Publications by authors named "Alexander W Friedrich"

148 Publications

Insight Into the Anti-staphylococcal Activity of JBC 1847 at Sub-Inhibitory Concentration.

Front Microbiol 2021 5;12:786173. Epub 2022 Jan 5.

Department of Veterinary and Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.

Multidrug-resistant pathogens constitute a serious global issue and, therefore, novel antimicrobials with new modes of action are urgently needed. Here, we investigated the effect of a phenothiazine derivative (JBC 1847) with high antimicrobial activity on , using a wide range of assays, flow cytometry, and RNA transcriptomics. The flow cytometry results showed that JBC 1847 rapidly caused depolarization of the cell membrane, while the macromolecule synthesis inhibition assay showed that the synthesis rates of DNA, RNA, cell wall, and proteins, respectively, were strongly decreased. Transcriptome analysis of exposed to sub-inhibitory concentrations of JBC 1847 identified a total of 78 downregulated genes, whereas not a single gene was found to be significantly upregulated. Most importantly, there was downregulation of genes involved in adenosintrifosfat (ATP)-dependent pathways, including histidine biosynthesis, which is likely to correlate with the observed lower level of intracellular ATP in JBC 1847-treated cells. Furthermore, we showed that JBC 1847 is bactericidal against both exponentially growing cells and cells in a stationary growth phase. In conclusion, our results showed that the antimicrobial properties of JBC 1847 were primarily caused by depolarization of the cell membrane resulting in dissipation of the proton motive force (PMF), whereby many essential bacterial processes are affected. JBC 1847 resulted in lowered intracellular levels of ATP followed by decreased macromolecule synthesis rate and downregulation of genes essential for the amino acid metabolism in . Bacterial compensatory mechanisms for this proposed multi-target activity of JBC 1847 seem to be limited based on the observed very low frequency of resistance toward the compound.
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http://dx.doi.org/10.3389/fmicb.2021.786173DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8766816PMC
January 2022

[We still have to continue to test, test, test].

Ned Tijdschr Geneeskd 2021 10 11;165. Epub 2021 Oct 11.

UMC Groningen, afd. Medische Microbiologie, Groningen.

Testing for COVID-19 is performed curatively for disease detection and as screening tool to prevent spread for public health and infection prevention. Testing for access is an important additional measure to provide information about asymptomatic or pre-symptomatic infection. In a recent meta-analysis 42.8% showed no symptoms with an estimated percentage of asymptomatic infections of 35.1%. False positive testing is not completely preventable but allows events such as the Eurovision Song Contest, professional football, the Olympic Games or Formula 1. Their trade-off is not purely medical but also political-economical. The choice to carry out many Dutch tests in foreign laboratories is inefficient, while at the same time the opportunity to strengthen regional innovation, training and infrastructure is missed. Cooperation between municipal health departments and regional laboratories and extra service from regional laboratories (such as fast variant PCR) are frustrated this way. We hope this situation will improve rapidly.
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October 2021

Long-read sequencing-based in silico phage typing of vancomycin-resistant Enterococcus faecium.

BMC Genomics 2021 Oct 23;22(1):758. Epub 2021 Oct 23.

Department of Medical Microbiology and Infection Prevention, University of Groningen, University Medical Center Groningen, Groningen, the Netherlands.

Background: Vancomycin-resistant enterococci (VRE) are successful nosocomial pathogens able to cause hospital outbreaks. In the Netherlands, core-genome MLST (cgMLST) based on short-read sequencing is often used for molecular typing. Long-read sequencing is more rapid and provides useful information about the genome's structural composition but lacks the precision required for SNP-based typing and cgMLST. Here we compared prophages among 50 complete E. faecium genomes belonging to different lineages to explore whether a phage signature would be usable for typing and identifying an outbreak caused by VRE. As a proof of principle, we investigated if long-read sequencing data would allow for identifying phage signatures and thereby outbreak-related isolates.

Results: Analysis of complete genome sequences of publicly available isolates showed variation in phage content among different lineages defined by MLST. We identified phage present in multiple STs as well as phages uniquely detected within a single lineage. Next, in silico phage typing was applied to twelve MinION sequenced isolates belonging to two different genetic backgrounds, namely ST117/CT24 and ST80/CT16. Genomic comparisons of the long-read-based assemblies allowed us to correctly identify isolates of the same complex type based on global genome architecture and specific phage signature similarity.

Conclusions: For rapid identification of related VRE isolates, phage content analysis in long-read sequencing data is possible. This allows software development for real-time typing analysis of long-read sequencing data, which will generate results within several hours. Future studies are required to assess the discriminatory power of this method in the investigation of ongoing outbreaks over a longer time period.
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http://dx.doi.org/10.1186/s12864-021-08080-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8542323PMC
October 2021

Molecular Characterisation of Vancomycin-Resistant Isolates Belonging to the Lineage ST117/CT24 Causing Hospital Outbreaks.

Front Microbiol 2021 27;12:728356. Epub 2021 Sep 27.

Department of Medical Microbiology and Infection Prevention, University of Groningen, University Medical Center Groningen, Groningen, Netherlands.

Vancomycin-resistant (VREfm) is a successful nosocomial pathogen. The current molecular method recommended in the Netherlands for VREfm typing is based on core genome Multilocus sequence typing (cgMLST), however, the rapid emergence of specific VREfm lineages challenges distinguishing outbreak isolates solely based on their core genome. Here, we explored if a detailed molecular characterisation of mobile genetic elements (MGEs) and accessory genes could support and expand the current molecular typing of VREfm isolates sharing the same genetic background, enhancing the discriminatory power of the analysis. The genomes of 39 VREfm and three vancomycin-susceptible (VSEfm) isolates belonging to ST117/CT24, as assessed by cgMLST, were retrospectively analysed. The isolates were collected from patients and environmental samples from 2011 to 2017, and their genomes were analysed using short-read sequencing. Pangenome analysis was performed on assemblies, which were also screened for known predicted virulence factors, antimicrobial resistance genes, bacteriocins, and prophages. Two representative isolates were also sequenced using long-read sequencing, which allowed a detailed analysis of their plasmid content. The cgMLST analysis showed that the isolates were closely related, with a minimal allelic difference of 10 between each cluster's closest related isolates. The vanB-carrying transposon 1549 was present in all VREfm isolates. However, in our data, we observed independent acquisitions of this transposon. The pangenome analysis revealed differences in the accessory genes related to prophages and bacteriocins content, whilst a similar profile was observed for known predicted virulence and resistance genes. In the case of closely related isolates sharing a similar genetic background, a detailed analysis of MGEs and the integration point of the -carrying transposon allow to increase the discriminatory power compared to the use of cgMLST alone. Thus, enabling the identification of epidemiological links amongst hospitalised patients.
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http://dx.doi.org/10.3389/fmicb.2021.728356DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8503688PMC
September 2021

Misidentification of meticillin-resistant by the Cepheid Xpert MRSA NxG assay, the Netherlands, February to March 2021.

