التشخيص المباشر للبكتريا المسببة لالتهاب الأذن الوسطى باستخدام تفاعل البلمرة المتسلسل في الوقت الحقيقي بالمقارنة مع طرق أخرى

الباحث

آمنة سامي احمد شهاب

المشرف

جاسم فتحي علي

الجامعة

جامعة الموصل

الكلية / القسم

كلية التربية للعلوم الصرفة / قسم علوم الحياة

الشهادة

ماجستير

السنة

2024

لغة الرسالة

انكليزي

الكلمات المفتاحية

Otitis media ; Real-time PCR ; Bacterial identification ; Diagnostic methods

140 swabs were collected from people with middle ear infections arriving at the ENT consultation at: Al-Salam Teaching Hospital, Al- Jumhuriyya Teaching Hospital, and Ibn Al-Atheer Teaching Hospital, whose ages ranged from 5 months - 80 years.Duration time 1October, 2023 to 1March. 2024 100 samples were used for traditional culture and biochemical tests, and 40 swabs were used for direct DNA extraction for direct molecular diagnosis. The study showed the predominance of gram-negative bacteria at a rate of 53.45% over gram-positive bacteria at a rate of 46.55%. As for molecular diagnostic smears, they showed a predominance of gram- positive bacteria at a rate of 34% On Gram-negative bacteria at a rat 24.5%. The results of the study showed that there are some factors that influence and help in its occurrence and spread Infection.the rate of infection among children was higher than that of adults. The geographical region also has an impact on the incidence of infection, so the percentage of infected people in rural areas was higher by 51%. of urbanized areas by 49%. Smoking also has a significant impact on causing infection. The results showed that the infection rate among smokers reached 17%, and the rate increased among people exposed to passive smoking by 22%. The males were more affected than females by 55.29%, and the highest incidence was among the age group 16-45 years. The most effective antibiotic recorded in the study was Imipenem with a rate of 86.84%, and the least effective antibiotic recorded in the study was Cieftidizim with a rate of 40.78% The results of the study showed that there is a difference between the results of bacteria diagnosed by culture methods and biochemical tests,and the bacteria diagnosed by direct molecular diagnosis. The results showed Culture methods: The bacteria causing otitis media is Pseudomonase aeruginosa at a rate of 32.29%, Staphlococcus aureus 11.76%, Staphlococcus epidermidis at a rate of 10.58%, Proteus mirabilis 9.41%, klebsiella pneumonia 7.05%, 4.70%, while the results of direct molecular diagnosis without culture showed that the bacteria causing otitis media are Streptococcus pneumonae 87.5%, Staphlococcus aureus 82.5%, Pseudomonase aeruginosa 80%, Haemophilus influenza 22.5%, Moraxella catarrhalis 22%, which both are did not appear at all in the culture diagnosis. there is a difference in the results of the molecular diagnosis itself. The results of the diagnosis using qPCR showed the presence of most of the bacteria that cause otitis media, in numbers very close to the numbers known in other studies, while the results of the PCR showed an almost absence of pathogens from the isolated samples despite the presence of DNA of pathogenic bacteria. The reason for this is that the qPCR technology can detect very small concentrations of nucleic acids, unlike the PCR technology, which requires high concentrations of nucleic acids and cannot detect low concentrations. Content List NO. Subject Pag Content list Tables list Figures& Pictures List Appendix list Abbreviations & Symbols List Chapter One/ Introduction Introduction The study aimed to: Chapter Two/ Literature review Literature review Definition of Otitis Media Anatomy of the ear Pinna The External Auditory canal The Tympanic Cavity Complications of Otitis Media: Classification of Otitis Media: Depending on The Duration of The Disease 2-4-1-1 Acute otitis media (AOM) 2-4-1-2 Chronic otitis media (COM) Depending on the nature of the fluid/discharge 2-4-2-1 Suppurative otitis media: (SOM) 2-4-2-2 Nonsuppurative otitis media (NSOM) otitis media with effusion Aerobic otitis media NO. Subject Pag Adhesive otitis media Acording to causative organisms Diagnosis of otitis media Incidence Factors affecting the occurrence of otitis media The Gender The Age The Socio Economic Status Level of Education Geographic Location The Infectious Agent Pseudomonas aeruginosa Moraxella catarrhalis Haemophilus influenzae Klebsiella pneumonia Escherichia coli Proteus mirabilis Staphylococcus aureus Streptococcus pneumonia: Polymerase chain reaction Principle of the PCR The denaturation Hybridization NO. Subject Pag Elongation Real time PCR (qPCR): Absoulute Quantitation Relative quantitation Chapter Three / Materials & methods Materials & methods Materials Laboratory equipments and instruments Chemicals and reagents Culture media Antibiotics Equipments Used to Extract DNA Materials Used in Extraction Polymerase Chain Reaction PCR devices 3-1-8-1 The devices of traditional PCR 3-1-8-2 The devices of qPCR Methods Sample collection Preparing the cultivation media 3-2-2-1 Ready-made Culture Media 3-2-2-2 synthetic Culture Media 3-2-2-2-1 Blood Agar Medium 3-2-2-2-2 Levienthal Media 3-2-2-2-3 Urea Agar Medium Preparation of reagents and solutions

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