Seminario: Phagetyping + PFGE

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Application of phage typing and pulsed-field gel electrophoresis to analyse Salmonella enterica isolates from a suspected outbreak in Lagos, Nigeria

Kabir O. Akinyemi et al

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Seminario: Phagetyping + PFGE

  1. 1. APPLICATION OF PHAGE TYPING AND PULSED-FIELD GEL ELECTROPHORESIS TO ANALYSE SALMONELLA ENTERICA ISOLATES FROM A SUSPECTED OUTBREAK IN LAGOS, NIGERIA KABIR O. AKINYEMI ET AL . María Alejandra Ramírez Ruiz María Carolina Portela Fernández UPB - Medellín
  2. 2. INTRODUCTION
  3. 3. Salmonella
  4. 4. Salmonella <ul><li>Taxonomic classification: </li></ul><ul><ul><li>Kingdom: Bacteria </li></ul></ul><ul><ul><li>Class: Gammaproteobacteria </li></ul></ul><ul><ul><li>Order: Enterobacteriales </li></ul></ul><ul><ul><li>Family: Enterobacteriaceae </li></ul></ul><ul><ul><li>Genus: Salmonella </li></ul></ul>
  5. 5. Salmonella enterica
  6. 6. Pulsed-Field Gel Electrophoresis (PFGE) <ul><li>Used to analyze molecules  separates them according to their size & molecular weight. </li></ul><ul><li>Uses electric flow to ease migration. </li></ul><ul><li>ONE OF THE MOST USED TECHNIQUES IN MOLECULAR BIOLOGY RESEARCH </li></ul>
  7. 7. Relationship in between
  8. 8. GENERAL OBJECTIVE
  9. 9. General objective <ul><ul><li>To identify the prevalences of the different Salmonella enterica serovars using phagotyping and PFGE techniques in Lagos, Nigeria. </li></ul></ul>
  10. 10. MATERIALES Y MÉTODOS
  11. 11. Bacterias Condiciones óptimas de conservación * Las muestras eran de S. enteritidis y/ o de S. typhimurium.
  12. 12. Test de Susceptibilidad Bacteriana
  13. 13. Test de Susceptibilidad Bacteriana <ul><li>Susceptibilidad a 13 antibióticos fue evaluada: </li></ul><ul><ul><li>Ampicilina, amoxicilina, gentamicina, tetraciclina, cloramfenicol, sulfato de colecistina, trimetoprim-sulfometoxazol, trimetoprim, cefoperazona, ofloxacina, ciprofloxacina, perfloxacina y cefotaxima. </li></ul></ul><ul><li>Controles: </li></ul><ul><ul><li>Escherichia coli ATCC 25922 </li></ul></ul><ul><ul><li>Staphylococcus aureus ATCC 29213 </li></ul></ul><ul><ul><li>Enterococcus faecalis ATCC 29212 </li></ul></ul>
  14. 14. Fagotipificación <ul><li>Usa bacteriófagos </li></ul><ul><li>ID serotipos comunes </li></ul><ul><li>Mismo fagotipo Relación epidemiológica </li></ul>
  15. 15. Electroforesis en gel de campo pulsado (PFGE) Ha demostrado ser altamente discriminatoria en epidemiología
  16. 16. RESULTADOS
  17. 17. Salmonella tymphimurium
  18. 18. Salmonella tymphimurium <ul><li>PFGE: </li></ul><ul><ul><li>X1 y X2: 96.8% similitud  grupo 1. </li></ul></ul><ul><ul><li>X3: mayor prevalencia (57.9%)  grupo 2. </li></ul></ul><ul><ul><li>X4  grupo 3. </li></ul></ul><ul><li>Fagotipos </li></ul><ul><ul><li>Grupo 1: RDNC </li></ul></ul><ul><ul><li>Grupo 2: DT71 </li></ul></ul><ul><ul><li>Grupo 3: UT </li></ul></ul>Total: 19 muestras Similitud total: 71%
  19. 19. Salmonella tymphimurium RDNC: Reaccionaron pero no se conformaron DT71: Reaccionaron y se conformaron con ese fago. UT: No se tipificaron SEROTIPO UBICACIÓN FAGOTIPIFICACIÓN PFGE CANTIDAD Salmonella typhimurium Humanos RDNC X1 2 Humanos RDNC X2 2 Agua RDNC X2 1 Humanos DT71 X3 5 Ganado DT71 X3 2 Agua DT71 X3 2 Aguas sucias DT71 X3 2 Humanos UT X4 3 TOTAL: 19
  20. 20. Salmonella enteritidis
