Reply to Furlan et al., “Importance of Sequencing To Determine Functional bla_TEM Variants”

REPLY

Furlan et al. (1) have made the useful observation that bla_TEM-116 contamination is still a problem with contemporary sequencing reagents. Their finding of bla_TEM-116 sequence only if another bla_TEM is absent is the expected result for sequencing software identifying the predominant nucleotides at positions 84 and 184 when a competing bla_TEM is in higher concentration than the contaminant. Hence, it is important that a control lacking sample DNA yields no PCR product.

We point out that, contrary to Furlan et al. (1), TEM-116 is not an extended-spectrum β-lactamase for the reasons elaborated in our original letter (2); we note that, unlike their claim that TEM variants on the now inactive Lahey site (http://www.lahey.org/studies) included unnatural ones, the requirements for assignment of a β-lactamase number specifically included the statement “To qualify for a new number the enzyme must have been found in nature and not created in the laboratory.” We also required sequencing of the full gene as a basis for β-lactamase molecular characterization. While no one would disagree with the conclusion by Furlan et al. (1) that “sequencing is essential for the determination of functional β-lactamase TEM variants,” it is equally essential to analyze function when a new sequencing variant is discovered, as this is increasingly an issue in today's era of whole-genome sequencing.