Environmental Microbiota Represents a Natural Reservoir for Dissemination of Clinically Relevant Metallo-β-Lactamases

Abstract

A total of 10 metallo-␤-lactamase-producing isolates of six different species, including Brevundimonas diminuta (n ‫؍‬ 3), Rhizobium radiobacter (n ‫؍‬ 2), Pseudomonas monteilii (n ‫؍‬ 1), Pseudomonas aeruginosa (n ‫؍‬ 2), Ochrobactrum anthropi (n ‫؍‬ 1), and Enterobacter ludwigii (n ‫؍‬ 1), were detected in the sewage water of a hospital. The presence of bla VIM-13 associated with a Tn1721-class 1 integron structure was detected in all but one of the isolates (E. ludwigii, which produced VIM-2), and in two of them (R. radiobacter), this structure was located on a plasmid, suggesting that environmental bacteria represent a reservoir for the dissemination of clinically relevant metallo-␤-lactamase genes.Metallo-␤-lactamases (MBLs) have emerged worldwide as a major source of acquired broad-spectrum ␤-lactam resistance. They hydrolyze virtually all classes of ␤-lactams (except monobactams), including carbapenems, which often represent the last option for the treatment of infections with multidrugresistant Gram-negative bacteria.There are two dominant types of transferable MBLs among clinical isolates, IMP and VIM (12). Most of the IMP-and VIM-type MBL genes are present as gene cassettes inserted into integrons located on the chromosome or on plasmids. These integrons may be associated with transposon-like structures which may contribute to their variable location and spread (12). Interestingly, these MBL genes have been found almost exclusively in the hospital setting and the role of nonclinical habitats as a reservoir for bacteria that carry these acquired resistance determinants has been poorly investigated (9,10).In this study, we evaluated the presence of MBL-producing bacteria in the sewage water of Son Llatzer Hospital (Mallorca, Spain) in order to obtain epidemiological data which could complement the results from a previous survey that investigated the incidence of MBL-producing Pseudomonas aeruginosa strains in this hospital (4).For this purpose, duplicates of up to six 10-fold dilutions of 1 liter of sewage water collected 20 m downstream of the hospital wastewater discharge site were plated on three different media specially designed for the selection of pseudomonads (Gould S1 agar [1], King B agar [6], and cetrimide agar [Merck, Darmstadt, Germany]) containing 30 g/ml ceftazidime (Combinopharm, Madrid, Spain). A total of 37 bacterial isolates corresponding to different colony morphologies were collected using this strategy and tested for the presence of MBL using the MBL Etest according to the manufacturer's instructions (bioMérieux, Marcy l'Etoile, France). Only 16 isolates were positive by this test.Although these 16 MBL Etest-positive isolates exhibited different colony morphologies, their identification by 16S rRNA gene amplification and sequencing as previously described (13) showed that they belonged to only eight different species, including Brevundimonas diminuta, Rhizobium radiobacter, Pseudomonas monteilii, P. aeruginosa, Ochrobactrum anthropi, Enterobacter ludwigii, Acinetobacter johnsonii, a...

Detection and Characterization of VIM-31, a New Variant of VIM-2 with Tyr224His and His252Arg Mutations, in a Clinical Isolate of Enterobacter cloacae

“…The occurrence of VIM-2 has nevertheless also been reported for clinical isolates of Enterobacteriaceae from Asia (17,24,39), Mexico (28), Argentina (16), and Tunisia (10). Moreover, a recent report showed the presence of VIM-2 in Enterobacter ludwigii isolated from sewage water of a hospital, highlighting the involvement of environmental bacteria as potential reservoirs for the dissemination of clinically relevant metallo-␤-lactamase resistance genes (37). In many subclass B1 (e.g., IMP and NDM) and B2 (e.g., CphA) metallo-␤-lactamases, the amino acid at position 224 (a conserved Lys residue), close to the active site, plays an important role in substrate binding (2,27).…”

The Complete Nucleotide Sequence of the Carbapenem Resistance-Conferring Conjugative Plasmid pLD209 from a Pseudomonas putida Clinical Strain Reveals a Chimeric Design Formed by Modules Derived from Both Environmental and Clinical Bacteria

“…In this context, Tn3-like elements such as Tn4652 have been described in the chromosome of a P. putida strain encompassing only tnpA and tnpC genes bracketed by IRs (30), a situation resembling the proposed precursor element of the arrangement now found in pLD209. Thus, we provide in this work further evidences of mobile genetic elements inserting into one another, thus generating novel antimicrobial resistance-conferring platforms (31,32). Additional studies are certainly necessary to determine whether the mobilizable Tn402-like class 1 integron element present in pLD209 has definite limits in the internal IRs or whether it may additionally extend also to the external IRs, alternatives that are in any case not mutually exclusive.…”

“…In this context, we noted previously (3) that the 5= region of the Tn402-like integron is identical to a class 1 integron found in carbapenem-resistant P. aeruginosa clinical strains (33), thus suggesting the clinical microbiota as its most recent source. Whether this bla VIM-2 module was directly provided by clinical bacteria or indirectly through environmental bacteria in contact with hospital effluents, representing reservoirs of clinically relevant M␤L genes (11,32), requires more comprehensive sequence analyses.…”

First Report of an Extensively Drug-Resistant VIM-2 Metallo-β-Lactamase-Producing Brevundimonas diminuta Clinical Isolate

“…The presence of bla VIM-13 associated with a Tn1721-class 1 integron structure was detected in all B. diminuta isolates. This structure was located on a plasmid, suggesting that environmental bacteria represent a reservoir for the dissemination of clinically relevant MBL genes (13).…”