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OUL6602.Pdf (10.82Mo) République Algérienne Démocratique et Populaire Ministère de l’Enseignement Supérieur et de la Recherche Scientifique Université Constantine 1 Faculté des Sciences de la Nature et de la Vie Département de Microbiologie N° d’ordre : 91/DS/2014 N° de série : 05/MBIO/2014 Thèse présentée par : OULMI Lamia En vue de l’obtention du Diplôme de : DOCTORAT EN SCIENCES Option : Biochimie et Microbiologie Appliquées ÉTUDE DES INFECTIONS CAUSÉES PAR LES ACTINOMYCÈTES AÉROBIE AUTRES QUE LES MYCOBACTÉRIES DANS LA RÉGION DE CONSTANTINE Soutenue le 22/09/2014 devant le jury Présidente : Pr. MIHOUBI Ilhem Univ. Constantine 1 Directeur de thèse : Pr. BOULAHROUF Abderrahmane Univ. Constantine 1 Examinateur : Pr. AITKAKI Zahia Univ. Constantine 3 Pr. KARAM Noureddine Univ. Essania-Oran Pr. MOULAHEM Tayeb Univ. Constantine 3 Année universitaire 2013/2014 DEDICACES A mes très chers parents En hommage à tous les sacrifices que vous avez consenti pour moi. Je vous remercie d'avoir fait de moi ce que je suis. Aucune dédicace, aucun mot, ne saurait exprimer réellement mon profond amour, mon respect et ma vive gratitude. Veuillez trouver dans ce travail le fruit de toutes vos espérances et vos sacrifices. A mon très cher époux qui m’a encouragée, qui m’a donné la force et la volonté de surmonter tous les obstacles et les difficultés. Merci pour m'avoir soutenu dans les moments les plus difficiles. A mon cher poussin, Moncef A ma famille, je dédie ce mémoire Quoique je fasse, je ne pourrais vous rendre ce que vous avez fait pour moi. Si j’en suis là, c’est bien grâce à vous. Que Dieu vous donne longue vie et vous protège pour moi. Lamia REMERCIMENTS Que Monsieur Abderrahmane BOULAHROUF (Professeur au département de Microbiologie de l’Université Constantine 1) trouve ici l’expression de ma très vive reconnaissance pour avoir accepté la responsabilité de ce travail. Je garderais toujours les meilleurs souvenirs de votre immense savoir et de votre sens du devoir. Qu'il me soit permis de vous exprimer ici mon profond respect et mon intense admiration pour vos qualités humaines exceptionnelles. Je remercie Madame Ilhem MIHOUBI (Professeur à l’Université Constantine 1) de m’avoir fait l’honneur de présider ce jury. Mes remerciements vont également à Madame Zahia AITKAKI (Professeur à l’Université Constantine 3), Monsieur Noureddine KARAM (Professeur à l’Université Essania Oran) et Monsieur Tayeb MOULAHEM (Professeur à l’Université Constantine 3) d’avoir accepté de siéger dans ce jury. Je dois une profonde reconnaissance à Monsieur Mahmoud KITOUNI, qui a suivi de très près ce travail avec une grande attention et une disponibilité remarquable. C’est avec gentillesse et patience qu’il m’a toujours fait bénéficier de ses connaissances. Un grand merci d'avoir supporté mes sauts d'humeur. Toute la partie isolement a été réalisée à l’unité de tuberculose du laboratoire de bactériologie C.H.U. BENBADIS de Constantine. Je suis très reconnaissante à Madame Zahia AITKAKI qui a veillé à la réalisation de ces travaux et avec qui les discussions ont toujours été fructueuses. Je tiens à remercier le Professeur Didier Raoult pour m'avoir accueillie au sein de son Unité de Recherche sur les Maladies Infectieuses et Tropicales Emergentes, UMR CNRS. Faculté de Médecine (Marseille). Un grand merci pour Véronique Roux pour m'avoir formée et assisté à la biologie moléculaire et à la spectrométrie de masse. Merci pour la qualité de son encadrement et la pertinence de ses remarques. Je la remercie également pour sa disponibilité quotidienne et pour son degré d'implication dans ces travaux. Merci enfin pour son amitié. Merci à tous les membres du laboratoire de Bactériologie de l’Hôpital de La Timone (Marseille) et le laboratoire de bactériologie du C.H.U. BENBADIS (Consantine), que j'ai pu côtoyer. Difficile de citer tout le monde, alors merci à tous. Que mes remercîments aillent égualement à mes collègues et amies de l’Université de Constantine. Table des matières Remerciements Liste des abréviations Liste des figures Liste des tableaux Résumés Introduction ........................................................................................................ 1 Revue bibliographique 1°/ Caractéristiques générales des actinomycétales .......................................... 4 1-1 Définition et caractéristiques principales .................................................. 4 1-2 Caractères morphologiques ...................................................................... 6 1-2-1 Le mycélium du substrat .................................................................... 7 1-2-2 Le mycélium aérien ............................................................................ 8 1-2-3 Les spores .......................................................................................... 