Abstract
Arcanobacterium (Actinomyces) pyogenes, are animal pathogens, they produce hemolytic exotoxin, pyolysin (PLO) and are opportunistic pathogens of economically important livestock such as dairy, beef cattle and sheep. Samples were collected from pleural fluid of 10 diseased and 10 healthy sheep. Samples were from neighboring village to gazelle breeding center, Riyadh Saud Arabia. The objectives of this study were to investigate A. pyogenes in these sheep. The isolates were grown on brain heart infusion (BHI; Difco) agar plates, supplemented with 5% bovine blood at 37°C and 5% CO2. Biochemical identification of 20 isolates revealed beta-hemolytic on blood agar, hydrolyzed gelatin, starch, were unable to hydrolyze esculin or reduce nitrate and did not produce urease. DNA was extracted from isolates using DNeasy Blood and Tissue Kits (Qiagen, NY, USA). Molecular tool using 16S r- deoxyribonucleic acid (DNA) universal eubacterial forward primer 16F27 (5´-AGA GTT TGA TCC TGG CTC AG-3´) and reverse primer 16SrDNA R1525 (5´-AAG GAG GTG ATC CAG CCG CA-3´), MWG Biotech AG (Ebersberg, Germany) identified the isolates successfully as A. pyogenes. Key words: Sheep, pathogen, esculin, urease, 16SrDNA, Antinomies pyogenes.
Highlights
Archanobacterium pyogenes is an anaerobic Grampositive bacilli and it is a part of the normal flora in many domestic animals (Carlos et al, 2009)
The results indicated that all isolates were non-motile, betahemolytic on blood agar, able to hydrolyze gelatin and starch, in addition, the isolates were unable to hydrolyze esculin or reduce nitrate and did not produce urease. 16S rDNA was amplified by polymerase chain reaction (PCR) successfully (Figure 3)
A number of assays such as cell culture method, enzyme-linked immunosorbent assay (ELISA), reversed passive latex agglutination (RPLA), hybridization and PCR methods are available for the detection of bacterial infection
Summary
Archanobacterium pyogenes is an anaerobic Grampositive bacilli and it is a part of the normal flora in many domestic animals (Carlos et al, 2009). In investigation of bacteria in human samples, polymerase chain reaction (PCR) based on bacterial 16S r-ribonucleic acid (RNA) genes were used by Fahriye et al (2011). A previous study had attempted to analyze the Abbreviations: PLO, Pyolysin; DNA, deoxyribonucleic acid; PCR, polymerase chain reaction; RNA, ribonucleic acid; HUS, hemolytic uremic syndrome; BHI, brain heart infusion; SEM, scanning electron microscope; TSB, trypticase soy broth medium; TEM, transmission electron microscope; ELISA, enzyme-linked immunosorbent assay. The traditional identification of bacteria on the basis of phenotypic characteristics is generally not as accurate as identification based on genotypic method of 16S rDNA sequencing is an effective means for the identification of aerobic gram-positive rods which are difficult to identify by conventional techniques. In this study primers specific for 16S rDNA in PCR reaction using template DNA from isolates were applied
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