The spike (S) glycoprotein gene (which encodes two subunits, S1 and S2), the membrane (M) glycoprotein gene and the gene which encodes the products of gene 3 are situated in the infectious bronchitis virus (IBV) genome in the order SI-S2-3-M. The S1 gene of four isolates of the Massachusetts (Mass) serotype, isolated between 1970 and 1984, each differed by only 2 to 3% from that of older Mass serotype isolates, including M41 and H120. Similarly, sequencing of the end of gene 3 and the beginning of the M gene showed that three of the isolates differed from the older Mass isolates by 2% or less. In contrast, the fourth Mass strain, Portugal/322/82, differed by 11% and 24% in gene 3, and the first part of gene M, respectively, suggesting that this strain was a recombinant. It was not possible to identify a putative recombination site but the finding that the S2 gene of Portugal/322/82 differed from other strains much less than did the S1 gene suggests that recombination might have occurred in S2 as a consequence of the high S2 nucleotide homology among IBV strains.