Euro Surveill 2021 09;26(37)

University of Groningen, University Medical Center Groningen, Department of Medical Microbiology, Groningen, the Netherlands.

We describe two false-negative results in the detection of meticillin-resistant (MRSA) of sequence type 398 and type t011 using the Cepheid Xpert MRSA NxG assay. The isolates were recovered in late February and early March 2021 from two patients in different hospitals in the northern Netherlands. Variations between the two isolate genomes indicate that this MRSA strain might have been spreading for some time and could have disseminated to other regions of the Netherlands and other European countries.
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http://dx.doi.org/10.2807/1560-7917.ES.2021.26.37.2100800DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8447827PMC
September 2021

and virulence factors as biomarkers of infection.

Biol Chem 2021 11 10;402(12):1565-1573. Epub 2021 Sep 10.

Department of Medical Microbiology, University of Groningen, University Medical Center Groningen, Hanzeplein 1, NL-9713 GZ Groningen, The Netherlands.

The gold standard for the diagnosis of bacterial infections in clinical samples is based on culture tests that are time-consuming and labor-intense. For these reasons, an extraordinary effort has been made to identify biomarkers as the tools for sensitive, rapid and accurate identification of pathogenic microorganisms. Moreover, biomarkers have been tested to distinguish colonization from infection, monitor disease progression, determine the clinical status of patients or predict clinical outcomes. This mini-review describes and biomarkers, which contribute to pathogenesis and have been used in culture-independent bacterial identification directly from patient samples.
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http://dx.doi.org/10.1515/hsz-2021-0243DOI Listing
November 2021

Staphylococcal cassette chromosome mec containing a novel mec gene complex, B4.

J Antimicrob Chemother 2021 07;76(8):1986-1990

University of Groningen, University Medical Center Groningen, Department of Medical Microbiology, Groningen, The Netherlands.

Objectives: To describe a new subclass of mec class B complex identified in Staphylococcus epidermidis.

Methods: Four S. epidermidis isolates obtained from bloodstream infections in patients at University Medical Center Groningen (UMCG) were analysed by phenotypic antibiotic susceptibility testing and WGS.

Results: Sequence analysis revealed a new staphylococcal cassette chromosome mec (SCCmec) structure in isolate UMCG335. In this structure, plasmid pUB110 was found to be integrated into SCCmec IVc, creating a new SCCmec subtype, IVUMCG335. SCCmec IVc and a copy of plasmid pUB110 were found in other isolates, UMCG364 and UMCG341, respectively, indicating a probability that SCCmec IVUMCG335 could have evolved at the UMCG. SCCmec of UMCG337 contained a new genetic organization of the mec complex (IS431-ΔmecR1-mecA-IS431-pUB110-IS431-ψIS1272) that we have named B4. This new subclass of mec class B complex originated by IS431-mediated inversion of the DNA segment encompassing the plasmid and most of the genes of the mec complex with the exception of IS1272. As the SCCmec organization in UMCG337 differed by the inversion of an ∼10 kb sequence compared with SCCmec IVUMCG335, we have named it SCCmec subtype IVUMCG337. Isolates UMCG335 and UMCG337 carrying SCCmec IVUMCG335 and IVUMCG337, respectively, were associated with a restriction-modification system and a CRISPR-Cas system, creating a composite island of almost 70 kb.

Conclusions: Our findings highlight the importance of IS431 in the evolution of the SCCmec region. The increasing genetic diversity identified in the SCCmec elements imposes a great challenge for SCCmec typing methods and highlights possible difficulties with the SCCmec nomenclature.
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http://dx.doi.org/10.1093/jac/dkab154DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8283725PMC
July 2021

: a potential 'living antibiotic' to control bacterial pathogens.

Crit Rev Microbiol 2021 Sep 1;47(5):630-646. Epub 2021 May 1.

Department of Medical Microbiology and Infection Prevention, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands.

is a small Deltaproteobacterium which, since its discovery, has distinguished itself for the unique ability to prey on other Gram-negative bacteria. The studies on this particular "predatory bacterium", have gained momentum in response to the rising problem of antibiotic resistance, because it could be applied as a potential probiotic and antibiotic agent. Hereby, we present recent advances in the study of , comprehending fundamental aspects of its biology, obligatory intracellular life cycle, predation resistance, and potential applications. Furthermore, we discuss studies that pave the road towards the use of as a "living antibiotic" in human therapy, focussing on its interaction with biofilms, the host immune response, predation susceptibility and application models. The available data imply that it will be possible to upgrade this predator bacterium from a predominantly academic interest to an instrument that could confront antibiotic resistant infections.
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http://dx.doi.org/10.1080/1040841X.2021.1908956DOI Listing
September 2021

Predominance of CTX-M-15-producing ST131 strains among ESBL-producing Escherichia coli isolated from asylum seekers in the Netherlands.

J Antimicrob Chemother 2021 01;76(1):70-76

University of Groningen, University Medical Center Groningen, Department of Medical Microbiology, Groningen, The Netherlands.

Objectives: Numerous studies show increased prevalence of MDR bacteria amongst asylum seekers, but data on the molecular profiles of such strains are limited. We aimed to evaluate the molecular profiles of ESBL-producing Escherichia coli (ESBL-E. coli) strains isolated from asylum seekers and investigate their phylogenetic relatedness.

Methods: WGS data of ESBL-E. coli isolates from asylum seekers, retrieved from 1 January to 31 December 2016, were analysed to assess MLST STs, fim types, phylogroups and resistance genes. Fifty-two ESBL-E. coli isolates from the Dutch-German border region were used for genome comparison purposes as a control group.

