Molecular characterization of extended-spectrum β-lactamase-producing escherichia coli and clones causing extraintestinal infections in humans. Animals as reservoir of high-risk e. Coli clones pathogenic for humans

Resumen

Overall, we undertook a total of six studies of molecular epidemiology as well as a genomic analysis of Eschericha coli strains. Within the four first studies, we analysed a total of 394 strains which were producers of extraintestinal infections retrieved from two different hospitals (Lucus Augusti hospital in Lugo and the Beaujon hospital in París), whereas in the remaining two studies we analysed isolated strains coming from canine and porcine origin. In the first study, we undertake a molecular analysis of a total of 196 E. coli strains derived from consecutive cases of extraintestinal infections throughout 2016. We observed that 10% of the human infections were caused by ST131 clone and approximately 60% of these infections were caused by strains belonging to only 10 STs (ST10, ST12, ST58, ST69, ST73, ST88, ST95, ST127, ST131, ST141). A 37% of the strains were found to by multidrug-resistant (MDR) to antibiotics, of which an approximate of 40% belonged to four dominant clones: B2-CH40-30-ST131, B2-CH40-41-ST131, C-CH4-39-ST88 and D-CH35-27-ST69. In the second study, we completed a molecular analysis of a total of 188 extended-spectrum β-lactamase-producing Escherichia coli (ESBLEC) collected throughout 2015. Amongst ESBLEC, a 42% of strains belonged to clone ST131, among which more than 50% of the strains belonged to subclade C2, a 28% belonged to subclade C1 (20% C1M27 and 8% no-C1M27) and a 5% belonged to clade A. The remaining 58% included 57 different STs, the following being represented by at least three isolates: ST10 (8 isolates), ST23 (3), ST38 (4), ST58 (3), ST88 (5), ST95 (4), ST167 (3), ST354 (5), ST361 (3), ST410 (6), ST648 (4), ST744 (3), and ST1615 (6). Furthermore, we described two strains belonging to the emergent clone B2-CH14-64-ST1193. In the third study, we analyzed the early biofilm formation (EBF), of the 394 strains which produced extraintestinal infections in humans. Less than a 10% of the strains were able to quickly and persistently produce high levels of biofilm (G1 category). We described an association between clones B2-CH14-2-ST127, B2-CH40-22-ST131, B2-CH52-5/14-ST141 and E-CH100-96-ST362, some virulence factors encoding genes, the P2 protein variant of the fimB allele and the G1 group of biofilm producers. In the fourth study, we carried out the whole genome sequencing (WGS) of 75 ESBLEC strains, of which 30 belonged to different clades of the clone ST131. We performed the core genome analysis observing correlation with the phylogenetic groups of the genomes. This analysis allowed us to described two new clades among the ST131 clonal complex represented by strains with the fimH5 and fimH35 alleles. We noticed that the average of virulence genes is superior in genomes which belong to ST131 (STC131) against genomes no-STC131 and we identified a total of 228 plasmids. Moreover, we localized the genes encoding for ESBL enzymes in 67 out of the 75 genomes, of which 21 were integrated in the chromosome (31%) and 46 of them were carried by plasmids (69%). The 92% of the plasmid belonged to MOBF, MOBP and MOBQ families of relaxases. Besides, Col-like and IncF incompatibility group of plasmid were the most abundant. Furthermore, we observed a correlation between the presence of specific types of IncF and the different ST131 clades. In the fifth study, we analyzed a total of 197 strains recovered from the faeces of healthy dogs between the years 2013 and 2017. The 46% of strains were molecularly classified as extraintestinal pathogenic E. coli (ExPEC) and/or uropathogenic E. coli (UPEC), including 50 clones, among which four clones were dominant (B2-CH14-180-ST127, B2-CH52-14-ST141, B2-CH103-9-ST372 and F-CH4-58-ST648) and 15 were previously identified among isolates causing extraintestinal infections in humans. Besides, 14% were classified as MDR. We also analyzed the whole genome of 23 ST372 strains (21 from canine origin and two from human origin). Additionally, we performed a phylogenetic study including 174 genomes of ST372 strains obtained from public databases. These 197 genomes were segregated into six branches (cluster 1 to 6). Cluster 1 comprised 75% of the strain genomes, mostly composed of canine strain genomes. Clusters 4 and 6 also included canine strain genomes, while clusters 2, 3, and 5 were significantly associated with human strain genomes. In the sixth study, we performed the whole genome analysis of 11 strains belonging to the clone ST131 O25b:H4 clade B from porcine origin (seven isolates collected from diarrhoeic piglets and four from pork meat). We undertook its phylogenetic analysis including 73 ST131 reference genomes from different origins. These were taken from public databases. We observed that clade B genomes constituted nine phylogenetic branches (9 sub-clusters), allowing us to describe five new subclades (B6, B6-like, B7, B8, B9). The genomes from porcine strains belonged to subclades B6 and B7 and where closely related to some genomes from human strains. Besides, the genomes from porcine strains accommodated a great diversity of plasmids, resistance and virulence associated genes. We described the presence of the mcr-1.1 gene that confer colistin resistance in 5 of 11 genomes from porcine strains associated with the presence of the epidemic plasmids IncX4 and IncHI2.