بررسی مولکولی برخی ژن‌ها در ایزوله‌های کلبسیلا پنومونیه از موارد بالینی مختلف

نوع مقاله : مقاله پژوهشی

نویسندگان

1 گروه زیست‌شناسی، دانشکده آموزش، دانشگاه القادسیه، القادسیه، عراق

2 گروه زیست‌شناسی، دانشکده آموزش، دانشگاه القادسیه، القادسیه، عراق.

10.22103/jab.2025.25209.1702

چکیده

هدف: این مطالعه با هدف بررسی شیوع کلبسیلا پنومونیه به‌عنوان عامل اصلی ذات‌الریه در استان المثنی، عراق، و بررسی پروفایل مولکولی ژن‌های بیماری‌زایی مرتبط با هایپرموکوویسکوزیته (مانند magA و rmpA) در سویه‌های اپیدمیک جدا شده از نمونه‌های دستگاه تنفسی تحتانی بیماران بستری مبتلا به ذات‌الریه در بیمارستان الرمیثه انجام شد. علاوه بر این، شیوع سویه‌های هایپرویرولنت کلبسیلا پنومونیه (hvKp) و سویه‌های کلاسیک کلبسیلا پنومونیه (cKp) نیز برآورد گردید.
مواد و روش‌ها: در مجموع 100 ایزوله باکتریایی از نمونه‌های دستگاه تنفسی تحتانی بیماران در بیمارستان الرمیثه بین 18 آوریل 2024 و 18 سپتامبر 2024 جمع‌آوری شد. ایزوله‌ها از طریق گامه‌های زیر شناسایی شدند: مورفولوژی کلونی روی آگار مَک‌کانکی، مورفولوژی سلولی از طریق رنگ‌آمیزی گرم و کپسول و مشاهده زیر میکروسکوپ نوری، آزمایش‌های فیزیولوژیکی، آزمایش‌های بیوشیمیایی، شناسایی مولکولی با استفاده از توالی‌یابی ژن 16S rRNA، و تشخیص ژن‌های بیماری‌زایی (magA و rmpA) از طریق واکنش زنجیره‌ای پلیمراز (PCR) با استفاده از پرایمرهای اختصاصی ژن که قطعاتی به طول 1283 جفت‌باز برای magA و 409 و 340 جفت‌باز برای rmpA را تکثیر می‌کردند.
نتایج: از 100 ایزوله، 20 مورد پروفایل معمولی کلبسیلا پنومونیه را نشان دادند که هویت آن‌ها با توالی‌یابی ژن 16SrRNA تأیید شد. توالی‌های 15 سویه از این ایزوله‌ها در GenBank تحت شماره‌های دسترسی PQ814166 تا PQ814180 ثبت شدند. تحلیل فیلوژنتیک این 15 سویه را به 10 کلاد تقسیم کرد: کلاد A (PQ814167, PQ814177)، کلاد B (PQ814171)، کلاد C (PQ814166, PQ814176)، کلاد D (PQ814179, PQ814180)، کلادهای E، F، و G (به ترتیب PQ814169، PQ814173، و PQ814174)، کلاد H (PQ814170, PQ814175)، کلاد I (PQ814172) و کلاد J (PQ814168, PQ814178). ژن‌های هایپرموکوویسکوزیته magA و rmpA به‌ترتیب در 55% و 50% از 20 ایزوله کلبسیلا پنومونیه شناسایی شدند. شیوع سویه‌های هایپرویرولنت کلبسیلا پنومونیه (hvKp) 50% بود.
نتیجه‌گیری: این مطالعه تأیید می‌کند که کلبسیلا پنومونیه یک عامل مهم ذات‌الریه در استان المثنی، با حضور قابل‌توجه سویه‌های هایپرویرولنت (50%) که با ژن‌های magA و rmpA مشخص می‌شوند است. این یافته‌ها اهمیت بررسی مولکولی در درک بیماری‌زایی و اپیدمیولوژی کلبسیلا پنومونیه در محیط‌های بالینی را برجسته می‌کند.

کلیدواژه‌ها


عنوان مقاله [English]

Molecular characterization of some genes in Klebsiella pneumoniae isolates from different clinical cases

نویسندگان [English]

  • Marwa Faez Abd Al-Munem 1
  • Mithal K. A. Al-Hassani 2
1 Department of Biology, College of Education, University of Al-Qadisiyah, Al-Qadisiyah, Iraq
2 Department of Biology, College of Education, University of Al-Qadisiyah, Al-Qadisiyah, Iraq.
چکیده [English]

Objective
This investigation aimed to investigate the prevalence of Klebsiella pneumoniae as a primary causative agent of pneumonia in the Al-Muthanna governorate, Iraq, and to describe the molecular profile of virulence genes related to hypermucoviscosity (e.g., magA and rmpA) in epidemic strains extracted from lower respiratory tract samples of hospitalized patients with pneumonia at Al-Rumaytha Hospital. Additionally, we estimated the prevalence of hypervirulent K. pneumoniae (hvKp) and classical K. pneumoniae (cKp) strains.

Materials and Methods
A total of 100 bacterial isolates were gathered from lower respiratory tract samples of patients at Al-Rumaytha Hospital between April 18, 2024, and September 18, 2024. Isolates were identified through below steps: colonial morphology on MacConkey agar, cell morphology via Gram and capsule staining observed under a light microscope, physiological experiments, biochemical experiments, molecular identification applying 16S rRNA gene sequencing, and diagnosis of virulence genes (magA and rmpA) via polymerase chain reaction (PCR) with gene-specific primers amplifying fragments of 1283 bp for magA and 409 bp and 340 bp for rmpA.

Results
Of the 100 isolates, 20 exhibited a typical K. pneumoniae profile, with identity affirmed by 16S rRNA gene sequencing. Sequences from 15 of these strains were deposited in GenBank under accession numbers PQ814166 to PQ814180. Phylogenetic analysis grouped these 15 strains into 10 clades: Clade A (PQ814167, PQ814177), Clade B (PQ814171), Clade C (PQ814166, PQ814176), Clade D (PQ814179, PQ814180), Clades E, F, and G (PQ814169, PQ814173, PQ814174, respectively), Clade H (PQ814170, PQ814175), Clade I (PQ814172), and Clade J (PQ814168, PQ814178). The hypermucoviscosity genes magA and rmpA were detected in 55% and 50% of the 20 K. pneumoniae isolates, respectively. The prevalence of hypervirulent K. pneumoniae (hvKp) strains was 50%.

Conclusions
This investigation affirms K. pneumoniae as a meaningful cause of pneumonia in the Al-Muthanna governorate, with a notable presence of hypervirulent strains (50%) described by magA and rmpA genes. These results highlight the importance of molecular characterization in understanding the pathogenicity and epidemiology of K. pneumoniae in clinical settings.

کلیدواژه‌ها [English]

  • 16S rRNA
  • Klebsiella pneumoniae
  • magA gene
  • rmpA gene
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