Results: Among 112 ESBL-E. coli isolates from asylum seekers, originating mostly from Syria (n = 40) and Iraq (n = 15), the majority belonged to ST131 (21.4%) and ST10 (17.0%). The predominant gene for β-lactam resistance was blaCTX-M-15 (67.9%), followed by the often co-detected blaTEM-1B (39.3%). No mcr or carbapenemase genes were detected. The majority of the strains belonged to phylogroups B2 (38.4%) and A (32.1%), carrying fimH27 (25%) and fimH30 (19.6%). A core genome MLST minimum spanning tree did not reveal clusters containing strains from the asylum seekers and the control group. Five clusters were formed within the asylum seeker group, by strains isolated from people originating from different countries.

Conclusions: The most frequently isolated clones in this study were isolated on a regular basis within the Dutch population before the increase in the asylum seeker population. No mcr- or carbapenemase-producing clones were detected among the asylum seeker population. Minor clustering was observed amongst the asylum seeker strains.
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http://dx.doi.org/10.1093/jac/dkaa395DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7729386PMC
January 2021

Enterococcus faecium: from microbiological insights to practical recommendations for infection control and diagnostics.

Antimicrob Resist Infect Control 2020 08 10;9(1):130. Epub 2020 Aug 10.

Department of Medical Microbiology, University of Groningen, University Medical Center Groningen, Groningen, The Netherlands.

Early in its evolution, Enterococcus faecium acquired traits that allowed it to become a successful nosocomial pathogen. E. faecium inherent tenacity to build resistance to antibiotics and environmental stressors that allows the species to thrive in hospital environments. The continual wide use of antibiotics in medicine has been an important driver in the evolution of E. faecium becoming a highly proficient hospital pathogen.For successful prevention and reduction of nosocomial infections with vancomycin resistant E. faecium (VREfm), it is essential to focus on reducing VREfm carriage and spread. The aim of this review is to incorporate microbiological insights of E. faecium into practical infection control recommendations, to reduce the spread of hospital-acquired VREfm (carriage and infections). The spread of VREfm can be controlled by intensified cleaning procedures, antibiotic stewardship, rapid screening of VREfm carriage focused on high-risk populations, and identification of transmission routes through accurate detection and typing methods in outbreak situations. Further, for successful management of E. faecium, continual innovation in the fields of diagnostics, treatment, and eradication is necessary.
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http://dx.doi.org/10.1186/s13756-020-00770-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7418317PMC
August 2020

Detection of extended-spectrum beta-lactamase (ESBL) genes and plasmid replicons in using PlasmidSPAdes assembly of short-read sequence data.

Microb Genom 2020 07 26;6(7). Epub 2020 Jun 26.

Department of Medical Microbiology and Infection Prevention, University of Groningen, University Medical Center Groningen, Groningen, The Netherlands.

Knowledge of the epidemiology of plasmids is essential for understanding the evolution and spread of antimicrobial resistance. PlasmidSPAdes attempts to reconstruct plasmids using short-read sequence data. Accurate detection of extended-spectrum beta-lactamase (ESBL) genes and plasmid replicon genes is a prerequisite for the use of plasmid assembly tools to investigate the role of plasmids in the spread and evolution of ESBL production in . This study evaluated the performance of PlasmidSPAdes plasmid assembly for in terms of detection of ESBL-encoding genes, plasmid replicons and chromosomal wgMLST genes, and assessed the effect of k-mer size. Short-read sequence data for 59 ESBL-producing were assembled with PlasmidSPAdes using different k-mer sizes (21, 33, 55, 77, 99 and 127). For every k-mer size, the presence of ESBL genes, plasmid replicons and a selection of chromosomal wgMLST genes in the plasmid assembly was determined. Out of 241 plasmid replicons and 66 ESBL genes detected by whole-genome assembly, 213 plasmid replicons [88 %; 95 % confidence interval (CI): 83.9-91.9] and 43 ESBL genes (65 %; 95 % CI: 53.1-75.6) were detected in the plasmid assemblies obtained by PlasmidSPAdes. For most ESBL genes (83.3 %) and plasmid replicons (72.0 %), detection results did not differ between the k-mer sizes used in the plasmid assembly. No optimal k-mer size could be defined for the number of ESBL genes and plasmid replicons detected. For most isolates, the number of chromosomal wgMLST genes detected in the plasmid assemblies decreased with increasing k-mer size. Based on our findings, PlasmidSPAdes is not a suitable plasmid assembly tool for short-read sequence data for ESBL-encoding plasmids of .
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http://dx.doi.org/10.1099/mgen.0.000400DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7478632PMC
July 2020

Improved diagnostic policy for respiratory tract infections essential for patient management in the emergency department.

Future Microbiol 2020 05 3;15:623-632. Epub 2020 Jun 3.

The University of Groningen, University Medical Center Groningen, Department of Medical Microbiology & Infection Prevention, Division of Clinical Virology, Groningen, The Netherlands.

Establishing an optimal diagnostic policy for patients with respiratory tract infections, at the emergency department (ED) of a university hospital in The Netherlands. Adult patients were sampled at admission, during the respiratory season (2014-2015). The FilmArray-RP was implemented at the clinical virology laboratory. Diagnostics were provided from 8 am to 10 pm, weekends included. 436/492 (89%) results were available while patients were still at the ED. Median TAT from admission to test result was 165 min (IQR: 138-214). No antibiotics were prescribed in 94/207 (45%) patients who tested positive for a virus. 185/330 (56%) hospitalized patients did not need admission with isolation measures. The value-based measure, expressed in euro-hour (€h), increased to tenfold compared with previous policy. An optimal policy is essential for patient management, by providing timely, reliable diagnostics.
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http://dx.doi.org/10.2217/fmb-2019-0119DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7426768PMC
May 2020

Prevention and Control of Multidrug-Resistant Bacteria in The Netherlands and Germany-The Impact of Healthcare Structures.

Int J Environ Res Public Health 2020 03 30;17(7). Epub 2020 Mar 30.

Department of Medical Microbiology and Infection Prevention, University of Groningen, University Medical Center Groningen, Hanzeplein 1, 9700RB Groningen, The Netherlands.

