Comparative genome analysis of Streptococcus suis strains isolated from swine in Brazil

Author(s)

Carlos Emilio Cabrera Matajira

• Document type: Doctoral Thesis
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Faculdade de Medicina Veterinária e Zootecnia (FMVZ/SBD)
• Defense date: 2020-04-08

Examining board members

Andrea Micke Moreno; Simone de Carvalho Balian; Franco Ferraro Calderaro; Terezinha Knöbl; Luís Guilherme de Oliveira

• Advisor: Andrea Micke Moreno; Luisa Zanolli Moreno

Abstract

Streptococcus suis is a pathogen of great importance in the pig industry and can cause diverse clinical conditions in animals of different ages, in addition of this, is also considered a zoonotic agent. Thus, is essential the characterization of strains that causes disease in swine from Brazil. The current study aimed phenotypic and genotypic characterization of 215 S.suis strains isolated between 2001 and 2016 from diseased swine of different Brazilian states. Isolated strains were stored at -86°C and were posteriorly reactivated for confirmation through PCR and MALDI-TOF mass spectrometry. Subsequently, molecular serotyping and virulence gene identification were performed by PCR, and genotyping was performed by AFLP and PFGE techniques. To identify the resistant profiles, the minimum inhibitory concentration (MIC) was determined using the broth microdilution technique. Based on the results obtained from the phenotypic and genotypic characterization, twenty-four strains were selected for genome sequencing using the Illumina® MiSeq platform. The most frequently identified serotype was 2 / ½ (82.8%); followed to a lesser extent by serotypes 3, 4, 6, 7, 8, 9, 18, 27, 28, 1/14, with only six non- typeable strains (NT). For virulence profile characterization, four genes - sly, arcA, epf and mrp variants were screened, and were identified 11 virulence profiles in the studied strains. The genotyping by AFLP technique identified 30 genotypic profiles and the PFGE technique identified 64 pulsotypes. For both techniques, no clear correlation between epidemiological and genotypic data could be identified, but a tendency to group strains per year of isolation can be observed. The MIC results identified nine antimicrobial susceptibility profiles and 72.1% of the strains were classified as multidrug-resistant. High rates of resistance to tetracyclines, macrolides, clindamycin and sulfadimethoxine were observed, while beta-lactams and florfenicol were the most effective antimicrobials. In the genomic analysis, no plasmid markers were detected and only three resistance genes were identified in the S. suis Brazilian genomes; the ermB gene, which encodes resistance to macrolides and lincosamides, was detected in 79.2% of the 24 studied genomes. The MLST analysis identified eight new allele sequences and one new allelic combination, such that six new type sequences (STs) were identified among the 24 genomes analyzed. It was identified a mobile genetic element (CMGETZ080501) related to antimicrobial resistance in seven from nine strains of serotype 2; the two Brazilian serotype 2 strains that differ from the others (45 and 47) were the most susceptible strains to antimicrobials. Regarding genetic mapping of the strains from serotype 3, it was observed that two of them differ from the others and show greater similarity to the reference genotype of serotype 4 strains. This result was both verified in wgSNP and MLST analysis, confirming that the capsular characterization of S. suis should not be used as an indication of genetic similarity. Given the genetic variability and the high frequency of multidrug resistance identified in the S. suis strains circulating in the country, it is possible to evaluate the reasons for the control of the agent to remain a major challenge for the national intensive pig production systems. (AU)

• FAPESP's process: 15/26159-1 - Comparative genome analysis of Streptococcus suis strains isolated from swine in Brazil
• Grantee: Carlos Emilio Cabrera Matajira
• Support Opportunities: Scholarships in Brazil - Doctorate