Differentiation of Infectious Bronchitis Virus Serotypes Using Polymerase Chain Reaction and Restriction Fragment Length Polymorphism Analysis

Polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analysis were used to differentiate between serotypes of several infectious bronchitis virus (IBV) strains. A sequence of 1720 base pairs (bp) that contains the S1 glycoprotein gene of IBV was amplified by PCR, purified, and digested with restriction enzymes. Eleven reference IBV strains were grouped according to the RFLP patterns. The IBV Holte, Arkansas DPI, SE 17, Md 27, and Iowa 97 strains could be differentiated from the other IBV strains using the restriction enzyme HaeIII. The Beaudette, Massachusetts 41, Connecticut, and Florida 88 strains had the same HaeIII RFLP pattern but could be differentiated using XcmI and BstYI restriction enzymes. The Gray and JMK strains could not be differentiated by their RFLP patterns following digestion with 23 different restriction enzymes. Twenty-six samples (field isolates and reference strains) of IBV, previously serotypes by the virus-neutralization (VN) test in embryonating eggs, were analyzed in a blind fashion. The results using the PCR and RFLP analysis agreed with the serotype for traditional and variant IBV viruses as determined by the VN test.