Biology subjects

Walter, M. C.

Publications and source records attributed to Walter, M. C..

3 recordsLinked to original sources

bioRxiv2021

A scalable, clinically severe pig model for Duchenne muscular dystrophy

Large animal models for Duchenne muscular dystrophy (DMD) are crucial for preclinical evaluation of novel diagnostic procedures and treatment strategies. Pigs cloned from male cells lacking DMD exon 52 (DMD{Delta}52) resemble molecular, clinical and pathological hallmarks of DMD, but cannot be propagated by breeding due to death before sexual maturity. Therefore, female DMD+/- carriers were generated. A single founder animal had 11 litters with 29 DMDY/-, 34 DMD+/- as well as 36 male and 29 female wild-type (WT) offspring. Breeding with F1 and F2 DMD+/- carriers resulted in additional 114 DMDY/- piglets. The majority of them survived for 3-4 months, providing large cohorts for experimental studies. Pathological investigations and proteome studies of skeletal muscles and myocardium confirmed the resemblance of human disease mechanisms. Importantly, DMDY/- pigs reveal progressive fibrosis of myocardium and increased expression of connexin-43, associated with significantly reduced left ventricular fractional shortening and ejection fraction already at age 3 months. Furthermore, behavioral tests provided evidence for impaired cognitive ability of DMDY/- pigs. Our breeding cohort of DMD{Delta}52 pigs and standardized tissue repositories from DMDY/- pigs, DMD+/- carriers, and WT littermate controls provide important resources for studying DMD disease mechanisms and for testing novel diagnostic procedures and treatment strategies.

pathology↗

bioRxiv2021

In-depth analysis of Bacillus anthracis 16S rRNA genes and transcripts reveals intra- and intergenomic diversity and facilitates anthrax detection

Analysis of 16S ribosomal RNA (rRNA) genes provides a central means of taxonomic classification of bacterial species. Based on presumed sequence identity among species of the Bacillus cereus sensu lato group, the 16S rRNA genes of B. anthracis have been considered unsuitable for diagnosis of the anthrax pathogen. With the recent identification of a single nucleotide polymorphism in some 16S rRNA gene copies, specific identification of B. anthracis becomes feasible. Here, we designed and evaluated a set of in situ-, in vitro- and in silico-assays to assess the yet unknown 16S-state of B. anthracis from different perspectives. Using a combination of digital PCR, fluorescence in situ hybridization, long-read genome sequencing and bioinformatics we were able to detect and quantify a unique 16S rRNA gene allele of B. anthracis (16S-BA-allele). This allele was found in all available B.anthracis genomes and may facilitate differentiation of the pathogen from any close relative. Bioinformatics analysis of 959 B. anthracis genome data-sets inferred that abundances and genomic arrangements of the 16S-BA-allele and the entire rRNA operon copy-numbers differ considerably between strains. Expression ratios of 16S-BA-alleles were proportional to the respective genomic allele copy-numbers. The findings and experimental tools presented here provide detailed insights into the intra- and intergenomic diversity of 16S rRNA genes and may pave the way for improved identification of B. anthracis and other pathogens with diverse rRNA operons.

microbiology↗

bioRxiv2021

Catching SARS-CoV-2 by sequence hybridization: a comparative analysis

Controlling and monitoring the still ongoing SARS-CoV-2 pandemic regarding geographical distributions, evolution and emergence of new mutations of the SARS-CoV-2 virus is only possible due to continuous next-generation sequencing (NGS) and worldwide sequence data sharing. Efficient sequencing strategies enabling the retrieval of the maximum number of high quality, full-length genomes are hence indispensable. Here, we describe for the first time a combined approach of digital droplet PCR (ddPCR) and NGS to evaluate five commercially available sequence capture panels targeting SARS-CoV-2. In doing so, we were not only able to determine the most sensitive and specific capture panel, but to discriminate their mode of action and number of read pairs needed to recover a high quality full length genome. Thereby, we are providing essential information for all sequencing laboratories worldwide striving for maximizing the sequencing output and simultaneously minimizing time, costs and sequencing resources.

genomics↗