Abstract
Short Sequence Repeat (SSR) typing of Mycobacterium avium subspecies paratuberculosis (Map) isolates is one of the most commonly used method for genotyping this pathogen. Currently used techniques have challenges in analyzing mononucleotide repeats >15 bp, which include some of the Map SSRs. Fragment analysis is a relatively simple technique, which can accurately measure the size of DNA fragments and can be used to calculate the repeat length of the target SSR loci. In the present study, fragment analysis was used to analyze 4 Map SSR loci known to provide sufficient discriminatory power to determine the relationship between Map isolates. Eighty-five Map isolates from 18 animals from the island of Newfoundland were successfully genotyped using fragment analysis. To the best of our knowledge, this is the first report on Map SSR diversity from Newfoundland dairy farms. Previously unreported Map SSR-types or combinations were also identified during the course of the described work. In addition, multiple Map SSR-types were isolated from a single animal in many cases, which is not a common finding.
Highlights
Mycobacterium avium subspecies paratuberculosis (Map) is a slow growing bacterium and is the cause of Johne’s disease, which is associated with chronic debilitating granulomatous enteritis that affects the small intestine of cattle, sheep, goats, farmed deer and {Li, 2005 #1017}other ruminants [1,2,3,4,5,6]
The sequencing of DNA repeats using conventional methods is often challenging and is prone to artifacts, which is further exacerbated by repeats with high GC content such as those found in Map
In the case of Map, it has been previously reported that the analysis of 4 Short Sequence Repeat (SSR) (L1/L2: monucleotide, and L3/L4 trinucleotide) provides enough sequence information for strain discrimination, and that one of the mononucleotide SSRs (L1) can be >14 bp in length depending on the isolate [5,14]
Summary
Mycobacterium avium subspecies paratuberculosis (Map) is a slow growing bacterium and is the cause of Johne’s disease, which is associated with chronic debilitating granulomatous enteritis that affects the small intestine of cattle, sheep, goats, farmed deer and {Li, 2005 #1017}other ruminants [1,2,3,4,5,6]. In the current study we used fragment analysis to analyze Map isolates from five Newfoundland dairy farms, the results of which are described below. The DNA sequences of the four SSR repeats were determined for a handful of Map isolates obtained as part of a separate study as described previously [5,14].
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