Characterization of Third-Generation Cephalosporin-Resistant Escherichia coli Isolated from Pigs in Cuba Using Next-Generation Sequencing.

Resistance to third-generation cephalosporins (3GC) in Escherichia coli has been reported worldwide from humans and animals, but the situation in Cuba is still poorly understood. This study aimed to gain new insights into the phenotypic and genotypic characteristics of third-generation cephalosporin-resistant (3GC-R) E. coli isolated from pigs in Cuba. Rectal swabs from 215 healthy pigs were taken from different municipalities in the western region of Cuba and spread on MacConkey agar supplemented with cefotaxime and ceftazidime. Ninety-six isolates were identified as 3GC-R E. coli and 87.5% of them were resistant to at least three antibiotic classes as determined by the measurement of the minimum inhibitory concentration (MIC) of 14 antibiotics. Twenty-seven different isolates were selected for Illumina next-generation sequencing, and subsequent in silico analysis was performed for the detection of antibiotic resistance and virulence genes, plasmid incompatibility (Inc) groups, multilocus sequence typing (MLST), and core genome MLST (cgMLST). The sequenced isolates contained extended-spectrum ß-lactamase genes blaCTX-M-32 (n = 17), blaCTX-M-15 (n = 5), and blaCTX-M-55 (n = 4) as well as with pAmpC gene blaCMY-2 (n = 2). They also harbored genes for resistance to other clinically important classes of antibiotics, as well as several diverse virulence factors. The 3GC-R E. coli were genetically highly diverse, belonging to 16 different sequence types. IncX1 was the most frequent Inc group. The presence of 3GC-R E. coli in pigs from Cuba containing several different antibiotic resistance mechanisms emphasizes the need for surveillance programs and the establishment of strategies for the prudent use of antibiotics in food-producing animals.

Genetic Features of Extended-Spectrum ß-Lactamase-Producing Escherichia coli from Poultry in Mayabeque Province, Cuba.

A total of 434 poultry cloacal samples were collected from seven different farms in different years (2013-2015) in the Cuban province of Mayabeque and analyzed for the presence of third-generation cephalosporin-resistant Escherichia coli (3GC-R-Ec). Sixty-two 3GC-R-Ec isolates were recovered in total from the farms, with detection rates of 2.9% in 2013, 10.3% in 2014, and 28.7% in 2015. Characterization of 32 3GC-R-Ec isolates revealed the presence of the extended-spectrum ß-lactamase (ESBL) genes blaCTX-M-1 (n = 27), blaCTX-M-15 (n = 4), and blaCTX-M-1 together with blaLAP-2 (n = 1). The isolates also contained different proportions of genes conferring decreased susceptibility to sulfonamides (sul1, sul2, sul3), trimethoprim (dfrA1, dfrA7, dfrA12, dfrA14, dfrA17), tetracyclines (tet(A), tet(B)), aminoglycosides (aac(6')-Ib-cr, strA, strB), chloramphenicol (cmlA1, floR), macrolides (mph(A), mph(D)), and quinolones (qnrS, qnrB, aac(6')-Ib-cr) as well as mutations in the fluoroquinolone-resistance determining regions of GyrA (S83L, D87N, D87Y) and ParC (S80I, E84G). The isolates belonged to 23 different sequence types and to phylogroups A (n = 25), B1 (n = 5), and D (n = 2), and they contained plasmid-associated incompatibility groups FII, X1, HI1, HI2, N, FIA, and FIB. These findings reveal a genetically diverse population of multiresistant ESBL-producing E. coli in poultry farms in Cuba, which suggests multiple sources of contamination and the acquisition of antibiotic resistance genes.

Complete Circular Genome Sequence of a Multidrug-Resistant Escherichia coli Strain from Cuba Obtained with Nanopore and Illumina Hybrid Assembly.

The complete genome sequence of a multidrug-resistant Escherichia coli strain isolated from a healthy pig in Cuba was determined using short and long reads. This strain carried four plasmids, including a 42,683-kb IncX1 plasmid, which contains the third-generation cephalosporin resistance gene bla CTX-M-32 together with other disinfectant and antibiotic resistance genes.