Dataset Information

Molecular analysis and epidemiological typing of Vancomycin-resistant Enterococcus outbreak strains.

ABSTRACT: Outbreaks of multidrug resistant bacteria including vancomycin-resistant enterococci (VRE) in healthcare institutions are increasing in Norway, despite a low level of resistance compared to other European countries. In this study, we describe epidemiological relatedness of vancomycin-resistant Enterococcus faecium isolated during an outbreak at a Norwegian hospital in 2012-2013. During the outbreak, 9454 fecal samples were screened for VRE by culture and/or PCR. Isolates from 86 patients carrying the vanA resistance gene were characterized using pulsed-field gel electrophoresis (PFGE), MALDI-TOF mass spectrometry and single nucleotide polymorphism typing. PFGE revealed two main clusters, the first comprised 56 isolates related to an initial outbreak strain, and the second comprised 21 isolates originating from a later introduced strain, together causing two partly overlapping outbreaks. Nine isolates, including the index case were not related to the two outbreak clusters. In conclusion, the epidemiological analyses show that the outbreak was discovered by coincidence, and that infection control measures were successful. All typing methods identified the two outbreak clusters, and the experiment congruence between the MALDI-TOF and the PFGE clustering was 63.2%, with a strong correlation (r?=?72.4%). Despite lower resolution compared to PFGE, MALDI-TOF may provide an efficient mean for real-time monitoring spread of infection.

SUBMITTER: Rangberg A

PROVIDER: S-EPMC6695431 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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Molecular analysis and epidemiological typing of Vancomycin-resistant Enterococcus outbreak strains.

Rangberg Anbjørg A Larsen Astri Lervik AL Kacelnik Oliver O Sæther Hanne Skarpodde HS Bjørland Marthe M Ringstad Jetmund J Jonassen Christine Monceyron CM

Scientific reports 20190815 1

Outbreaks of multidrug resistant bacteria including vancomycin-resistant enterococci (VRE) in healthcare institutions are increasing in Norway, despite a low level of resistance compared to other European countries. In this study, we describe epidemiological relatedness of vancomycin-resistant Enterococcus faecium isolated during an outbreak at a Norwegian hospital in 2012-2013. During the outbreak, 9454 fecal samples were screened for VRE by culture and/or PCR. Isolates from 86 patients carryin ...[more]

PMID: 31417140

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Project description:We studied the clinical isolates Enterococcus faecium NEF1, resistant to high levels of vancomycin (MIC, 512 microg/ml) and teicoplanin (MIC, 64 microg/ml); Enterococcus faecium BM4653 and BM4656 and Enterococcus avium BM4655, resistant to moderate levels of vancomycin (MIC, 32 microg/ml) and to low levels of teicoplanin (MIC, 4 microg/ml); and Enterococcus faecalis BM4654, moderately resistant to vancomycin (MIC, 16 microg/ml) but susceptible to teicoplanin (MIC, 0.5 microg/ml). The strains were distinct, were constitutively resistant via the synthesis of peptidoglycan precursors ending in D-alanyl-D-lactate, and harbored a chromosomal vanD gene cluster that was not transferable. New mutations were found in conserved domains of VanS(D): at T(170)I near the phosphorylation site in NEF1, at V(67)A at the membrane surface in BM4653, at G(340)S in the G2 ATP-binding domain in BM4655, in the F domain in BM4656 (a 6-bp insertion), and in the G1 and G2 domains of BM4654 (three mutations). The mutations resulted in constitutivity, presumably through the loss of the phosphatase activity of the sensor. The chromosomal Ddl D-Ala:D-Ala ligase had an IS19 copy in NEF1, a mutation in the serine (S(185)F) or near the arginine (T(289)P) involved in D-Ala1 binding in BM4653 or BM4655, respectively, and a mutation next to the lysine (P(180)S) involved in D-Ala2 binding in BM4654, leading to the production of an impaired enzyme. In BM4653 vanY(D), a new insertion sequence, ISEfa9, belonging to the IS3 family, resulted in the absence of D,D-carboxypeptidase activity. Strain BM4656 had a functional D-Ala:D-Ala ligase, associated with high levels of both VanX(D) and VanY(D) activities, and is the first example of a VanD-type strain with a functional Ddl enzyme. Study of these five clinical isolates, displaying various assortments of mutations, confirms that all VanD-type strains isolated so far have undergone mutations in the vanS(D) or vanR(D) gene, leading to constitutive resistance, but that the Ddl host ligase is not always impaired. Based on sequence differences, the vanD gene clusters could be assigned to two subtypes: vanD-1 and vanD-4.

