Determination of hepatitis B virus genotypes by DNA sequence analysis in patients from ankara, Turkey

Hepatit B virusu (HBV) genotiplerinin dünya üzerindeki dağılımı coğrafi bölgelere göre farklılık göstermektedir. Ülkemizde yapılan çalışmalarda belirlenen HBV genotipi ise genotip D olup, homojen ve tek tip şeklinde yayılım saptanmaktadır. Bu çalışmanın amacı, Ankara’da Gazi Üniversitesi Hastanesine başvuran HBV enfeksiyonlu hastalarda HBV genotiplerinin belirlenmesidir. Çalışmaya, tümü HBsAg pozitif ve anti-HBs negatif olan 84 (52 erkek, 32 kadın) hasta örneği dahil edilmiştir. Hastaların %95.2’sinde anti-HBc, %47.6’sında HBeAg, %11.9’unda ise anti-HBe belirteçleri pozitif olup, HBV-DNA düzeyleri ortalama 5.7 x 107 ± 4.6 x 107 IU/ml; ALT değerleri ortalama 131 ± 171 IU/ml ve AST değerleri ortalama 98 ± 170 IU/ml olarak saptanmıştır. Örneklerden HBV-DNA ekstraksiyonu, fenokloroform yöntemi ile yapılmış, HBV-DNA S gen bölgesi polimeraz zincir reaksiyonu (PCR) ile amplifiye edilmiştir. PCR ürünlerinin döngüsel dizi analizi reaksiyonu; dideoksi zincir sonlanması yöntemine dayalı olan ticari bir kit (Cy5/5.5 Dye Primer Cycle Sequencing Kit; Visible Genetics, Kanada) kullanılarak gerçekleştirilmiştir. Elde edilen DNA dizileri floresans temelli otomatik bir DNA dizileme sisteminde (Long-Read Tower System, Visible Genetics, Kanada) okutulmuş ve analiz edilmiştir. Gen bankasından alınan yayınlanmış tüm genotiplerin S gen bölgesine ait DNA dizileriyle, hasta örneklerinden elde edilen DNA dizilerinin karşılaştırılarak değerlendirilmesi sonucunda, 84 örneğin tümünün (%100) HBV D genotipi ve ayw alttipi ile ilişkili olduğu görülmüştür. PHYLIP filogenetik analiz paket programı kullanılarak yapılan analiz sonucunda oluşturulan filogenetik ağaçlarda, çalışılan örneklerin D genotipine ait grupta kümelendiği izlenmiştir. Sonuç olarak, ülkemizdeki HBV suşlarının moleküler epidemiyolojisi ve ülkemizin içinde bulunduğu coğrafi konum ile uyumlu olarak, hastanemize başvuran hasta grubunda da başlıca genotipin D olduğu saptanmış ve HBV genotip tayininin, klinik yaklaşımların daha bilinçli olmasına olanak sağlayacağı düşünülmüştür.Hepatitis B virus (HBV) genotypes vary depending on the geographical region. The HBV genotype determined in Turkey has been genotype D which is found as the homogenously disseminated single genotype. The aim of this study was to determine HBV genotypes in a group of HBV infected patients who were admitted to a university hospital in Ankara, Turkey. Serum samples from HBsAg positive and anti-HBs negative 84 (52 male, 32 female) patients with HBV infection were included into the study. Anti-HBc was positive in 95.2%, HBeAg was positive in 47.6% and anti-HBe was positive in 11.9% of the patients. Mean HBV-DNA levels of the patients were 5.7 x 107 ± 4.6 x 107 IU/ml; mean ALT levels were 131 ± 171 IU/ml and mean AST levels were 98 ± 170 IU/ml. HBV-DNA was extracted from serum by the phenol-chloroform method and PCR was performed to amplify the S gene region of HBV-DNA. Cycle sequencing of PCR products was performed by a commercial “Cy5/Cy5.5 Dye Primer Cycle Sequencing Kit” (Visible Genetics, Canada) based on dideoxy chain termination method. The sequences were read and analyzed in an automated fluorescence-based DNA-sequencing system (Long-Read Tower System, Visible Genetics, Canada). The nucleotide sequences of the patient samples were compared with the previously reported sequences in gene bank for each genotype. According to the comparative analysis of S-sequences of all patient samples with the published sequences of the genotypes in gene bank, all of the 84 hepatitis B strains (100%) were shown to be related to D genotypic group, subtype ayw. A phylogenetic analysis was performed and phylogenetic trees were constructed using programs in the PHYLIP phylogeny inference package. The patient samples clustered within the genotypic group D. According to these results, the main HBV genotype in our patients was genotype D in accordance with the previous molecular epidemiologic information on HBV in this geographic area. HBV genotype determination may help to establish more rational clinical approach in the evaluation of HBV infected patients

Nasopharyngeal pneumococcal carriage in healthy Turkish children after 13-valent conjugated pneumococcal vaccine implementation in the national immunization program.

In Turkey, pneumococcal conjugated vaccine (PCV) was introduced to the national immunization program as PCV7 in 2008, and was replaced with PCV13 in 2011. The aims of this study were to investigate the effects of PCV13 on nasopharyngeal pneumococcal carriage (NPC) by determining the serotype distribution, and to identify risk factors for carriage, in healthy Turkish children