Open Access Open Access  Restricted Access Subscription Access

Using Real-Time Polymerase Chain Reaction as an Alternative Rapid Method for Enumeration of Colony Count in Live Brucella Vaccines


Affiliations
1 Central Laboratory for Evaluation of Veterinary Biologics, Cairo, Abbasia, Egypt
2 Department of Microbiology and Immunology, National Research Center, Egypt
 

Aim: Brucellosis is a major bacterial zoonosis of global importance affecting a range of animal species and man worldwide. It has economic, public health, and bio-risk importance. Control and prevention of animal brucellosis mainly depend on accurate diagnostic tools and implementation of effective and safe animal vaccination program. There are three types of animal Brucella live vaccines-Brucella melitensis Rev-1 vaccine, Brucella abortus S19, and B. abortus RB51. Evaluation of these vaccines depends mainly on enumeration of Brucella viable count. At present, used colony count method is time consuming, costly and requires especial skills. Hence, the aim of this study is to use and standardize real-time polymerase chain reaction (RT-PCR) as an alternative, quantitative, sensitive, and rapid method to detect the colony count of Brucella in live Brucella vaccine.
Materials and Methods: Four batches of different live Brucella vaccines were evaluated using of conventional bacterial count and RT-quantitative PCR (RT-qPCR) using BSCP31 gene specific primers and probe. Standard curve was generated from DNA template extracted from 10-fold serial dilution of living B. abortus RB51 vaccine to evaluate the sensitivity of RT-qPCR.
Results: Results revealed that three batches of living Brucella vaccines were acceptable for Brucella colony count when traditional bacterial enumeration method was used. Results of RT-qPCR were identical to that of conventional bacterial count.
Conclusions: Results concluded that RT-qPCR was relatively sensitive compared to traditional bacterial colony count of these vaccines.

Keywords

Brucella, Colony Count, RB51, Rev-1, Real-Time Polymerase Chain Reaction, S19, Vaccines.
User
Notifications
Font Size

Abstract Views: 136

PDF Views: 0




  • Using Real-Time Polymerase Chain Reaction as an Alternative Rapid Method for Enumeration of Colony Count in Live Brucella Vaccines

Abstract Views: 136  |  PDF Views: 0

Authors

Waleed S. Shell
Central Laboratory for Evaluation of Veterinary Biologics, Cairo, Abbasia, Egypt
Mahmoud L. Sayed
Central Laboratory for Evaluation of Veterinary Biologics, Cairo, Abbasia, Egypt
A. A. Samy
Department of Microbiology and Immunology, National Research Center, Egypt
Ghada Mohamed Al-Sadek
Central Laboratory for Evaluation of Veterinary Biologics, Cairo, Abbasia, Egypt
Gina Mohamed Mohamed Abd El-Hamid
Central Laboratory for Evaluation of Veterinary Biologics, Cairo, Abbasia, Egypt
Abdel Hakam M. Ali
Central Laboratory for Evaluation of Veterinary Biologics, Cairo, Abbasia, Egypt

Abstract


Aim: Brucellosis is a major bacterial zoonosis of global importance affecting a range of animal species and man worldwide. It has economic, public health, and bio-risk importance. Control and prevention of animal brucellosis mainly depend on accurate diagnostic tools and implementation of effective and safe animal vaccination program. There are three types of animal Brucella live vaccines-Brucella melitensis Rev-1 vaccine, Brucella abortus S19, and B. abortus RB51. Evaluation of these vaccines depends mainly on enumeration of Brucella viable count. At present, used colony count method is time consuming, costly and requires especial skills. Hence, the aim of this study is to use and standardize real-time polymerase chain reaction (RT-PCR) as an alternative, quantitative, sensitive, and rapid method to detect the colony count of Brucella in live Brucella vaccine.
Materials and Methods: Four batches of different live Brucella vaccines were evaluated using of conventional bacterial count and RT-quantitative PCR (RT-qPCR) using BSCP31 gene specific primers and probe. Standard curve was generated from DNA template extracted from 10-fold serial dilution of living B. abortus RB51 vaccine to evaluate the sensitivity of RT-qPCR.
Results: Results revealed that three batches of living Brucella vaccines were acceptable for Brucella colony count when traditional bacterial enumeration method was used. Results of RT-qPCR were identical to that of conventional bacterial count.
Conclusions: Results concluded that RT-qPCR was relatively sensitive compared to traditional bacterial colony count of these vaccines.

Keywords


Brucella, Colony Count, RB51, Rev-1, Real-Time Polymerase Chain Reaction, S19, Vaccines.