The Netherlands and Germany are neighbouring countries within the European Union but are differently affected by multidrug-resistant microorganisms (MDRO). In this narrative review, we summarize data about antibiotic use, the occurrence of MDRO and healthcare-associated infections in these two countries, as well as data about organizational and structural differences between the Dutch and German healthcare systems. These results are discussed with a focus on whether or how the organization of healthcare influences MDRO prevention. We found that from the point of view of MDRO prevention, a higher density of inpatient care, a higher number of hospitals, a longer length of stay and lower staffing ratios might facilitate MDRO dissemination in German hospitals.
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http://dx.doi.org/10.3390/ijerph17072337DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7178045PMC
March 2020

Decreasing prevalence of contamination with extended-spectrum beta-lactamase-producing Enterobacteriaceae (ESBL-E) in retail chicken meat in the Netherlands.

PLoS One 2019 31;14(12):e0226828. Epub 2019 Dec 31.

Department of Infection Control, Amphia Hospital, Breda, the Netherlands.

Retail chicken meat is a potential source of extended-spectrum beta-lactamase-producing Enterobacteriaceae (ESBL-E). In the past decade, vast national efforts were undertaken to decrease the antibiotic use in the veterinary sector, resulting in a 58% decrease in antibiotic sales in the sector between 2009 and 2014. This decrease in antibiotic use was followed by a decrease in ESBL-E prevalence in broilers. The current study investigates the prevalence of contamination with ESBL-E in retail chicken meat purchased in the Netherlands between December 2013 and August 2015. It looks at associations between the prevalence of contamination with ESBL-E and sample characteristics such as method of farming (free-range or conventional), supermarket chain of purchase and year of purchase. In the current study, 352 chicken meat samples were investigated for the presence of ESBL-E using selective culture methods. Six samples were excluded due to missing isolates or problems obtaining a good quality sequence leaving 346 samples for further analyses. Of these 346 samples, 188 (54.3%) were positive for ESBL-E, yielding 216 ESBL-E isolates (Escherichia coli (n = 204), Klebsiella pneumoniae (n = 11) and Escherichia fergusonii (n = 1)). All ESBL-E isolates were analysed using whole-genome sequencing. The prevalence of contamination with ESBL-E in retail chicken meat decreased from 68.3% in 2014 to 44.6% in 2015, absolute risk difference 23.7% (95% confidence interval (CI): 12.6% - 34.1%). The ESBL-E prevalence was lower in free-range chicken meat (36.4%) compared with conventional chicken meat (61.5%), absolute risk difference 25.2% (95% CI: 12.9% - 36.5%). The prevalence of contamination with ESBL-E varied between supermarket chains, the highest prevalence of contamination was found in supermarket chain 4 (76.5%) and the lowest in supermarket chain 1 (37.8%). Pairwise isolate comparisons using whole-genome multilocus sequence typing (wgMLST) showed that clustering of isolates occurs more frequently within supermarket chains than between supermarket chains. In conclusion, the prevalence of contamination with ESBL-E in retail chicken in the Netherlands decreased over time; nevertheless, it remains substantial and as such a potential source for ESBL-E in humans.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0226828PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6938319PMC
April 2020

Incidence, clinical implications and impact on public health of infections with Shigella spp. and entero-invasive Escherichia coli (EIEC): results of a multicenter cross-sectional study in the Netherlands during 2016-2017.

BMC Infect Dis 2019 Dec 9;19(1):1037. Epub 2019 Dec 9.

Department of Medical Microbiology and Infection Prevention, University of Groningen, University Medical Center Groningen, Groningen, the Netherlands.

Background: Shigella spp. and entero-invasive E. coli (EIEC) use the same invasive mechanism to cause diarrheal diseases. Public health regulations apply only to Shigella spp. infections, but are hampered by the lack of simple methods to distinguish them from EIEC. In the last decades, molecular methods for detecting Shigella spp. and EIEC were implemented in medical microbiological laboratories (MMLs). However, shigellosis cases identified with molecular techniques alone are not notifiable in most countries. Our study investigates the impact of EIEC versus Shigella spp. infections and molecular diagnosed shigellosis versus culture confirmed shigellosis for re-examination of the rationale for the current public health regulations.

Methods: In this multicenter cross-sectional study, fecal samples of patients suspected for gastro-enteritis, referred to 15 MMLs in the Netherlands, were screened by PCR for Shigella spp. or EIEC. Samples were cultured to discriminate between the two pathogens. We compared risk factors, symptoms, severity of disease, secondary infections and socio-economic consequences for (i) culture-confirmed Shigella spp. versus culture-confirmed EIEC cases (ii) culture positive versus PCR positive only shigellosis cases.

Results: In 2016-2017, 777 PCR positive fecal samples with patient data were included, 254 of these were culture-confirmed shigellosis cases and 32 were culture-confirmed EIEC cases. EIEC cases were more likely to report ingestion of contaminated food and were less likely to be men who have sex with men (MSM). Both pathogens were shown to cause serious disease although differences in specific symptoms were observed. Culture-negative but PCR positive cases were more likely report travel or ingestion of contaminated food and were less likely to be MSM than culture-positive cases. Culture-negative cases were more likely to suffer from multiple symptoms. No differences in degree of secondary infections were observed between Shigella spp. and EIEC, and culture-negative and culture-positive cases.

Conclusions: No convincing evidence was found to support the current guidelines that employs different measures based on species or detection method. Therefore, culture and molecular detection methods for Shigella spp. and EIEC should be considered equivalent for case definition and public health regulations regarding shigellosis. Differences were found regarding risks factors, indicating that different prevention strategies may be required.
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http://dx.doi.org/10.1186/s12879-019-4659-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6902317PMC
December 2019

Development of a reference data set for assigning and species based on next generation sequencing of the 16S-23S rRNA region.

Antimicrob Resist Infect Control 2019 15;8:178. Epub 2019 Nov 15.

2Department of Medical Microbiology, University of Groningen, University Medical Center Groningen, Groningen, The Netherlands.

Background: Many members of and genera are clinically relevant opportunistic pathogens warranting accurate and rapid identification for targeted therapy. Currently, the developed method based on next generation sequencing (NGS) of the 16S-23S rRNA region proved to be a rapid, reliable and precise approach for species identification directly from polymicrobial and challenging clinical samples. The introduction of this new method to routine diagnostics is hindered by a lack of the reference sequences for the 16S-23S rRNA region for many bacterial species. The aim of this study was to develop a careful assignment for streptococcal and enterococcal species based on NGS of the 16S-23S rRNA region.

