Loop-mediated isothermal amplification for rapid detection of Streptococcus suis in hemoculture and clinical isolates

Abstract

Introduction: Streptococcus suis is a zoonotic gram-positive cocci causing systemic infection and severe acute meningitis in humans. Infection is involved in the exposure of contaminated pigs and raw pork. Endemic usually occurs in northern area of Thailand where people eat raw pork-derived products. Due to lacking of effective vaccines, an early diagnosis of S. suis infection is extremely important for disease control and prevention. Objective: To develop a rapid and high sensitive loop-mediated isothermal amplification (LAMP) technique for investigation of S. suis. Materials and methods: Based on conserved sequences, glutamate dehydrogenase gene of S. suis was used as a target for LAMP primers design. The detection limit was compared with a conventional PCR. Thirty-four serotypes consisting of serotype 1-31, 32, 34 and 1/2 of S. suis were examined with the developed method. Specificity determination was tested with several blood-borne bacteria including other alpha and beta hemolytic Streptococcus spp. In addition, 25 clinical isolates and 30 positive hemocultures were resolved by established LAMP assay. Results: LAMP assay exhibited approximately 1,000 times more sensitive than the conventional PCR with a final bacterial load of approximately 12 colony forming unit (CFU). Thirty-four serotypes of S. suis were tested and 24 of those – serotype 2-4, 6-12, 14, 15, 17-19, 21, 24-26, 28-31 and 1/2 could be detected by LAMP technique. The assay demonstrated a high specificity without cross-reactivity to other bacteria. In addition, 25 clinical isolates and 2 out of 30 cases from positive hemocultures were successfully amplified by this method. Conclusion: LAMP assay developed is rapid and sensitive, and can be used for routine diagnosis of various serotypes of S. suis infection in clinical specimens. Bull Chiang Mai Assoc Med Sci 2016; 49(2): 207-217. Doi: 10.14456/jams.2016.23

Keywords

• Streptococcus suis
• glutamate dehydrogenase
• LAMP
• hemoculture
• PCR