Genetic Diversity of Carbapenem-Hydrolyzing Metallo-β-Lactamases from Chryseobacterium ( Flavobacterium ) indologenes

Abstract

ABSTRACT The class B carbapenem-hydrolyzing β-lactamase IND-1 has been characterized for Chryseobacterium indologenes strain 001. With internal primers for the bla gene for IND-1 ( bla IND-1 ) and an internal bla IND-1 probe, PCR amplifications failed, while hybridization results were positive when DNA from another C. indologenes isolate, strain CIP101026, was used as a template. Thus, a bla IND -related gene was cloned from this C. indologenes reference strain. Sequencing of the insert of a recombinant plasmid conferring resistance to carbapenems revealed an open reading frame with a G + C content of 39.9% and coding for a 243-amino-acid preprotein named IND-2. IND-2 shared 80% amino acid identity with IND-1 and had a similar broad-spectrum resistance profile, including resistance to carbapenems. It was classified in functional subgroup 3a of class B carbapenem-hydrolyzing β-lactamases. IND-1 and IND-2, despite their genetic diversity, possessed similar kinetic parameters, except that ceftazidime was hydrolyzed less by IND-2. To obtain the entire bla IND -related gene sequences of eight other C. indologenes isolates, PCR was performed using internal and external primers, followed by inverse PCR techniques. The likely chromosome-mediated metallo-β-lactamases of the 10 C. indologenes isolates were divided into several groups and subgroups. IND-1, IND-2, IND-2a, IND-3, and IND-4 shared 77 to 99% amino acid identity.