Molecular Identification Local Isolates of Staphylococcus aureus by Polymerase Chain Reaction (PCR) Method

Abstract

Foodborne disease is a major concern worldwide, and S. aureus is continuously involved in foodborne disease outbreaks. Staphylococcal enterotoxins (SE) are important virulence factor for the bacterial intoxications. This research aimed to identify local isolates of S. aureus based on universal 16S rRNA gene and to identify whether they possess SE A and C1 genes. Bacterial DNA isolation was conducted using Doyle and Doyle extraction method with some modification. Amplification of genes encoding for 16S rRNA, SEA and SEC1 were investigated using primers 63f/1387r, SEA-1/SEA-2 and SEC1-1/SEC1-2, consecutively. The amplification products of 16S rRNA were sequenced and then analyzed for their relatedness to S. aureus. The resulted sequence was analyzed using BLAST (Basic Local Alignment Search Tool) program and it was found that 5 out of 14 isolates were confirmed as S. aureus. The local isolates of NU4, NU5 and NU9 have 87 – 92% similarity, but all of the local isolates do not have similarity with S. aureus ATCC 25923. Based on amplification of the SE genes, 1 local isolates of S. aureus (NU5) was capable of producing both SEA and SEC1 while 1 local isolate (NU1) was only capable of producing SEA.

Bakteri S. aureus telah banyak dilaporkan sebagai patogen asal pangan yang dapat menyebabkan keracunan pada manusia. Karakteristik S. aureus antara lain termasuk bakteri bulat Gram positif yang bergerombol seperti anggur, membentuk pigmen kuning keemasan, tidak membentuk spora, non-motil, bersifat aerob dan anaerob fakultatif serta membentuk koagulase dan katalase positif. Umumnya S. aureus tidak memiliki kemampuan untuk berkompetisi baik dengan mikroflora normal pada makanan. Keberadaan S. aureus pada pangan umumnya disebabkan oleh kontaminasi silang dari pekerja maupun peralatan pengolahan yang digunakan serta perlakuan pangan setelah diolah. Bakteri ini sendiri ditemukan di dalam saluran pernapasan, permukaan kulit dan rambut hewan berdarah panas termasuk manusia. Lebih dari 30 – 50% populasi manusia adalah “carrier” S. aureus (Le Loir et al., 2003).