Molecular analysis of Brazilian infectious bronchitis field isolates by reverse transcription-polymerase chain reaction, restriction fragment length polymorphism, and partial sequencing of the N gene

Abstract

Molecular analysis of 15 Brazilian infectious bronchitis virus (IBV) isolates, obtained from clinical outbreaks of the disease in chickens (broilers or layers) in the state of Minas Gerais (Brazil) between 1972 and 1989, is reported. Using the N protein gene as target, IBVs were analyzed by reverse transcription-polymerase chain reaction/restriction fragment length polymorphism (RT-PCR/RFLP) with the restriction enzymes AvaII, HphI, Sau96I, and Tsp509I and cDNA sequencing. Results obtained from those isolates were compared to 19 sequences available in GenBank. N gene RFLP profiles, cDNA sequences, and predicted amino acid composition were used for the construction of dendrograms. Brazilian isolates were grouped into one distinct group. Identity of predicted N protein amino acid composition varied from 45% (between isolates G and 208) up to 99% (PM 1 and PM2), and, when compared to the other IBVs, the amino acid identity was from 42% (Q3/88 and G) up to 97% (D41 and PM1). The great genetic diversity was shown to occur before the official use of vaccination in Brazil and has remained thereafter.