Investigation of a 24-Hour Culture Step to Determine the Viability of Streptococcus equi Subspecies equi Via Quantitative Polymerase Chain Reaction in Nasal Secretions From Horses With Suspected Strangles

Abstract

Polymerase chain reaction (PCR)-based detection assays for Streptococcus equi subspecies equi often overestimate the prevalence of samples containing viable organisms. The objective of this study was to determine if viability could be determined using genome quantitation and detection of messenger RNA (mRNA) transcripts for the SeM gene of S. equi in pre- and post-cultured samples. Nasal secretions collected from 42 horses with suspected strangles were tested by culture and by quantitative PCR (qPCR) before and 24 hours after a culture step. Viable S. equi was determined based on the detection of S. equi via culture, the detection of mRNA transcripts for the SeM gene of S. equi by qPCR, and/or an increase in absolute number of SeM target genes of S. equi between pre- and post-cultured samples. Viability was determined in 28/42 samples based on isolation of S. equi (11 samples), the presence of mRNA transcripts for the SeM gene of S. equi (25), and/or an increase in absolute quantitation of the SeM gene of S. equi between pre- and post-culture (17). The overall agreement between culture alone and the three criteria to determine viability was 59%. The overall agreement for the detection of mRNA transcripts and increase in absolute target genes was 88% and 74%, respectively. The combination of mRNA transcripts and increase in absolute target genes was able to determine the viability status in all 42 samples. In the absence of a culture-positive result for S. equi, the determination of viability can be achieved by using molecular strategies applied to samples undergoing a 24-hour culture step.

Keywords: Culture; DNA; Streptococcus equi subspecies equi; Viability; mRNA.