Methods: Thirty two strains recovered from clinical samples and 19 reference strains representing 42 streptococcal species and nine enterococcal species were subjected to bacterial identification by four Sanger-based sequencing methods targeting the genes encoding (i) 16S rRNA, (ii) , (iii) and (iv) ; and NGS of the 16S-23S rRNA region.

Results: This study allowed obtainment and deposition of reference sequences of the 16S-23S rRNA region for 15 streptococcal and 3 enterococcal species followed by enrichment for 27 and 6 species, respectively, for which reference sequences were available in the databases. For , NGS of the 16S-23S rRNA region was as discriminative as Sanger sequencing of the and genes allowing for an unambiguous identification of 93% of analyzed species. For , , and genes sequencing allowed for identification of all species, while the NGS-based method did not allow for identification of only one enterococcal species. For both genera, the sequence analysis of the 16S rRNA gene was endowed with a low identification potential and was inferior to that of other tested identification methods. Moreover, in case of phylogenetically related species the sequence analysis of only the intergenic spacer region was not sufficient enough to precisely identify strains at the species level.

Conclusions: Based on the developed reference dataset, clinically relevant streptococcal and enterococcal species can now be reliably identified by 16S-23S rRNA sequences in samples. This study will be useful for introduction of a novel diagnostic tool, NGS of the 16S-23S rRNA region, which undoubtedly is an improvement for reliable culture-independent species identification directly from polymicrobially constituted clinical samples.
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http://dx.doi.org/10.1186/s13756-019-0622-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6858756PMC
July 2020

Reduced Fitness Costs of Compared to Mutated in Isogenic Colistin-Resistant KPC-3-Producing Klebsiella pneumoniae.

mSphere 2019 11 6;4(6). Epub 2019 Nov 6.

University of Groningen, University Medical Center Groningen, Department of Medical Microbiology, Groningen, Netherlands.

In the present study, we provide the results of a detailed genomic analysis and the growth characteristics of a colistin-resistant KPC-3-producing sequence type 512 (ST512) isolate (the colR-KPC3-KP isolate) with a mutated and isogenic isolates of colR-KPC3-KP with isolated from an immunocompromised patient. From 2014 to 2017, four colR-KPC3-KP isolates were detected in rectal swab samples collected from a pediatric hematology patient at the Azienda Ospedaliero-Universitaria Pisana in Pisa, Italy. Whole-genome sequencing was performed by MiSeq sequencing (Illumina). Growth experiments were performed using different concentrations of colistin. The growth lag phases both of an isolate harboring a deletion in and of clonal variants with were assessed by the use of real-time light-scattering measurements. In the first isolate (isolate 1000-Δ, recovered in September 2014), a 17-nucleotide deletion in was detected. In subsequent isolates, the gene associated with the plasmid pIncX4-AOUP was found, while was intact. Additionally, plasmid pIncQ-AOUP, harboring aminoglycoside resistance genes, was detected. The growth curves of the first three isolates were identical without colistin exposure; however, at higher concentrations of colistin, the growth curves of the isolate with a deletion in showed longer lag phases. We observed the replacement of mutated colR-KPC3-KP by isogenic isolates with multiple resistance plasmids, including carrying pIncX4, probably due to coselection under gentamicin treatment in a patient with prolonged colR-KPC3-KP carriage. The carriage of these isolates persisted in follow-up cultures. Coselection and the advantages in growth characteristics suggest that the plasmid-mediated resistance conferred by has fewer fitness costs in colR-KPC3-KP than mutations in chromosomal , contributing to the success of this highly resistant hospital-adapted epidemiological lineage. Our study shows a successful prolonged human colonization by a colistin-resistant isolate harboring An intense antibiotic therapy contributed to the maintenance of this microorganism through the acquisition of new resistance genes. The isolates carrying showed fewer fitness costs than isogenic isolates with a mutation in the chromosome. Coselection and reduced fitness costs may explain the replacement of isolates with the mutation by other isolates and the ability of the microorganism to persist despite antibiotic treatment.
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http://dx.doi.org/10.1128/mSphere.00551-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6835208PMC
November 2019

Development and Validation of a Reference Data Set for Assigning Species Based on Next-Generation Sequencing of the 16S-23S rRNA Region.

Front Cell Infect Microbiol 2019 7;9:278. Epub 2019 Aug 7.

Department of Medical Microbiology, University of Groningen, University Medical Center Groningen, Groningen, Netherlands.

Many members of the genus are clinically relevant opportunistic pathogens that warrant accurate and rapid identification for targeted therapy. The aim of this study was to develop a careful assignment scheme for staphylococcal species based on next-generation sequencing (NGS) of the 16S-23S rRNA region. All reference staphylococcal strains were identified at the species level using Sanger sequencing of the 16S rRNA, and genes and NGS of the 16S-23S rRNA region. To broaden the database, an additional 100 staphylococcal strains, including 29 species, were identified by routine diagnostic methods, 16S rRNA Sanger sequencing and NGS of the 16S-23S rRNA region. The results enabled development of reference sequences encompassing the 16S-23S rRNA region for 50 species (including one newly proposed species) and 6 subspecies of the genus. This study showed and targets were the most discriminative but NGS of the 16S-23S rRNA region was more discriminative than gene sequencing and much more discriminative than 16S rRNA gene sequencing. Almost all species could be distinguished when the max score was 99.0% or higher and the sequence similarity between the best and second best species was equal to or >0.2% (min. 9 nucleotides). This study allowed development of reference sequences for 21 staphylococcal species and enrichment for 29 species for which sequences were publicly available. We confirmed the usefulness of NGS of the 16S-23S rRNA region by identifying the whole species content in 45 clinical samples and comparing the results to those obtained using routine diagnostic methods. Based on the developed reference database, all staphylococcal species can be reliably detected based on the 16S-23S rRNA sequences in samples composed of both single species and more complex polymicrobial communities. This study will be useful for introduction of a novel diagnostic tool, which undoubtedly is an improvement for reliable species identification in polymicrobial samples. The introduction of this new method is hindered by a lack of reference sequences for the 16S-23S rRNA region for many bacterial species. The results will allow identification of all species, which are clinically relevant pathogens.
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http://dx.doi.org/10.3389/fcimb.2019.00278DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6698797PMC
July 2020

Applied shotgun metagenomics approach for the genetic characterization of dengue viruses.

