DETERMINATION OF GROWTH CHARACTERISTICS OF THE FOOT AND MOUTH VIRUSES (A, O, ASIA-1) IN BHK-21 AN30 AND BHK-21 AN73 CELL CULTURES

Öz

Bu çalışmada, BHK-21 An₇₃ hücre kültürü ile BHK-21 An₃₀ hücre kültürlerinin üreme hızı, miktarı ve virus titresi üzerine etkilerinin araştırılması amaçlandı. Bu amaçla BHK-21 An₃₀ ve BHK-21 An₇₃ süspanse hücre kültürlerinde, hücrelerin üreme kinetikleri belirlendi. Bu hücre kültürlerinde A TUR 11, O TUR 07 ve Asia-1/11 şap aşı suşlarının üretimi gerçekleştirildi. BHK-21 An₃₀ hücre kültürlerinin 7 günlük inkubasyon periyodu esnasında monolayer formda 4. günde 1,3x10⁶/ml hücre sayısıyla, süspanse formda ise 4. günde 4,1x10⁵ hücre sayısı ile pik noktaya ulaşırken BHK-21 An₇₃  kültürlerinde ise monolayer formda 6. günde 2,2x10⁶/ml hücre sayısına ulaşılırken, süspanse formda 3. günde 2,8 x10⁵ hücre sayısına ulaştığı belirlendi.  Süspanse BHK-21 An₃₀ hücre kültürlerinde A, O ve Asia-1 aşı virusunun üretimi sonrası 146S değerleri sırasıyla ortalama; 0,51 μg/ml, 0,18 μg/ml ve 0,16 μg/ml olarak saptanırken enfektif titreleri ortalama 6,87 pfu/ml, 6,22 pfu/ml ve 6,49 pfu/ml olarak belirlendi.

Süspanse BHK-21 An₇₃ hücre kültürlerinde A, O ve Asia-1 aşı virusunun üretimi sonrası değerleri sırasıyla ortalama; 2,11 μg/ml, 2,59 μg/ml ve 0,53 μg/ml olarak saptanırken enfektif titreleri ortalama 6,99 pfu/ml, 7,82 pfu/ml ve 6,37 pfu/ml olarak tespit edildi.

Sonuç olarak, aşı üretiminde kullanılan A, O ve Asia-1 aşı viruslarının BHK-21 An₇₃ hücre kültüründe daha yüksek 146S ve enfektif titrede üreme gösterdiği ve bundan dolayı şap aşısı üretiminde BHK-21 An₇₃ hücre kültürlerinin kullanılmasının gerek üretim zamanı gerekse maliyet olarak BHK-21 An₃₀ hücre kültürlerinden daha uygun olduğu kanaatine varılmıştır. 

Anahtar Kelimeler

• Şap Hastalığı
• Şap Virusu suşları
• BHK-21 hücre kültürü

DETERMINATION OF GROWTH CHARACTERISTICS OF THE FOOT AND MOUTH VIRUSES (A, O, ASIA-1) IN BHK-21 AN30 AND BHK-21 AN73 CELL CULTURES

Abstract

In this study, it was aimed to investigate BHK-21An₇₃ and BHK-21An₃₀ growth rate of cell cultures and the effects on foot and mouth disease (FMD) vaccine virus strains titers. For this purpose, growth rate of suspended cell cultures of BHK-21An₇₃ and BHK-21An₃₀ were determined. A TUR 11, O TUR 07 and Asia-1/11 strains of foot and mouth disease (FMD) vaccine virus strains were produced separately on BHK-21An₃₀ and BHK-21An_73. BHK-21An₃₀ cell amounts during seven incubation days of reached to peak level in monolayer form at the 4th day with 1,3x10⁶/ml cell numbers, in suspension form at the 4th day with 4,1x10⁵ cell numbers. BHK-21An₇₃ reached to peak level in monolayer form at the 6th day 2,2x10⁶/ml cell numbers while in suspension form at the 3rd day with 2,8 x10⁵ cell numbers. After the production of A, O and Asia-1 vaccine viruses, in suspension BHK-21An₃₀ cell cultures average of 146S values respectively; 0,51 μg/ml, 0,18 μg/ml and 0,16 μg/ml were detected while infective titers were determined as average 6,87 (Plaque Forming Units) pfu/ml, 6,22 pfu/ml and 6,49 pfu/ml. After the production of A, O and Asia-1 vaccine viruses, in suspended BHK-21An₇₃ cell cultures average of 146S values respectively; 2,11 μg/ml, 2,59 μg/ml and 0,53 μg/ml were detected while infective titers were determined as average 6,99 pfu/ml, 7,82 pfu/ml and 6,37 pfu/ml. As a result, in the production of A, O and Asia-1 vaccine viruses in BHK-21An_73, resulted in higher 146S values and higher infective titers than BHK-21An_30. It is concluded that for FMD vaccine production the using of BHK-21An₇₃ cell culture is more appropriate for both of cost of manufacture and productive time.

