Candida auris in Greek healthcare facilities: Active surveillance results on first cases and outbreaks from eleven hospitals within Attica region.

BACKGROUND: Candida auris was sporadically detected in Greece until 2019. Thereupon, there has been an increase in isolations among inpatients of healthcare facilities. AIM: We aim to report active surveillance data on MALDI-TOF confirmed Candida auris cases and outbreaks, from November 2019 to September 2021. METHODS: A retrospective study on hospital-based Candida auris data, over a 23-month period was conducted, involving 11 hospitals within Attica region. Antifungal susceptibility testing and genotyping were conducted. Case mortality and fatality rates were calculated and p-values less than 0.05 were considered statistically significant. Infection control measures were enforced and enhanced. RESULTS: Twenty cases with invasive infection and 25 colonized were identified (median age: 72 years), all admitted to hospitals for reasons other than fungal infections. Median hospitalisation time until diagnosis was 26 days. Common risk factors among cases were the presence of indwelling devices (91.1 %), concurrent bacterial infections during hospitalisation (60.0 %), multiple antimicrobial drug treatment courses prior to hospitalisation (57.8 %), and admission in the ICU (44.4 %). Overall mortality rate was 53 %, after a median of 41.5 hospitalisation days. Resistance to fluconazole and amphotericin B was identified in 100 % and 3 % of tested clinical isolates, respectively. All isolates belonged to South Asian clade I. Outbreaks were identified in six hospitals, while remaining hospitals detected sporadic C. auris cases. CONCLUSION: Candida auris has proven its ability to rapidly spread and persist among inpatients and environment of healthcare facilities. Surveillance focused on the presence of risk factors and local epidemiology, and implementation of strict infection control measures remain the most useful interventions.

A Community Waterborne Salmonella Bovismorbificans Outbreak in Greece.

BACKGROUND: In August 2022, the Hellenic National Public Health Organisation was notified about a gastroenteritis outbreak in town A in Southern Greece. Investigations aimed to identify the source and implement control measures. METHODS: Case definition categories were used in a 1:3 case-control study. Cases and controls were interviewed about various exposures. Cases' stool samples were cultured on agar plates and characterised by serotyping, antimicrobial susceptibility testing and Pulse Field Gel Electrophoresis (PFGE). Environmental investigations included tap water sampling for microbiological and chemical analysis in town A and inspection of the water supply system. RESULTS: We identified 33 cases (median age: 17 years). Tap water consumption was the only significant risk factor for gastroenteritis (OR = 5.46, 95% CI = 1.02-53.95). Salmonella (S.) Bovismorbificans isolated from eight stool and one tap water samples had identical PFGE profiles. No resistant isolates were identified. Residual chlorine levels were lower than the acceptable limits before and during the outbreak. We advised consumption of bottled water and adherence to strict hand hygiene rules until tap water was declared suitable for drinking. CONCLUSIONS: Epidemiological and molecular data revealed a waterborne S. Bovismorbificans outbreak in town A. We recommend local water safety authorities to ensure that residual chlorine levels comply with the legislation towards water safety planning, to mitigate risks.

The role and importance of epidemiology in transfusion medicine.

Transfusion therapy is an indispensable form of treatment, and an important element of the public health system. Due to its origin, blood's clinical use is associated with various risks that may cause adverse reactions and events. Progress in quality and safety of blood components has eliminated numerous risks, especially those of infectious origin. However, some risks cannot be predicted, while others cannot always be prevented. Globalisation and climate change constantly favour the spread of infectious agents. Against this, epidemiology plays a central role in ensuring the safety of transfusion treatment, by continuous surveillance and timely identification of risks, and in the development of routine and additional tests as measures for risk mitigation. As a quantitative discipline based on research methods, epidemiology is a method of reasoning; it relies on the generation and testing of hypotheses; it utilises other scientific resources, particularly in the field of blood donation and blood transfusion, thus having many applications. The main focus falls on transfusion-transmissible infections, and on environmental or occupational diseases, injuries, disabilities and death causes at large. The practice of epidemiology relies on a systematic approach and measurement of disease frequencies. Surveillance is a key element, involving continuously gathering, analysing, and evaluating data regarding diseases, morbidity and mortality, and disseminating the conclusions of the analyses to relevant competent authorities; in this way, action is taken for disease prevention and control. Surveillance systems also provide an important tool for risk assessment, a method to assess and characterise the critical parameters in the functionality of equipment, systems or processes of using scientific data in order to estimate the magnitude of any health effect that derives from decisions of policy makers. Epidemiological surveillance, particularly for the incidence of adverse reactions and adverse events associated with blood transfusion at the national and international levels, has demonstrated the importance of multidisciplinary cooperation between blood and public health services.

