Antibiotics and antibiotic resistant bacteria enter wastewater treatment plants (WWTPs), an environment where resistance genes can potentially spread and exchange between microbes. Several antibiotic resistance genes (ARGs) were quantified using qPCR in three WWTPs of decreasing capacity located in Helsinki, Tallinn, and Tartu, respectively: sulphonamide resistance genes (sul1 and sul2), tetracycline resistance genes (tetM and tetC), and resistance genes for extended spectrum beta-lactams (blaoxa-58, blashv-34, and blactx-m-32). To avoid inconsistencies among qPCR assays we normalised the ARG abundances with 16S rRNA gene abundances while assessing if the respective genes increased or decreased during treatment. ARGs were detected in most samples; sul1, sul2, and tetM were detected in all samples. Statistically significant differences (adjusted p<0.01) between the inflow and effluent were detected in only four cases. Effluent values for blaoxa-58 and tetC decreased in the two larger plants while tetM decreased in the medium-sized plant. Only blashv-34 increased in the effluent from the medium-sized plant. In all other cases the purification process caused no significant change in the relative abundance of resistance genes, while the raw abundances fell by several orders of magnitude. Standard water quality variables (biological oxygen demand, total phosphorus and nitrogen, etc.) were weakly related or unrelated to the relative abundance of resistance genes. Based on our results we conclude that there is neither considerable enrichment nor purification of antibiotic resistance genes in studied conventional WWTPs.

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A plasmid carrying the colistin resistance gene mcr-1 was isolated from a pig slurry sample in Estonia. The gene was present on a 33,311-bp plasmid of the IncX4 group. mcr-1 is the only antibiotic resistance gene on the plasmid, with the other genes mainly coding for proteins involved in conjugative DNA transfer (taxA, taxB, taxC, trbM, and the pilX operon). The plasmid pESTMCR was present in three phylogenetically very different Escherichia coli strains, suggesting that it has high potential for horizontal transfer.A plasmid containing the mcr-1 gene causing colistin resistance was originally described in Escherichia coli strains from animals, animal products, and human samples in South China (1). This plasmid could cause serious problems when transferred into strains for which colistin is the last treatment option. Since the first report, sequence information from several antibiotic resistance programs has been screened for the presence of the mcr-1 gene (2-25). The gene was found in several samples from around the world. The mcr-1 genes described are present in several plasmid backbones. In addition to mcr-1, two new mcr gene variants were detected (14,20). Although by now we are aware that mcr-1 is not restricted to China, we need more information for a better

In this study, we aimed to characterize the population structure, drug resistance mechanisms, and virulence genes of Enterococcus isolates in Estonia. Sixty-one Enterococcus faecalis and 34 Enterococcus faecium isolates were collected between 2012 and 2014 across the country from various sites and sources, including farm animals and poultry (n = 53), humans (n = 12), environment (n = 24), and wild birds (n = 44). Clonal relationships of the strains were determined by whole-genome sequencing and analyzed by multi-locus sequence typing. We determined the presence of acquired antimicrobial resistance genes and 23S rRNA mutations, virulence genes, and also the plasmid or chromosomal origin of the genes using dedicated DNA sequence analysis tools available and/or homology search against an ad hoc compiled database of relevant sequences. Two E. faecalis isolates from human with vanB genes were highly resistant to vancomycin. Closely related E. faecalis strains were isolated from different host species. This indicates interspecies spread of strains and potential transfer of antibiotic resistance. Genomic context analysis of the resistance genes indicated frequent association with plasmids and mobile genetic elements. Resistance genes are often present in the identical genetic context in strains with diverse origins, suggesting the occurrence of transfer events.

BackgroundWe aimed to identify the main spreading clones, describe the resistance mechanisms associated with carbapenem- and/or multidrug-resistant P. aeruginosa and characterize patients at risk of acquiring these strains in Estonian hospitals.MethodsNinety-two non-duplicated carbapenem- and/or multidrug-resistant P. aeruginosa strains were collected between 27th March 2012 and 30th April 2013. Clinical data of the patients was obtained retrospectively from the medical charts. Clonal relationships of the strains were determined by whole genome sequencing and analyzed by multi-locus sequence typing. The presence of resistance genes and beta-lactamases and their origin was determined. Combined-disk method and PCR was used to evaluate carbapenemase and metallo-beta-lactamase production.ResultsForty-three strains were carbapenem-resistant, 11 were multidrug-resistant and 38 were both carbapenem- and multidrug-resistant. Most strains (54%) were isolated from respiratory secretions and caused an infection (74%). Over half of the patients (57%) were ≥ 65 years old and 85% had ≥1 co-morbidity; 96% had contacts with healthcare and/or had received antimicrobial treatment in the previous 90 days.Clinically relevant beta-lactamases (OXA-101, OXA-2 and GES-5) were found in 12% of strains, 27% of which were located in plasmids. No Ambler class B beta-lactamases were detected. Aminoglycoside modifying enzymes were found in 15% of the strains. OprD was defective in 13% of the strains (all with CR phenotype); carbapenem resistance triggering mutations (F170 L, W277X, S403P) were present in 29% of the strains. Ciprofloxacin resistance correlated well with mutations in topoisomerase genes gyrA (T83I, D87N) and parC (S87 L). Almost all strains (97%) with these mutations showed ciprofloxacin-resistant phenotype.Multi-locus sequence type analysis indicated high diversity at the strain level – 36 different sequence types being detected. Two sequence types (ST108 (n = 23) and ST260 (n = 18)) predominated. Whereas ST108 was associated with localized spread in one hospital and mostly carbapenem-resistant phenotype, ST260 strains occurred in all hospitals, mostly with multi-resistant phenotype and carried different resistance genotype/machinery.ConclusionsDiverse spread of local rather than international P. aeruginosa strains harboring multiple chromosomal mutations, but not plasmid-mediated Ambler class B beta-lactamases, were found in Estonian hospitals.Trial registrationThis trial was registered retrospectively in ClinicalTrials.gov (NCT03343119).Electronic supplementary materialThe online version of this article (10.1186/s12879-018-3421-1) contains supplementary material, which is available to authorized users.