Keywords

• Foot and Mouth Disease
• FMDV vaccine strains
• BHK-21 cell culture

References

1. 1. Abbas, F., Khan, F, A., Ahmad, F., Hussain, A., Ahmad, M., Awan, M.A., Tariq, M.,M., Kakar, M. A., Wadood, A., Ali, M. (2011). Production of Foot and Mouth Disease Virus Vaccine (O Type) on BHK-21 Cell Line, Iğdır Univ. J. Ins. Sci. Tech. 1(2), 155-159.
2. 2. Ali, S.,M., Ismail, A.,H., Soliman, E.,M., Hanaa, A., M. (2013). Studies on Growth Kinetics of the FMDV Serotype SAT-2 Egyptian Strain in Cell Culture, J.Vet.Adv. 3(2), 92-97
3. 3. Alexandersen, S., Zhang, Z., Donaldson, A., I., Garland, A., J., M. (2003). The Pathogenesis and Diagnosis of Foot and Mouth Disease. J.Comp Path. 129, 1-36.
4. 4. Bartelling, S., J., Vreeswijk, J. (1991). Developments in foot- and-mouth disease vaccines. Central Veterinary Institute, Lelystad, The Netherlands. Vaccine. Feb, 9(2),5-88.
5. 5. Bartelling, S., J. (2002). Development and performance of inactivated vaccines against foot and mouth disease. Rev. Sci. Tech. Off.int. Epiz, 21, 577-588.
6. 6. Grubman, M.,J., Baxt, B. (2004). Foot-and-Mouth Disease. Clin. Microbiol. Rev., 17 (2), 465–493
7. 7. Harmsen, M.,M., Fitjen, H.,P.,D., Westra, D.,F., Coco-martin, J., M. (2011). Effect of thiomersal on dissociation of intact (146S) foot-and-mouth disease virions into 12S particles as assessed by novel ELISAs specific for either 146S or 12S particles. Vaccine, 29, 2682-2690.
8. 8. Institute of Foot and Mouth Disease Protocol, 2010.

9. 9. Radwan MEI, Khalifa NO, Fahmy HA (2016) Molecular epidemiology of foot and mouth disease virus during 2014 with references to biochemical changes in Egyptian buffaloes . World Journal of Biology and Medical Sciences. 3(1)68-81.
10. 10. Rahman, S., U., Rabbani, M., Sahidullah, K., Muhammed, Z. (2007). Studies on In Vitro Culture Characteristics of Adherent Baby Hamster Kidney-21(BHK-21) Cell Line. Int. J. of Agri-Biol. 1560-8530, 821-826.
11. 11. Rweyeamu, M.,M., Umehara, O., Giorci, W., Medeiros, R., Lucca, D.,N., Balzatar, M. (1989), Effect of formaldehyde and binary ethyleneimine (BEI) on the Integrity of Foot and Mouth Disease Virus Capsid. Rev. Sci. Tech. Off. Int. Epiz., 8(3), 747-764.
12. 12.Shirai, J., Chatchawanchonteera, A., Sinsuwongwat, W., Makarasen, P., Sugimura,T. (1990), Estimation of 146S Paticles in Foot and Mouth Disease Virus (FMDV) Vaccine by Using Computer Analysing System. Jpn.J.Vet., Sci.52(3):621-630.