~6.8 ka. A combination of primers targeting marsupial and placental mammals, reptiles and two universal plant primers were used to reveal genetic biodiversity for comparison with the mainland and with the morphological fossil record for Kelly Hill Cave. We demonstrate that Kelly Hill Cave has excellent long-term DNA preservation, back to at least 20 ka. This contrasts with the majority of Australian cave sites thus far explored for ancient DNA preservation, and highlights the great promise Kangaroo Island caves hold for yielding the hitherto-elusive DNA of extinct Australian Pleistocene species

A Pilot Study to Non-Invasively Track PIK3CA Mutation in Head and Neck Cancer

Background: PIK3CA pathways are the most frequently mutated oncogenic pathway in head and neck squamous cell carcinoma (HNSCC), including virally driven HNCs. PIK3CA is involved in the PI3K-PTEN-mTOR signalling pathway. PIK3CA has been implicated in HNSCC progression and PIK3CA mutations may serve as predictive biomarkers for therapy selection. Circulating tumour DNA (ctDNA) derived from necrotic and apoptotic tumour cells are thought to harbour tumour-specific genetic alterations. As such, the detection of PIK3CA alterations detected by ctDNA holds promise as a potential biomarker in HNSCC. Methods: Blood samples from treatment naïve HNSCC patients (n = 29) were interrogated for a commonly mutated PIK3CA hotspot mutation using low cost allele-specific Plex-PCRTM technology. Results: In this pilot, cross sectional study, PIK3CA E545K mutation was detected in the plasma samples of 9/29 HNSCC patients using the Plex-PCRTM technology. Conclusion: The results of this pilot study support the notion of using allele-specific technologies for cost-effective testing of ctDNA, and further assert the potential utility of ctDNA in HNSCC

Absence of germline mutations in BAP1 in sporadic cases of malignant mesothelioma

Malignant mesothelioma (MM) is a uniformly fatal tumour caused predominantly by exposure to asbestos. It is not known why some exposed individuals get mesothelioma and others do not. There is some epidemiological evidence of host susceptibility. BAP1 gene somatic mutations and allelic loss are common in mesothelioma and recently a BAP1 cancer syndrome was described in which affected individuals and families had an increased risk of cancer of multiple types, including MM. To determine if BAP1 mutations could underlie any of the sporadic mesothelioma cases in our cohort of patients, we performed targeted deep sequencing of the BAP1 exome on the IonTorrent Proton sequencer in 115 unrelated MM cases. No exonic germline BAP1 mutations of known functional significance were observed, further supporting the notion that sporadic germline BAP1 mutations are not relevant to the genetic susceptibility of MM

Speg Interacts With Myotubularin, And Its Deficiency Causes Centronuclear Myopathy With Dilated Cardiomyopathy

Centronuclear myopathies (CNMs) are characterized by muscle weakness and increased numbers of central nuclei within myofibers. X-linked myotubular myopathy, the most common severe form of CNM, is caused by mutations in MTM1, encoding myotubularin (MTM1), a lipid phosphatase. To increase our understanding of MTM1 function, we conducted a yeast two-hybrid screen to identify MTM1-interacting proteins. Striated muscle preferentially expressed protein kinase (SPEG), the product of SPEG complex locus (SPEG), was identified as an MTM1-interacting protein, confirmed by immunoprecipitation and immunofluorescence studies. SPEG knockout has been previously associated with severe dilated cardiomyopathy in a mouse model. Using whole-exome sequencing, we identified three unrelated CNM-affected probands, including two with documented dilated cardiomyopathy, carrying homozygous or compound-heterozygous SPEG mutations. SPEG was markedly reduced or absent in two individuals whose muscle was available for immunofluorescence and immunoblot studies. Examination of muscle samples from Speg-knockout mice revealed an increased frequency of central nuclei, as seen in human subjects. SPEG localizes in a double line, flanking desmin over the Z lines, and is apparently in alignment with the terminal cisternae of the sarcoplasmic reticulum. Examination of human and murine MTM1-deficient muscles revealed similar abnormalities in staining patterns for both desmin and SPEG. Our results suggest that mutations in SPEG, encoding SPEG, cause a CNM phenotype as a result of its interaction with MTM1. SPEG is present in cardiac muscle, where it plays a critical role; therefore, individuals with SPEG mutations additionally present with dilated cardiomyopathy.WoSScopu

Complete MHC Haplotype Sequencing for Common Disease Gene Mapping

The future systematic mapping of variants that confer susceptibility to common diseases requires the construction of a fully informative polymorphism map. Ideally, every base pair of the genome would be sequenced in many individuals. Here, we report 4.75 Mb of contiguous sequence for each of two common haplotypes of the major histocompatibility complex (MHC), to which susceptibility to >100 diseases has been mapped. The autoimmune disease-associated-haplotypes HLA-A3-B7-Cw7-DR15 and HLA-A1-B8-Cw7-DR3 were sequenced in their entirety through a bacterial artificial chromosome (BAC) cloning strategy using the consanguineous cell lines PGF and COX, respectively. The two sequences were annotated to encompass all described splice variants of expressed genes. We defined the complete variation content of the two haplotypes, revealing >18,000 variations between them. Average SNP densities ranged from less than one SNP per kilobase to >60. Acquisition of complete and accurate sequence data over polymorphic regions such as the MHC from large-insert cloned DNA provides a definitive resource for the construction of informative genetic maps, and avoids the limitation of chromosome regions that are refractory to PCR amplification