unique megaplasmid contributes to stress tolerance and pathogenicity of an emergent Salmonella enterica serovar Infantis strain
Of all known Salmonella enterica serovars, S. Infantis is one of the most commonly isolated and has been recently emerging worldwide. To understand the recent emergence of S. Infantis in Israel, we performed extensive comparative analyses between pre‐emergent and the clonal emergent S. Infantis popu...
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| Published in: | Environmental microbiology Vol. 16; no. 4; pp. 977 - 994 |
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| Main Authors: | , , , , , , , |
| Format: | Journal Article |
| Language: | English |
| Published: |
England
Blackwell Publishing Ltd
01-04-2014
Wiley Subscription Services, Inc |
| Subjects: | |
| Online Access: | Get full text |
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| Summary: | Of all known Salmonella enterica serovars, S. Infantis is one of the most commonly isolated and has been recently emerging worldwide. To understand the recent emergence of S. Infantis in Israel, we performed extensive comparative analyses between pre‐emergent and the clonal emergent S. Infantis populations. We demonstrate the fixation of adaptive mutations in the DNA gyrase (gyrA) and nitroreductase (nfsA) genes, conferring resistance to quinolones and nitrofurans, respectively, and the carriage of an emergent‐specific plasmid, designated pESI. This self‐transferred episome is a mosaic megaplasmid (∼280 kb), which increases bacterial tolerance to environmental mercury (mer operon) and oxidative stress, and provides further resistance to tetracycline, sulfamethoxazole and trimethoprim, most likely due to the presence of tetRA, sulI and dfrA genes respectively. Moreover, pESI carries the yersiniabactin siderophore system and two novel chaperone‐usher fimbriae. In vitro studies established that pESI conjugation into a plasmidless S. Infantis strain results in superior biofilm formation, adhesion and invasion into avian and mammalian host cells. In vivo mouse infections demonstrated higher pathogenicity and increased intestinal inflammation caused by an S. Infantis strain harboring pESI compared with the plasmidless parental strain. Our results indicate that the presence of pESI that was found only in the emergent population of S. Infantis in Israel contributes significantly to antimicrobials tolerance and pathogenicity of its carrier. It is highly likely that pESI plays a key role in the successful spread of the emergent clone that replaced the local S. Infantis community in the short time of only 2–3 years. |
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| Bibliography: | http://dx.doi.org/10.1111/1462-2920.12351 G.A.G - No. 249241 ark:/67375/WNG-CKSLLLC3-S istex:035916B853053B84EC9A20BD6D59B8365424C02B Appendix S1 Experimental procedures.Fig. S1. The emergence of S. Infantis in Israel. Salmonella is a mandatory reportable pathogen in Israel and each Salmonella isolate received at the National Salmonella Reference Center is being serotyped and documented. The prevalence of laboratory-confirmed S. Infantis isolates is shown as the percentage from all Salmonella isolates that were obtained and serotyped at the National Salmonella Reference Center 1995 to 2013. For the year 2013, S. Infantis proportion was calculated for the months of Jan-Sep. Data about S. Infantis prevalence between 1995-2009 were also included in our previous report (Gal-Mor et al., 2010) and are included here to provide a complete and updated description of S. Infantis epidemiology over 19 years.Fig. S2. A consensus pairwise comparison of the Phenotypic Microarray (PM) results that were generated from the analysis of 20 plates (1920 conditions) in duplicate runs. Each position represents the growth of isolate 119944 (red) and 335-3 (green) under a certain condition (nutrient source, antimicrobial, pH, salt concentration etc.) over 24 h (carbon and nutrient) or 48 h (sensitivity) time period. Yellow color indicates equal (and therefore overlapping) growth of both isolates.Fig. S3. The emergent strain harbors a single ∼280 kb plasmid conferring tetracycline, sulfamethoxazole and trimethoprim resistance. (A) Mating experiments between a plasmidless E. coli J5-3 strain and S. Infantis 119944 strain. Bacterial growth of the donor (S. Infantis 119944), recipient (E. coli J5-3) and two randomly selected transconjugant isolates grown on LB plates under different selections is shown. (B) S. Infantis emergent clone (119944), pre-emergent strain (335-3), E. coli transconjugant (TC-1), the recipient strain (J5-3) and E. coli R27 (carrying a 168 kb plasmid as a positive control) were digested with S1 nuclease followed by PFGE. The identified linearized plasmids are indicated by arrow heads.Fig. S4. Nalidixic acid resistance in S. Infantis is associated with point mutations at position 87 in GyrA. The amino acid sequence of the QRDR (positions 61 to 122) of the DNA gyrase subunit A (GyrA) in S. Infantis SARB27 (ZP_09726429) was compared against the sequence that was determined in pre-emergent (n = 11) and emergent (n = 13) isolates. Nalidixic acid resistance (+) or sensitive (−) phenotype is indicated. Amino acid substitutions all found at position 87 are highlighted in grey.Fig. S5. pESI harbors an IncP-1α oriV, but not an IncI1 origin of replication. (A) Southern blot analysis of the S. Infantis 119944, E. coli DH10BT1/ pCVM29188_101 (as a positive control), E. coli C600/pRK2 (as a negative control) and S. Infantis 335-3 genomes using a Dig-labeled IncI-1α oriV probe. (B) Multiple alignment of the oriV sequence present in pESI against the origin of replication of IncP-1α plasmids pBS228 (AM261760), RK2 (J01780) and pTB11 (AJ744860) is shown. Alignment was generated and formatted using Multalin version 5.4.1. High consensus is shown in red and low consensus sequences are in blue.Fig. S6. The virulence of the emergent S. Infantis clone in vivo. C57BL/6 mice were treated with streptomycin and 24 h later infected orally with the S. Infantis emergent strain (119944) and the pre-emergent strain (335-3), or given HEPES buffer only (uninfected). At 24 h post infection, colonization and bacterial burden were examined at gastrointestinal sites (cecum, colon and ileum) (A). Pathology scoring (0-25) was performed for H&E stained sections of the cecal mucosa (B). Representative micrographs at an original magnification of 40 × are shown (C). Bar = 100 μm; L indicates the lumen and E indicates submucosal edema. Two independent mice infection experiments were performed, the results of one representative experiment are shown.Table S1. Differences in antimicrobial resistance between the emergent and the pre-emergent strains.Table S2. Bacterial strains and plasmids used in this study.Table S3. Primers used in this study. GIF - No. 1096.39.11/2010 European Community's Seventh Framework program - No. PF7/2007-2013 ArticleID:EMI12351 ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 ObjectType-Article-2 ObjectType-Feature-1 |
| ISSN: | 1462-2912 1462-2920 |
| DOI: | 10.1111/1462-2920.12351 |