J Biotechnol 2019 15;306S:100009. Epub 2019 May 15.

University of Groningen, University Medical Center Groningen, Department of Medical Microbiology and Infection Prevention, Groningen, the Netherlands. Electronic address:

Dengue virus (DENV), an arthropod-borne virus, has rapidly spread in recent years. DENV diagnosis is performed through virus serology, isolation or molecular detection, while genotyping is usually done through Sanger sequencing of the envelope gene. This study aimed to optimize the use of shotgun metagenomics and subsequent bioinformatics analysis to detect and type DENV directly from clinical samples without targeted amplification. Additionally, presence of DENV quasispecies (intra-host variation) was revealed by detecting single nucleotide variants. Viral RNA was isolated with or without DNase-I treatment from 17 DENV (1-4) positive blood samples. cDNA libraries were generated using either a combination of the NEBNext® RNA to synthesize cDNA followed by Nextera XT DNA library preparation, or the TruSeq RNA V2 (TS) library preparation kit. Libraries were sequenced using both the MiSeq and NextSeq. Bioinformatic analysis showed complete ORFs for all samples by all approaches, but longer contigs and higher sequencing depths were obtained with the TS kit. No differences were observed between MiSeq and NextSeq sequencing. Detection of multiple DENV serotypes in a single sample was feasible. Finally, results were obtained within three days with associated reagents costs between €130-170/sample. Therefore, shotgun metagenomics is suitable for identification and typing of DENV in a clinical setting.
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http://dx.doi.org/10.1016/j.btecx.2019.100009DOI Listing
May 2019

Daptomycin Resistant Clinical Strain With Novel Non-synonymous Mutations in the and Genes: A New Insight Into Daptomycin Resistance.

Front Microbiol 2018 6;9:2705. Epub 2018 Nov 6.

Department of Medical Microbiology, University Medical Center Groningen, University of Groningen, Groningen, Netherlands.

Daptomycin (DAP) resistance in is uncommon but there are increasing reports of the emergence of resistance during DAP therapy. Most clinical DAP-resistant isolates investigated carried mutations in the gene. The aim of this study was to identify mutations between a clinical pair of methicillin-susceptible (MSSA) isolates (DAP-susceptible and DAP-resistant). Additionally, the activity of genes previously associated with DAP resistance was assessed. Two MSSA isolates from patient with left-sided endocarditis were analyzed by whole genome sequencing (WGS) and reverse transcription-quantitative real-time PCR (RT-qPCR). The first isolate, DAP-susceptible, was obtained before initiation of treatment and the second isolate, DAP-resistant, was recovered after 4 weeks of DAP therapy. Comparison of complete genomes of DAP-susceptible and its DAP-resistant variant identified two non-synonymous and one synonymous mutations. The non-synonymous mutations consisted of a S829L substitution in and a T331I substitution in . The RT-qPCR experiments revealed an increased expression of , and genes in DAP-resistant variant. Strikingly, the expression of and genes was significantly downregulated by DAP. The and genes were previously associated with DAP resistance, however, none of the mutations described in this study had been previously identified and linked to DAP resistance. Moreover, we provide a new insight into the DAP action on , in which the expression of key genes in DAP resistance is decreased by the antibiotic.
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http://dx.doi.org/10.3389/fmicb.2018.02705DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6232378PMC
November 2018

Antimicrobial Resistance in Class 1 Integron-Positive Shiga Toxin-Producing Isolated from Cattle, Pigs, Food and Farm Environment.

Microorganisms 2018 Sep 28;6(4). Epub 2018 Sep 28.

Laboratorio de Inmunoquímica y Biotecnología, Facultad de Ciencias Veterinarias, UNCPBA 7000, Argentina.

The aim of this study was to investigate the presence of class 1 integrons in a collection of Shiga toxin-producing (STEC) from different origins and to characterize pheno- and genotypically the antimicrobial resistance associated to them. A collection of 649 isolates were screened for the class 1 integrase gene () by Polymerase chain reaction The variable region of class 1 integrons was amplified and sequenced. Positive strains were evaluated for the presence of antimicrobial resistance genes with microarray and for antimicrobial susceptibility by the disk diffusion method. Seven out of 649 STEC strains some to serogroups, O26, O103 and O130 isolated from cattle, chicken burger, farm environment and pigs were identified as positive for . Different arrangements of gene cassettes were detected in the variable region of class 1 integron: , and . In almost all strains, phenotypic resistance to streptomycin, tetracycline, trimethoprim/sulfamethoxazole, and sulfisoxazole was observed. Microarray analyses showed that most of the isolates carried four or more antimicrobial resistance markers and STEC strains were categorized as Multridrug-resistant. Although antimicrobials are not usually used in the treatment of STEC infections, the presence of Multridrug-resistant in isolates collected from farm and food represents a risk for animal and human health.
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http://dx.doi.org/10.3390/microorganisms6040099DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6313391PMC
September 2018

Joint Genomic and Proteomic Analysis Identifies Meta-Trait Characteristics of Virulent and Non-virulent Strains.

Front Cell Infect Microbiol 2018 6;8:313. Epub 2018 Sep 6.

Department of Analytical Biochemistry, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Krakow, Poland.

is an opportunistic pathogen of humans and warm-blooded animals and presents a growing threat in terms of multi-drug resistance. Despite numerous studies, the basis of staphylococcal virulence and switching between commensal and pathogenic phenotypes is not fully understood. Using genomics, we show here that strains exhibiting virulent (VIR) and non-virulent (NVIR) phenotypes in a chicken embryo infection model genetically fall into two separate groups, with the VIR group being much more cohesive than the NVIR group. Significantly, the genes encoding known staphylococcal virulence factors, such as clumping factors, are either found in different allelic variants in the genomes of NVIR strains (compared to VIR strains) or are inactive pseudogenes. Moreover, the pyruvate carboxylase and gamma-aminobutyrate permease genes, which were previously linked with virulence, are pseudogenized in NVIR strain ch22. Further, we use comprehensive proteomics tools to characterize strains that show opposing phenotypes in a chicken embryo virulence model. VIR strain CH21 had an elevated level of diapolycopene oxygenase involved in staphyloxanthin production (protection against free radicals) and expressed a higher level of immunoglobulin-binding protein Sbi on its surface compared to NVIR strain ch22. Furthermore, joint genomic and proteomic approaches linked the elevated production of superoxide dismutase and DNA-binding protein by NVIR strain ch22 with gene duplications.
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http://dx.doi.org/10.3389/fcimb.2018.00313DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6136393PMC
August 2019

Extended-spectrum beta-lactamase producing Enterobacteriaceae (ESBL-E) isolated from bean sprouts in the Netherlands.