  21. 21. Salmonella enteritidis <ul><li>PFGE </li></ul><ul><ul><li>X1e: más frecuente (61.5%)  grupo 1. MULTIRRESISTENCIA </li></ul></ul><ul><ul><li>X2e y X3e: similitud 80%  grupo 2. </li></ul></ul><ul><ul><li>X4e  grupo 3. </li></ul></ul><ul><li>Fagotipos </li></ul><ul><ul><li>PT29 </li></ul></ul><ul><ul><li>UT </li></ul></ul><ul><ul><li>RDNC </li></ul></ul>Total: 13 muestras Similitud total: 64%
  22. 22. Salmonella enteritidis RDNC: Reaccionaron pero no se conformaron PT29: Reaccionaron y se conformaron con ese fago. UT: No se tipificaron SEROTIPO UBICACIÓN FAGOTIPIFICACIÓN PFGE CANTIDAD Salmonella enteritidis Humanos UT X1e 4 Aguas residuales UT X1e 1 Humanos RDNC X1e 3 Humanos RDNC X2e 1 Humanos RDNC X3e 1 Agua RDNC X4e 1 Ganado RDNC X4e 1 Agua PT29 X4e 1 TOTAL: 13
  23. 23. DISCUSSION
  24. 24. AUTHOR STATEMENT YES NO Laconcha I et al. PFGE and phagetyping increase the discrimination in the epidemiologic analysis of S. enteriditis strains. X Onyango MD et al. Phagetyping profiles cathegorized as 1, 3 & 5 may be found in S. typhimurium in Africa X Ward LR et al. Phage type PT4 is the most found within S. enteritidis samples. X Porwollik S et al. Serogroup, host specificity & disease characteristcs influence the isolates’ clustering. X
  25. 25. CONCLUSIONS
  26. 26. Conclussions <ul><li>The characterization and research about microorganisms as the Salmonella is absolutely necessary to improve the control of the diseases they cause. </li></ul><ul><li>It is alarming to see how the bacteria are getting each day stronger against antibiotics (out of 13 antibiotics just 3 showed to be effective against all serovars) and that’s mainly led by the self-medication with antibiotics, not following correct treatment schemes. </li></ul>
  27. 27. Conclussions <ul><li>The identification of different Salmonella serotypes can help to establish control on the transmission mechanisms used by these microorganisms generating a positive impact on the community’s public health. </li></ul><ul><li>Thanks to such studies we can getsome control over the use of antibiotics and try to educate people about their use and how NOT to use them, to prevent multidrug resistance. </li></ul>
  28. 28. BIBLIOGRAFÍA
  29. 29. <ul><li>Akinyemi KO, Phillip W, Beyer W, Böhm R. Application of phage typing and pulsed-field gel electrophoresis to analyse Salmonella enterica isolates from a suspected outbreak in Lagos, Nigeria. J Infect Dev Ctries 2010; 4 (12): 828-834. </li></ul><ul><li>Mancera-Martínez A, Vázquez Navarrete J, Heneidi-Zeckua A. Phage typing of Salmonella enteritidis strains isolated from poultry in Mexico. Téc Pecu Méx 2004; 42 (2): 287 – 294. </li></ul><ul><li>MARTINEZ SÁNCHEZ, Lina María [y otros]. Biología molecular. 6 ed. Medellín: UPB. Fac. de Medicina, 2011. 292 p. </li></ul><ul><li>Autor desconocido.“Pruebas de sensibilidad a agentes antimicrobianos”, disponible online en http://minnie.uab.es/~veteri/21273/Practica%202.2009-10.pdf [consultado 24/02/2011, 16:30 hrs] </li></ul><ul><li>Autor desconocido.“Prueba de susceptibilidad antimicrobiana por difusión en agar”, disponible online en http://www.ispch.cl/lab_sal/doc/man_suscep.pdf [consultado 24/02/2011, 19:00 hrs]. </li></ul>
  30. 30. MAPAS CONCEPTUALES
  31. 31. Por: María Carolina Portela Fernández
  32. 32. Por: María Alejandra Ramírez Ruiz
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