8 1-3 Ecologie .................................................................................................... 9 2°/ Pouvoir pathologique .......................... ........................................................ 10 2-1 l’infection pulmonaire .............................................................................. 12 2-2 l’infection du système nerveux central .................................................... 12 2-3 l’infection cutanée, sous-cutanée et lymphocutanée .............................. 13 2-4 L’infection extrapulmonaire (autre que le système nerveux et le tissu cutané) .......................................................................................................... 13 2-5 L’infection disséminée (impliquant deux ou plusieurs sites infectieux) ... 14 2-6 Transmission nosocomiale ..................................................................... 14 3°/ Les techniques conventionnelles d’identificatio n des actinomycètes .......... 15 3-1 Méthode morphologique ......................................................................... 15 3-2 Méthode numérique ................................................................................ 16 3-3 Méthode Chimiotaxinomique .................................................................. 16 3-3-1 Composition en acides aminés et spectre glucidique ....................... 16 3-3-2 Composition en lipides ..................................................................... 17 4°/ Le diagnostic rapide en bactériologie ......... ................................................ 20 4-1 Technologie du séquençage d’ADN ....................................................... 21 4-1-1 L’identification bactérienne par séquençage du gène ARN 16S. .... 23 4-1-2 L’identification bactérienne par séquençage des gènes domestiques (Housekeeping Gene HKG) ....................................................................... 25 4-2 La spectrométrie de masse .................................................................... 26 4-2-1 Historique ......................................................................................... 26 4-2-2 Principe ............................................................................................ 27 4-2-3 Application de la spectrométrie de masse au laboratoire de microbiologie ............................................................................................. 28 4-2-4 Conclusion ....................................................................................... 29 5°/ Classification des actinomycètes .............. .................................................. 29 Partie expérimentale 1°/ Isolement des actinomycètes à partir des prélèv ements clinique ............... 32 1-1 Les prélèvements ................................................................................... 32 1-1-1 Prélèvements contaminés ............................................................ 33 1-1-2 Prélèvements non contaminés ......................................................... 33 1-2 Traitements des prélèvements................................................................ 34 1-3 Recherche de BAAR à l’examen direct ................................................... 35 1-3-1 Préparation des frottis ...................................................................... 35 1-3-2 Coloration de Dugommier à l’auramine ............................................ 35 1-3-3 Coloration de Ziehl-Neelsen ............................................................. 35 1-4 stratégie d’isolement des actinomycètes ................................................ 36 1-4-1 Isolement des actinomycètes selon le premier protocole ................. 36 1-4-2 Isolement des actinomycètes selon le deuxième protocole .............. 37 2°/ Caractérisation morphologique des isolats actin omycétales ...................... 38 2-1 Préparation de l’inoculum ....................................................................... 38 2-2 Caractères morphologiques et culturaux ................................................ 38 2-3 Aspect des isolats en microscopie optique ............................................. 39 2-4 Aspect des isolats en microscopie électronique (Coloration négative) ... 39 2-5 Etude physiologique ............................................................................... 40 2-5-1 Production de pigments mélanoides ................................................ 40 2-5-2 Tolérance au chlorure de sodium ..................................................... 40 2-5-3 Croissance à différentes températures ............................................. 40 2-5-4 Croissance à différentes concentration de CO2................................ 40 2-5-5 Sensibilité au lysozyme ...................................................................
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