ESBL- and Carbapenemase-Producing Escherichia coli and Klebsiella pneumoniae among Bivalves from Portuguese Shellfish Production Areas

Abstract

Bivalves are filter-feeding organisms and biomarkers of bacterial pollution. Our study aimed to analyze the occurrence and characteristics of extended-spectrum β-lactamase (ESBL)- and carbapenemase-producing Escherichia coli among bivalves. A total of 522 bivalve samples were collected along Portuguese shellfish production areas. Homogenized samples were screened for E. coli contamination on corresponding selective plates, allowing for concomitant growth of Klebsiella pneumoniae. E. coli growth was observed in 39% of the samples. Subsequent selective screening identified nine samples (4.4%) contaminated with ESBL producers, corresponding to E. coli (n = 7) and K. pneumoniae (n = 2), while a single carbapenemase-producing K. pneumoniae (0.5%) was identified. ESBLs were all CTX-M-types commonly identified in human isolates, i.e., CTX-M-32 (n = 4), CTX-M-15 (n = 4), and CTX-M-14 (n = 1). The carbapenemase producer harbored the bla_GES-5 gene located on a ColE plasmid. Clonality was evaluated by multilocus sequence typing, identifying E. coli backgrounds as ST10, ST23, ST540, ST617, ST746, SLV206, and SLV2325, commonly identified among environmental and human strains. The K. pneumoniae isolates belonged to ST834, ST15, and DLV644. The occurrence of ESBL- and carbapenemase-producing Enterobacteriaceae in bivalves reveals how the marine environment constitutes a reservoir of critical bacterial pathogens, thus potentially representing a risk to human health.

Keywords: ESBL; Escherichia coli; Klebsiella pneumoniae; Portugal; bivalves; carbapenemases.

Figure 1

Localization of the 19 shellfish production areas. (a) Estuaries, (b) Coastal waters. ELM—Estuário do Lima; RIAV1—Ria de Aveiro, Triângulo das Correntes-Moacha; RIAV2—Ria de Aveiro, Canal de Mira; RIAV3—Ria de Aveiro, Canal Principal-Espinheiro; RIAV4—Ria de Aveiro, Canal de Ílhavo; EMN—Estuário do Mondego; LOB—Lagoa de Óbidos; ETJ1—Estuário do Tejo—Jusante Ponte Vasco da Gama; LAL—Lagoa de Albufeira; ESD1—Estuário do Sado, Esteiro da Marateca; ESD2—Estuário do Sado, Canal de Alcácer; EMR—Estuário do Rio Mira; L1—Litoral de Viana; L2—Litoral de Matosinhos; L3—Litoral de Aveiro; L4—Litoral Figueira da Foz—Nazaré; L5a—Litoral Peniche—Cabo Raso; L5b—Litoral Cabo Raso—Cabo Espichel; L6—Litoral Setúbal—Sines.