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Shigella sonnei Antimicrobial Resistance, Phage Typing And Molecular Characterisation

Shigella sonnei Antimicrobial Resistance, Phage Typing And Molecular Characterisation. Niall DeLappe 1 , D.Morris 2 , S.Fanning 3 , M.Daly 3 , T.Chiesty 4 , F.O’Holloran 3 ,G.Corbett-Feeney 1,2 , M.Cormican 1 ,2 . 1 Interim National Salmonella Reference Laboratory, UCHG.

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Shigella sonnei Antimicrobial Resistance, Phage Typing And Molecular Characterisation

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  1. Shigella sonneiAntimicrobial Resistance, Phage Typing And Molecular Characterisation Niall DeLappe1, D.Morris2, S.Fanning3, M.Daly3, T.Chiesty4, F.O’Holloran3 ,G.Corbett-Feeney1,2 , M.Cormican1 ,2. 1Interim National Salmonella Reference Laboratory, UCHG. 2National University of Ireland, Galway 3Cork Institute of Technology. 4LEP, PHLS, Colindale

  2. Abstract

  3. Introduction Dysentery • Amoebicand Bacillary • HistoricallyA dysenteriae 15 serotypes B boydii 18 serotypes C flexneri 6 serotypes & 2 variants D sonnei 1 serotype (5 biotypes) • Evolved from different ancestral strains of E.coli. • Virulence plasmid , Pinv (140 MDa) directs the entry of bacterium into host epithelial cells. • produce various enterotoxins

  4. Shigella sonnei • Evolved from E.coli O62 • Acquired O antigen from Pleisiomonasshigelloides O17 by lateral transfer • Gene cluster for O antigen is located on Pinv. • Incidence USA 1998 25,000 cases 1999 18,000 cases UK approx. 900 cases/year • Spread by person-to-person contact and contaminated food or water • Low infectious dose (10 cells)

  5. Isolates analysed in this study • Galway n = 42 (clusters of 4,2 &2 related) • Mayo n = 9 (cluster of 3 related) • Roscommon n = 8 (cluster of 2 related) • Dublin n = 6 (unknown) • Cork n = 2 (unknown )

  6. Materials and Methods • Pulsed Field Gel Electrophoresis- XbaI • 20 h at 5-50s , 16 h at 5-20s • Phage typing (8 isolates) • Sensitivity testing • NCCLS Disk Diffusion • Plasmid profiling • alkaline lysis method of Kado & Liu • Conjugation- liquid mating • Integron analysis (14 isolates) • PCR

  7. Location of predominant strain Number of Isolates Galway34 Mayo8 Roscommon4

  8. Seasonal Variation of Predominant strain PFGE A, ASSu *(Ampicillin,Streptomycin,Sulphonamide) Results 1998 1999

  9. Pulsed Field Gel Electrophoresis B A A B1 B4 A A A A B4 B3 B1 B B5 B B B5 B B B4 Kb 339.5 291.0 242.5 194.0 145.5 97.0 48.5 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 Restriction patterns interpreted by criteria of Tenover et al

  10. PFGE analysed by Bionumerics PT9 PT9 RDNC PT6 PT50 PT6 PT6

  11. PT 9 PFGE A (n=2) PT6 PFGE B (n=1) PFGE B1 (n=1) PFGE B4 (n=1) PFGE B5 (n=1) PT50 PFGE B3 (n=1) RDNC PFGE B (n=1) Phage Typing of S. sonnei isolates (performed by LEP, Colindale, London) Phage type 6 accounts for approximately 80% of U.K. infections. PT9 is quite rare.

  12. Conjugation of S.sonnei isolates 1 Isolated on Trptone Soya Agar , Oxoid 2 Isolated on Mueller Hinton Agar, BBL A = Ampicillin 100mg/ml agar Na = Nalidixic acid 30mg/ml agar T = Tetracycline 20mg/ml agar Tm = Trimethoprim 5mg/ml agar

  13. Plasmid Profiles B4 A A A A A A A1 A1 B A A B1 B4 pDu Kb 10 8 6 5 4 3 2 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 Most isolates had multiple plasmids ranging in size from 2kb to >50kb

  14. Antibiotic Resistance Patterns Antibiotics tested: Amp10, C 30, S10, Su300, Tet30, W5, Na30, Nt300, K30, Cip5 Oxoid

  15. Conclusions • Majority of isolates tested were from 1 PFGE group, A • Good concurrence between PFGE ,phage typing and resistance typing • High rate of resistance to antibiotics with several being encoded by conjugative plasmids or integrons • It would prove interesting to analyse isolates from other countries

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