During a survey of extended-spectrum beta-lactamases (ESBLs) in Bulgaria from 1996 to 2003, a TEM-3-like ESBL was detected in strains of Klebsiella pneumoniae, Escherichia coli, Citrobacter freundii and Klebsiella oxytoca from three centres in three different towns. The nucleotide sequence of the cloned gene was identical to that of TEM-3, except for one substitution (C347A) causing an amino acid exchange at position 49 from leucine to methionine. This TEM-3 variant with both a unique nucleotide and amino acid sequence was designated TEM-139. Transformants producing TEM-3 or TEM-139 expressed identical beta-lactam resistance phenotypes. TEM-139 was the only TEM-type ESBL detected in the surveyed hospitals (seven centres in three towns). TEM-139 is a natural variant of TEM-3 with an amino acid exchange without informational content, detectable only by molecular procedures, e.g. a nucleotide-specific polymerase chain reaction.