Biochemical and molecular identification of Gram-positive isolates with β-hemolysis activity isolated from the nasal swab of pigs during the human meningitis outbreak in Badung Regency, Bali-Indonesia

Abstract

Background and Aim: The nasal cavity of a pig serves as an entry point and a habitat for the colonization of commensal microbes and pathogenic bacteria. Based on biochemical and serological tests, Streptococcus β-hemolytic Group C was identified as the Gram-positive bacteria, which resulted in the 1994 outbreak and death of thousands of pigs in Bali. Furthermore, this agent is zoonotic and frequently results in the development of meningitis lesions in the infected pig. Recently, a meningitis outbreak in humans was also reported after the consumption of pig-derived foods at Sibang Kaja, Badung-Bali. This study aimed to identify and characterize Gram-positive β-hemolytic organisms collected from nasal swab of pigs from the outbreak area, as well as to compare API Kit and 16S rRNA gene analysis methods. Materials and Methods: This study commenced with the cultivation of two isolates, Punggul Swab Nasal (PSN) 2 and PSN 19, which were characterized by β-hemolysis activity. These samples were then conventionally and molecularly identified using Kit API 20 Strep and 16S ribosomal RNA (rRNA) gene primers, respectively. Results: Using the Kit API 20 Strep, both isolates were identified as Enterococcus faecium, which was previously classified as Group D Streptococci. Based on the 16S rRNA gene sequencing, PSN 2 and PSN 19 were molecularly confirmed to have 99 and 98.1% similarities with E. faecium (NR042054), respectively. Furthermore, both isolates share the same clade in the phylogenetic tree analysis. Conclusion: Using Kit API 20 Strep and 16S rRNA gene analysis, the PSN 2 and PSN 9 Gram-positive isolates with β-hemolysis activity from pig nasal swabs were identified as E. faecium.