Characterization of Spectinomycin Resistance in Streptococcus suis Leads to Two Novel Insights into Drug Resistance Formation and Dissemination Mechanism

Abstract

Spectinomycin is an aminocyclitol antibiotic used clinically to treat a variety of infections in animals. Here, we characterized drug resistance prevalence in clinical Streptococcus suis isolates and discovered a novel resistance mechanism in which the s5 mutation (Gly26Asp) results in high spectinomycin resistance. Additionally, a novel integrative and conjugative element encompassing a multidrug resistance spw_like-aadE-lnu(B)-lsa(E) cluster and a cadmium resistance operon were identified, suggesting a possible cause for the wide dissemination of spectinomycin resistance in S. suis. Streptococcus suis, the leading agent for human meningitis in several Asian countries, is a significant zoonotic pathogen worldwide. Recently, it has received growing attention from the global academic community not only for its increased incidence of infections in humans but also for its important implied role in drug resistance transmission (1). The aminocyclitol antibiotic spectinomycin, often combined with lincomycin, is widely used for the treatment of pathogen infections in farm animals, including swine (2). However, few studies have examined spectinomycin resistance in S. suis globally. In this study, 191 clinical S. suis isolates, collected from different provinces in China over the period from 2006 to 2012, were initially subjected to spectinomycin susceptibility analysis by Etest method. Based on the spectinomycin MIC breakpoints to cattle respiratory pathogens (3), 20 isolates (10.4%) were found to be resistant (MIC of Ն256 g/ml) (see Table S1 in the supplemental material). To investigate resistance mechanisms within them, multi-PCR experiments were conducted to detect an array of adenyltransferase genes, including spc, aad9, spw, spd, and various aadA genes (4-8). In addition, the complete 16S RNA gene and the s5 gene were amplified and sequenced for mutation analysis in all resistant strains as well as certain susceptible strains (see Table S3 in the supplemental material). Unexpectedly, none of the widely reported resistance genes could be detected among these strains with the exception of the spw gene and its variant spw_like, which demonstrated approximately 94% amino acid identity corresponding to 15 site mutations (see Fig. S1 in the supplemental material). The spw gene and its variant spw_like were present in 13 isolates and 6 isolates, respectively (Table 1). In contrast, there were a variety of mutations within the 16S RNA and s5 genes in the resistant isolates in comparison with the susceptible strains, and, among these, two mutations were present in the 16S rRNA regions of all four operons corresponding to sites 1069(C1069T) and 1188(A1188G) in Escherichia coli 16S rRNA and one site within the 26 amino acids (aa) of the s5 protein (GGT ¡ GAT, Gly26Asp). Interestingly, in the isolate with the s5 gene mutation, the spw or spw_like genes were also not detected, suggesting that spectinomycin resistance was likely caused by this mutation.To verify these potential mechanisms, we cloned the complete spw_like genes a...

Chromosomal Conjugative and Mobilizable Elements in Streptococcus suis: Major Actors in the Spreading of Antimicrobial Resistance and Bacteriocin Synthesis Genes

“…In particular, GI integrated into rpsI could be mobilizable by subverting the relaxase and mating apparatus of a co-resident ICE. Indeed, the transfer of such element-mis-annotated as an ICE even if it does not carry any conjugation gene-was described recently (at a very low frequency: 3.7×10 −9 per donor) [35]. It should be pointed out that such elements are more difficult to detect in the genomes and are likely overlooked.…”

Genomic Epidemiology of Streptococcus suis Sequence Type 7 Sporadic Infections in the Guangxi Zhuang Autonomous Region of China

“…Viridans Group and Streptococcus pneumoniae. No breakpoint values for streptomycin, kanamycin, gentamicin, and spectinomycin were available for Streptococci, and their breakpoints were taken from previous studies [38,39].…”

A novel genomic island harbouring lsa(E) and lnu(B) genes and a defective prophage in a Streptococcus pyogenes isolate resistant to lincosamide, streptogramin A and pleuromutilin antibiotics

“…A second lsa(E)lnu(B)-containing element was described in a human S. agalactiae isolate ( Fig 1C; 20162235) [7]. Beside this, the lnu(B) gene have been also found in streptococci and staphylococci with L and LS A P phenotypes isolated from animals and some food products [18], indicating that some streptococci serve as a reservoir for antimicrobial resistance genes [17]. When comparing the characterized structures harbouring lsa(E)lnu (B) genes with GAS2887HUB (Fig.…”

“…Among streptococci (Fig 1C), the MGE harbouring lsa(E)lnu(B) genes was first reported in a S. agalactiae structure (SGB76) also containing a multidrug resistance cluster (aadEaptspw) [8]. This combination was also described with different arrangement in S.suis [17,18]. A second lsa(E)lnu(B)-containing element was described in a human S. agalactiae isolate ( Fig 1C; 20162235) [7].…”

“…When comparing the characterized structures harbouring lsa(E)lnu (B) genes with GAS2887HUB (Fig. 1C), three findings were notable [8,16,18,19]. First, the orf4orf5lsa(E)lnu(B) structure was highly conserved (95% identity) among them.…”