Bruselloz aşısı için aday olarak Brucella abortus'un dış zar veziküllerinin ekstraksiyonu ve biyolojik değerlendirmesi

Öz

Amaç: Bruselloz, hayvan ve insan sağlığını etkileyen endemik bir zoonotik hastalıktır. Son birkaç on yılda, hastalığı kontrol etmek için daha güvenli Brucella aşıları geliştirmek için çok araştırma yapılmıştır. Dış zar vezikülleri (OMV'ler), insan brusellozunu önlemek için bir alt birim aşı için immünojenik yapılar olarak düşünülmüştür. Bu çalışmanın amacı Brusella abortus'un dış zar veziküllerini bruselloz aşısı için aday olarak değerlendirmektir.
Yöntemler: Bu çalışmada Brucella abortus, S99 suşu kullanıldı. Brucella abortusun kitle kültivasyonundan sonra OMV ekstraksiyonu, Ultracentrifugation ve Sodium Dosoxycotinate tarafından gerçekleştirildi. SDS-PAGE yöntemi, fizikokimyasal özellikleri değerlendirmek için protein paternini ve elektron mikroskopisini gözlemlemek için kullanıldı. OMV'deki LOS miktarı da LAL testi ile ölçüldü. Protein konsantrasyonunun belirlenmesinden sonra farelere belirli bir konsantrasyonda enjekte edildi ve antikoru değerlendirmek için kan örnekleri alındı. Son olarak serum izolasyonundan sonra antikor üretimi ELISA yöntemi ile ölçüldü.
Bulgular: OMV'de mevcut protein miktarı 0,1 mg/ml idi. OMV'nin boyutu 55 ila 150 nm idi ve SDS-PAGE değerlendirmesinde protein aralığı >25 kDa idi. LAL testindeki LOS değeri yetkili aralıkta rapor edildi. İlk enjeksiyondan sonra IgG düzeylerinde (P=0,001) kontrol grubuna kıyasla ve daha sonraki güçlendiricilerde anlamlı artış gözlendi. En yüksek tepki oranı ikinci güçlendiricide gözlenmiştir.
Sonuç: OMV ekstraksiyon sırasında uzamsal şeklini korumaktadır ve Brucella düşüklerine karşı yüksek derecede spesifik antikorların üretimini indükleme potansiyeline sahiptir. Bu nedenle klinik fazları inceledikten sonra aday bir aşı olarak düşünülebilir.

Anahtar Kelimeler

• Dış mukoza vezikülleri
• Brucella abortus S99
• Bruselloz aşısı

Extraction and biological evaluation of external membrane vesicles of Brucella abortus as a candidate for brucellosis vaccine

Öz

Aim: Brucellosis is an endemic zoonotic disease affecting animal and human health. In the last several decades, much research has been performed to develop safer Brucella vaccines to control the disease. Outer membrane vesicles (OMVs) have been considered as immunogenic structures for a subunit vaccine to prevent human brucellosis. The aim of this study was to evaluate the external membrane vesicles of Brucella abortus as a candidate for brucellosis vaccine.
Methods: In this study Brucella abortus, S99 strain was used. Extraction of OMV after mass cultivation of Brucella abortus was performed by Ultracentrifugation and Sodium Dosoxycotinate. The SDS-PAGE method was used to observe the protein pattern and electron microscopy, to evaluate the physicochemical properties. The amount of LOS in OMV was also measured by LAL test. After determination of the protein concentration, mice were injected at a specific concentration and blood samples were obtained to evaluate the antibody. Finally, after serum isolation, the antibody production was measured by the ELISA method.
Results: The amount of protein present in OMV was 0.1 mg/ml. The size of OMV was 55 to 150 nm, and in the SDS-PAGE assessment, the protein range was >25 kDa. The LOS value in the LAL test was reported within the authorized range. Significant increase in IgG levels was observed after the first injection (P=0.001) compared to the control group, and in subsequent boosters as well. The highest response rate was observed in the second booster.
Conclusion: OMV maintains its spatial shape during extraction and has the potential to induce the production of a high degree of specific antibodies against Brucella abortus. For this reason, it can be considered as a candidate vaccine after examining clinical phases.

Anahtar Kelimeler

• External mucous vesicles
• Brucella aborotus
• Brucellosis vaccine

Kaynakça

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