Streptococcus suis outbreak caused by an emerging zoonotic strain with acquired multi-drug resistance in Thailand

Abstract

Streptococcus suis is an emerging zoonotic swine pathogen which can cause severe infections in humans. In March 2021, an outbreak of S. suis infections with 19 confirmed cases of septicemia and meningitis leading to two deaths, occurred in Nakhon Ratchasima province, Thailand. We characterized the outbreak through an epidemiological investigation combined with Illumina and Nanopore whole genome sequencing (WGS). The source of the outbreak was traced back to a raw pork dish prepared from a single pig during a Buddhist ceremony attended by 241 people. WGS analysis revealed that a single S. suis serotype 2 strain belonging to a novel sequence type (ST) of the emergent Thai zoonotic clade CC233/379, was responsible for the infections. The outbreak clone grouped together with other Thai zoonotic strains from CC233/379 and CC104 in a global S. suis phylogeny and capsule switching events between serotype 2 zoonotic strains and serotype 7 porcine strains were identified. The outbreak strain showed reduced susceptibility to penicillin corresponding with mutations in key residues in the penicillin binding proteins (PBPs). Furthermore, the outbreak strain was resistant to tetracycline, erythromycin, clindamycin, linezolid and chloramphenicol, having acquired an integrative and conjugative element (ICE) carrying resistance genes tetO and ermB, as well as a transposon from the IS1216 family carrying optrA and ermA. This investigation demonstrates that multi-drug resistant zoonotic lineages of S. suis which pose a threat to human health continue to emerge.

Keywords: Streptococcus suis; antimicrobial resistance; outbreak; whole genome sequencing; zoonosis.

Fig. 1.

Overview of the S. suis Nakhon Ratchasima outbreak. The Nakhon Ratchasima province and the Dan Khun Thod district where the outbreak took place are highlighted in red. In total, 199 participants of the Buddhist ordination ceremony were examined at community hospital following local news of the two deaths caused by S. suis . Patient information was assessed by a team of epidemiologists from the Ministry of Public Health. The patients were also interviewed and questionnaires were conducted by the epidemiologists. From this, 88 participants were suspected cases of the outbreak and hemocultures were taken from them. Lumbar punctures were performed on suspected cases showing symptoms of meningitis (n=3) to collect cerebrospinal fluid (CSF). Nineteen cases were confirmed with positive cultures (hemocultures and CSF). Presumptive taxonomical identification of S. suis was carried out using Vitek-2 in the Maharaj Nakhon Ratchasima hospital as well as via a previously described PCR typing method [27, 28].

Fig. 2.

SNP reference mapped phylogeny of outbreak and post-outbreak S. suis isolates. The maximum-likelihood phylogeny was reconstructed using Snippy and IQ-TREE by mapping the filtered Illumina reads of each isolate to the complete genome of the index case (STC78). Branch lengths were corrected for recombination using ClonalFrameML. Outbreak isolates are marked with a black circle. Information regarding the ST and serotype of each isolate is included in the columns adjacent to the phylogenetic tree. The two lineages CC104 and CC233/379 are coloured blue and red respectively in the tree.

Fig. 3.

Emergence of outbreak clone within the context of the global S. suis population. (a) Core-genome maximum-likelihood phylogeny based on 1725 S . suis genomes reconstructed using Roary and IQ-TREE. Inner ring represents the host from which strains were isolated. The outer ring indicates whether strains were collected during this study or were part of a previously curated S. suis dataset [46]. The main S. suis CCs are annotated with various colours. The magnifying lens indicates the clade to which the outbreak strain belongs. (b) Whole genome SNP reference mapping based phylogeny of the outbreak and related clades reconstructed using Snippy and IQ-TREE. Branch lengths were corrected for recombination using ClonalFrameML. Support values for each node of the genome-wide SNP based tree used as an input for ClonalFrameML are shown in Fig. S5. The seven additional CC104 isolates were isolated from human (n=2) and pig (n=5). Human isolates were isolated in Thailand whereas the porcine isolates were isolated in the USA (n=3), China (n=1), and Canada (n=1) [33, 76]. Columns adjacent to the tree contain information regarding the host, source, ST and serotype for each isolate. The two different CCs are coloured in blue (CC104) and red (CC233/379) in the tree. The yellow circle indicates the acquisition of ICENsui78te. The blue star indicates the acquisition of the IS1216-optra-ermA. The red triangle indicates a capsule switching event. The green square indicates progressive acquisition of mutations in the PBPs.

Fig. 4.

MGEs containing AMR genes acquired by the outbreak clade. (a) Protein level alignment of the ICENsui78te with the ICENsui34te carrying tetO and ermB previously found in strains isolated in Thailand [62]. The ICENsui34te annotated sequence was extracted from strain DP_SS29 2 (NZ_WCIZ01000002.1 [63]). (b) Protein level alignment of the IS1216 Transposon with the φSC81 prophage (MK359990.1 [66]) and the STC78 genomic region where it was inserted. Genes of interest are coloured and defined in the legend. Alignments were made using clinker and clustermap.js.