PLoS One 2018 30;13(8):e0203338. Epub 2018 Aug 30.

Department of Infection Control, Amphia Hospital, Breda, the Netherlands.

Community-acquired carriage and infections due to extended-spectrum beta-lactamase producing Enterobacteriaceae (ESBL-E) are increasing worldwide, resulting in increased morbidity, mortality and healthcare costs. The origins of community-acquired ESBL-E carriage and infections remain unclear. Bean sprouts are a potential source of Enterobacteriaceae for the community, as illustrated by outbreaks of pathogenic Enterobacteriaceae in the past. The current study focuses on contamination of retail bean sprouts with ESBL-E in the Netherlands. Of 131 bean sprout samples purchased between 2013 and 2016, 25 (19%) were contaminated with ESBL-E. The detected isolates were almost exclusively Klebsiella spp. and co-resistance to other antibiotics was observed frequently. Over time there was substantial genetic diversity between isolates. On the other hand, isolates from samples closely matched in time were frequently clonally related, indicative of batch contamination. Remarkably, no Escherichia coli was found. In conclusion, bean sprouts frequently harbor ESBL-E, which is a potential source for consumers.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0203338PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6117087PMC
February 2019

European external quality assessments for identification, molecular typing and characterization of Staphylococcus aureus.

J Antimicrob Chemother 2018 10;73(10):2662-2666

Centre National de Référence des Staphylocoques, Institut des Agents Infectieux, Hospices Civils de Lyon, 103 Grande Rue de la Croix Rousse, Lyon, France.

Objectives: We present the results of two European external quality assessments (EQAs) conducted in 2014 and 2016 under the auspices of the Study Group on Staphylococci and Staphylococcal Infections of ESCMID. The objective was to assess the performance of participating centres in characterizing Staphylococcus aureus using their standard in-house phenotypic and genotypic protocols.

Methods: A total of 11 well-characterized blindly coded S. aureus (n = 9), Staphylococcus argenteus (n = 1) and Staphylococcus capitis (n = 1) strains were distributed to participants for analysis. Species identification, MIC determination, antimicrobial susceptibility testing, antimicrobial resistance and toxin gene detection and molecular typing including spa typing, SCCmec typing and MLST were performed.

Results: Thirteen laboratories from 12 European countries participated in one EQA or both EQAs. Despite considerable diversity in the methods employed, good concordance (90%-100%) with expected results was obtained. Discrepancies were observed for: (i) identification of the S. argenteus strain; (ii) phenotypic detection of low-level resistance to oxacillin in the mecC-positive strain; (iii) phenotypic detection of the inducible MLSB strain; and (iv) WGS-based detection of some resistance and toxin genes.

Conclusions: Overall, good concordance (90%-100%) with expected results was observed. In some instances, the accurate detection of resistance and toxin genes from WGS data proved problematic, highlighting the need for validated and internationally agreed-on bioinformatics pipelines before such techniques are implemented routinely by microbiology laboratories. We strongly recommend all national reference laboratories and laboratories acting as referral centres to participate in such EQA initiatives.
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http://dx.doi.org/10.1093/jac/dky260DOI Listing
October 2018

Evaluation of a Culture-Dependent Algorithm and a Molecular Algorithm for Identification of Shigella spp., Escherichia coli, and Enteroinvasive E. coli.

J Clin Microbiol 2018 10 25;56(10). Epub 2018 Sep 25.

Medical Microbiology, University of Groningen, University Medical Center Groningen, Groningen, The Netherlands.

Identification of spp., , and enteroinvasive (EIEC) is challenging because of their close relatedness. Distinction is vital, as infections with spp. are under surveillance of health authorities, in contrast to EIEC infections. In this study, a culture-dependent identification algorithm and a molecular identification algorithm were evaluated. Discrepancies between the two algorithms and original identification were assessed using whole-genome sequencing (WGS). After discrepancy analysis with the molecular algorithm, 100% of the evaluated isolates were identified in concordance with the original identification. However, the resolution for certain serotypes was lower than that of previously described methods and lower than that of the culture-dependent algorithm. Although the resolution of the culture-dependent algorithm is high, 100% of noninvasive , , and , 93% of and EIEC, and 85% of isolates were identified in concordance with the original identification. Discrepancy analysis using WGS was able to confirm one of the used algorithms in four discrepant results. However, it failed to clarify three other discrepant results, as it added yet another identification. Both proposed algorithms performed well for the identification of spp. and EIEC isolates and are applicable in low-resource settings, in contrast to previously described methods that require WGS for daily diagnostics. Evaluation of the algorithms showed that both algorithms are capable of identifying species and EIEC isolates. The molecular algorithm is more applicable in clinical diagnostics for fast and accurate screening, while the culture-dependent algorithm is more suitable for reference laboratories to identify spp. and EIEC up to the serotype level.
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http://dx.doi.org/10.1128/JCM.00510-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6156305PMC
October 2018

ESCMID postgraduate education course: regional capacity building for integration of next-generation sequencing in the clinical microlab.

Microbes Infect 2018 05 17;20(5):275-280. Epub 2018 Mar 17.

Department of Health System Management, School of Public Health, Faculty of Health Sciences, Ben-Gurion University of the Negev, Beer-Sheva, Israel; ESCMID Study Group for Genomic and Molecular Diagnostics (ESGMD), Basel, Switzerland. Electronic address:

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http://dx.doi.org/10.1016/j.micinf.2018.02.006DOI Listing
May 2018

Defining Multidrug Resistance of Gram-Negative Bacteria in the Dutch-German Border Region-Impact of National Guidelines.

Microorganisms 2018 Jan 26;6(1). Epub 2018 Jan 26.

