Genetic Characterization of Corynebacterium pseudotuberculosis Isolates in Egypt

Nadine El-Sebay,Dalia Abd El-Moaty,Elham El-Sergany,Ashraf M Abbas,Roukaya M Osman,Marwah Abd Elmonem

doi:10.21608/javs.2021.154576

Nadine El-Sebay, Dalia Abd El-Moaty + Show 4 more

Open Access

https://doi.org/10.21608/javs.2021.154576

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Abstract

Corynebacterium pseudotuberculosis is a small Gram-positive bacillus containing mycolic acid in the structure of the cell wall. The bacterium is responsible for Caseous Lymphadenitis (CLA) in small ruminants (sheep and goats). The bacteria are also responsible for Ulcerative Lymphangitis in equines. The disease causes great economic loss in the animal industry. This work aimed to check the ability of Quadruplex PCR (Q-PCR) for genotyping and identification of Egyptian isolates of C. pseudotuberculosis and sequence analysis of phospholipase D (PLD) gene of local isolates. Four of C. pseudotuberculosis local isolates previously biochemically identified were tested for narG gene (nitrate reductase gene). Both nitrate negative biovar (ovis) and nitrate positive biovar (equi) showed a positive result for 16S rRNA, rpoB and PLD genes of C. pseudotuberculosis species. The sequence analysis of our local isolates' PLD gene revealed minor changes in PLD proteins between ovine and equine strains compared with other published PLD sequences in GenBank. It was concluded that the Q-PCR method is able to differentiate between C. pseudotuberculosis equi and ovis biovars. Also, the sequence of PLD gene of local isolates representing the two biovars revealed some variation, which leads to an accurate diagnosis of C. pseudotuberculosis biovars and generates a mapping of immerged local isolates. Further, the PCR and sequence of these isolates provide rapid and accurate genotyping, especially with hyperimmune serum's unavailability.

Highlights

Corynebacterium pseudotuberculosis is a Gram-positive and small bacillus
When tested with DNA from bacterial isolates taxonomically related to C. pseudotuberculosis biovar ovis from goat and sheep and C. pseudotuberculosis biovar equi from buffalo and cattle gave a similar profile of Multiplex PCR, with the 816,446 and 203 bp amplicons corresponding to the 16S rRNA, rpoB and phospholipase D (PLD) genes, respectively Fig. (1)
The multiplex PCR assay that targets 16S rRNA, rpoB, and PLD genes was improved by adding C. pseudotuberculosis biovar-specific primers for the narG gene (Table 2) which was only found in C. pseudotuberculosis isolates biovar equi from cattle and buffalo (Fig. 2)

Summary

Introduction

Corynebacterium pseudotuberculosis is a Gram-positive and small bacillus It is the causative agent of Caseous Lymphadenitis (CLA) in goats and sheep, while in buffaloes and cattle, it causes a disease known as Oedematose Skin Disease (OSD), but in equine it causes abscesses and ulcerative lymphangitis. It was recorded in other hosts, including camels and rare in humans (Tejedor et al, 2008). Biovar equi strains are nitrate positive and infect both horses and buffalos, while biovar ovis are known to be nitrate negative and it is the causative agent for CLA in goats and sheep (Biberstein et al, 1971; Selim., 2001; Baird and Fontaine,2007).

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