Project description:Enterococcus faecium clinical isolates A902 and BM4538, which were resistant to relatively high levels of vancomycin (128 and 64 microg/ml, respectively) and to low levels of teicoplanin (4 microg/ml), and Enterococcus faecalis clinical isolates BM4539 and BM4540, which were resistant to moderate levels of vancomycin (16 microg/ml) and susceptible to teicoplanin (0.25 microg/ml), were studied. They were constitutively resistant by synthesis of peptidoglycan precursors ending with d-alanyl-d-lactate and harbored a chromosomal vanD gene cluster which was not transferable by conjugation to other enterococci. VanX(D) activity, which is not required in the absence of d-Ala-d-Ala, was low in the four strains, although none of the conserved residues was mutated; and the constitutive VanY(D) activity in the membrane fractions was inhibited by penicillin G. The mutations E(13)G in the region of d-alanine:d-alanine ligase (which is implicated in d-Ala1 binding in A902) and S(319)N of the serine involved in ATP binding in BM4538 and a 7-bp insertion at different locations in BM4539 and BM4540 (which led to putative truncated proteins) led to the production of an impaired enzyme and accounted for the lack of d-Ala-d-Ala-containing peptidoglycan precursors. The same 7-bp insertion in vanS(D) of BM4539 and BM4540 and a 1-bp deletion in vanS(D) of A902, which in each case led to a putative truncated and presumably nonfunctional protein, could account for the constitutive resistance. Strain BM4538, with a functional VanS(D), had a G(140)E mutation in VanR(D) that could be responsible for constitutive glycopeptide resistance. This would represent the first example of constitutive van gene expression due to a mutation in the structural gene for a VanR transcriptional activator. Study of these four additional strains that could be distinguished on the basis of their various assortments of mutations confirmed that all VanD-type strains isolated so far have mutations in the ddl housekeeping gene and in the acquired vanS(D) or vanR(D) gene that lead to constitutive resistance to vancomycin.

Project description:BackgroundEnterococcus faecium is ranked worldwide as one of the top ten pathogens identified in healthcare-associated infections (HAI) and is classified as one of the high priority pathogens for research and development of new antibiotics worldwide. Due to molecular biology techniques' higher costs, the approach for identifying and controlling infectious diseases in developing countries has been based on clinical and epidemiological perspectives. Nevertheless, after an abrupt vancomycin-resistant Enterococcus faecium dissemination in the Méderi teaching hospital, ending up in an outbreak, further measures needed to be taken into consideration. The present study describes the vancomycin-resistant Enterococcus faecium pattern within Colombian's largest installed-bed capacity hospital in 2016.MethodsThirty-three vancomycin-resistant Enterococcus faecium isolates were recovered during a 5-month period in 2016. Multilocus variable-number tandem-repeat analysis was used for molecular typing to determine clonality amongst strains. A modified time-place-sequence algorithm was used to trace VREfm spread patterns during the outbreak period and estimate transmission routes.ResultsFour clonal profiles were identified. Chronological clonal profile follow-up suggested a transitional spread from profile "A" to profile "B", returning to a higher prevalence of "A" by the end of the study. Antibiotic susceptibility indicated high-level vancomycin-resistance in most isolates frequently matching vanA gene identification.DiscussionTransmission analysis suggested cross-contamination via healthcare workers. Despite epidemiological control of the outbreak, post-outbreak isolates were still being identified as having outbreak-related clonal profile (A), indicating reduction but not eradication of this clonality. This study supports the use of combined molecular and epidemiological strategies in an approach to controlling infectious diseases. It contributes towards a more accurate evaluation of the effectiveness of the epidemiological measures taken regarding outbreak control and estimates the main cause related to the spread of this microorganism.

Dataset Information

Molecular analysis and epidemiological typing of Vancomycin-resistant Enterococcus outbreak strains.

Publications

Molecular analysis and epidemiological typing of Vancomycin-resistant Enterococcus outbreak strains.

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