Department of Medical Microbiology, University Medical Center Groningen (UMCG), University of Groningen, 9713 GZ Groningen, The Netherlands.

Preventing the spread of multidrug-resistant Gram-negative bacteria (MDRGNB) is a public health priority. However, the definition of MDRGNB applied for planning infection prevention measures such as barrier precautions differs depending on national guidelines. This is particularly relevant in the Dutch-German border region, where patients are transferred between healthcare facilities located in the two different countries, because clinicians and infection control personnel must understand antibiograms indicating MDRGNB from both sides of the border and using both national guidelines. This retrospective study aimed to compare antibiograms of Gram-negative bacteria and classify them using the Dutch and German national standards for MDRGNB definition. A total of 31,787 antibiograms from six Dutch and four German hospitals were classified. Overall, 73.7% were no MDRGNB according to both guidelines. According to the Dutch and German guideline, 7772/31,787 (24.5%) and 4586/31,787 (12.9%) were MDRGNB, respectively ( < 0.0001). Major divergent classifications were observed for extended-spectrum β-lactamase (ESBL) -producing , non-carbapenemase-producing carbapenem-resistant , and . The observed differences show that medical staff must carefully check previous diagnostic findings when patients are transferred across the Dutch-German border, as it cannot be assumed that MDRGNB requiring special hygiene precautions are marked in the transferred antibiograms in accordance with both national guidelines.
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http://dx.doi.org/10.3390/microorganisms6010011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5874625PMC
January 2018

Evaluation of an Accelerated Workflow for Surveillance of ESBL (CTX-M)-Producing Escherichia coli Using Amplicon-Based Next-Generation Sequencing and Automated Analysis.

Microorganisms 2018 Jan 11;6(1). Epub 2018 Jan 11.

Department of Medical Microbiology, University Medical Center Groningen, University of Groningen, 9713 GZ Groningen, The Netherlands.

Outbreak management of extended spectrum -lactamase (ESBL)-producing pathogens requires rapid and accurate diagnosis. However, conventional screening is slow and labor-intensive. The vast majority of the screened samples are negative and detection of non-outbreak-related resistant micro-organisms often complicates outbreak management. In a CTX-M-15-producing outbreak, 149 fecal samples and rectal eSwabs were collected by a cross-sectional survey in a Dutch nursing home. Samples were processed by routine diagnostic methods. Retrospectively, ESBL-producing bacteria and resistance genes were detected directly from eSwab medium by an accelerated workflow without prior enrichment cultures by an amplicon-based next-generation sequencing (NGS) method, and culture. A total of 27 (18.1%) samples were positive in either test. Sensitivity for CTX-M detection was 96.3% for the phenotypic method and 85.2% for the NGS method, and the specificity was 100% for both methods, as confirmed by micro-array. This resulted in a positive predictive value (PPV) of 100% for both methods, and a negative predictive value (NPV) of 99.2% and 96.8% for the phenotypic method and the NGS method, respectively. Time to result was four days and 14 h for the phenotypic method and the NGS method, respectively. In conclusion, the sensitivity without enrichment shows promising results for further use of amplicon-based NGS for screening during outbreaks.
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http://dx.doi.org/10.3390/microorganisms6010006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5874620PMC
January 2018

Diagnostic Evasion of Highly-Resistant Microorganisms: A Critical Factor in Nosocomial Outbreaks.

Front Microbiol 2017 3;8:2128. Epub 2017 Nov 3.

Department of Medical Microbiology, University Medical Center Groningen, University of Groningen, Groningen, Netherlands.

Highly resistant microorganisms (HRMOs) may evade screening strategies used in routine diagnostics. Bacteria that have evolved to evade diagnostic tests may have a selective advantage in the nosocomial environment. Evasion of resistance detection can result from the following mechanisms: low-level expression of resistance genes not resulting in detectable resistance, slow growing variants, mimicry of wild-type-resistance, and resistance mechanisms that are only detected if induced by antibiotic pressure. We reviewed reports on hospital outbreaks in the Netherlands over the past 5 years. Remarkably, many outbreaks including major nation-wide outbreaks were caused by microorganisms able to evade resistance detection by diagnostic screening tests. We describe various examples of diagnostic evasion by several HRMOs and discuss this in a broad and international perspective. The epidemiology of hospital-associated bacteria may strongly be affected by diagnostic screening strategies. This may result in an increasing reservoir of resistance genes in hospital populations that is unnoticed. The resistance elements may horizontally transfer to hosts with systems for high-level expression, resulting in a clinically significant resistance problem. We advise to communicate the identification of HRMOs that evade diagnostics within national and regional networks. Such signaling networks may prevent inter-hospital outbreaks, and allow collaborative development of adapted diagnostic tests.
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http://dx.doi.org/10.3389/fmicb.2017.02128DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5675861PMC
November 2017

PVL overexpression due to genomic rearrangements and mutations in the S. aureus reference strain ATCC25923.

BMC Res Notes 2017 Nov 7;10(1):576. Epub 2017 Nov 7.

Alere Technologies GmbH (Abbott Rapid Diagnostics), Jena, Germany.

Objective: ATCC25923 is a Staphylococcus aureus strain that is positive for the Panton Valentin leukocidin. It has been used for decades as reference strain. We observed that two separately maintained clones of ATCC25923 ("G477 and G478") differed grossly in the expression of this toxin. For that reason, both clones were sequenced using an Illumina MiSeq instrument. After assembling, the final sequences were analyzed and mapped to a previously published ATCC25923 sequence (GenBank CP009361) using bl2seq from the NCBI Blast2 package.

Results: The genomes of G477 and G478 size 2778,859 and 2792,213 nucleotides, respectively. Both genomes include a circular plasmid of 27,490 nucleotides. The sequence of the G477 chromosome maps nearly exactly to CP009361. G478 has a slightly larger size because of the presence of an additional transposable element tnp13k. The second copy of that tnp13k element is located in an intergenic region between the genes mazF and rsbU. The sequences of the ATCC25923 clones G477 and G478 differ mainly in the insertion of a second tnp13k element between the genes mazF and rsbU. That insertion may lead to a different transcription of that genome region resulting in upregulation of the expression of the Panton-Valentine leukocidin in the ATCC25923 clone G478.
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http://dx.doi.org/10.1186/s13104-017-2891-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5678758PMC
November 2017
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