Investigation of an international water polo tournament in Czechia as a potential source for early introduction of the SARS-CoV-2 Omicron variant into Belgium, Switzerland and Germany, November 2021.

BackgroundThe earliest recognised infections by the SARS-CoV-2 Omicron variant (Pango lineage B.1.1.529) in Belgium and Switzerland suggested a connection to an international water polo tournament, held 12-14 November 2021 in Brno, Czechia.AimTo study the arrival and subsequent spread of the Omicron variant in Belgium and Switzerland, and understand the overall importance of this international sporting event on the number of infections in the two countries.MethodsWe performed intensive forward and backward contact tracing in both countries, supplemented by phylogenetic investigations using virus sequences of the suspected infection chain archived in public databases.ResultsThrough contact tracing, we identified two and one infected athletes of the Belgian and Swiss water polo teams, respectively, and subsequently also three athletes from Germany. In Belgium and Switzerland, four and three secondary infections, and three and one confirmed tertiary infections were identified. Phylogenetic investigation demonstrated that this sporting event played a role as the source of infection, but without a direct link with infections from South Africa and not as a superspreading event; the virus was found to already be circulating at that time in the countries involved.ConclusionThe SARS-CoV-2 Omicron variant started to circulate in Europe several weeks before its identification in South Africa on 24 November 2021. Accordingly, it can be assumed that travel restrictions are usually implemented too late to prevent the spread of newly detected SARS-CoV-2 variants to other regions. Phylogenetic analysis may modify the perception of an apparently clear result of intensive contact tracing.

High prevalence of clonally related ST182 NDM-1-producing Enterobacter cloacae complex clinical isolates in Greece.

NDM-type metallo-ß-lactamase (MBL)-producing Enterobacterales remain uncommon in the European region, especially among species other than Klebsiella pneumoniae and Escherichia coli. The aim of this study was to describe epidemiological and molecular characteristics of a widespread NDM-1-producing Enterobacter cloacae complex outbreak in Greece. Over a 6-year period (March 2016-March 2022), a retrospective study was conducted in a tertiary care Greek hospital. Ninety single-patient carbapenem-non-susceptible E. cloacae complex clinical isolates were recovered consecutively. The isolates were subjected to further investigation, including antimicrobial susceptibility testing and combined disc tests for carbapenemase production, polymerase chain reaction and sequencing for resistance genes, molecular fingerprinting by pulsed-field gel electrophoresis (PFGE), plasmid profiling, replicon typing, conjugation experiments, genotyping by multi-locus sequence typing (MLST), whole-genome sequencing and phylogenetic analysis. Phenotypic and molecular testing confirmed the presence of blaNDM-1 in 47 (52.2%) of the E. cloacae complex isolates. MLST analysis clustered all but four of the NDM-1 producers into a single MLST sequence type (ST182), whereas single isolates belonged to different sequence types (ST190, ST269, ST443 and ST743). PFGE analysis revealed that ST182 isolates were clustered into a single clonal type, with three subtypes, which differed from the clonal types detected among the remaining carbapenem non-susceptible E. cloacae complex isolates identified during the study period. All ST182 blaNDM-1-carrying isolates also harboured the blaACT-16 AmpC gene, while the blaESBL, blaOXA-1 and blaTEM-1 genes were detected in most cases. In all clonal isolates, the blaNDM-1 gene was located on an IncA/C-type plasmid, and flanked upstream by an ISAba125 element and downstream by bleMBL. Conjugation experiments failed to produce carbapenem-resistant transconjugants, indicating a low dynamic for horizontal gene transfer. Application of enforced infection control measures led to the absence of new NDM-positive cases for periods of time during the survey. This study represents the largest clonal outbreak of NDM-producing E. cloacae complex in Europe.

Clonal outbreak caused by VIM-4-producing Proteus mirabilis in a Greek tertiary-care hospital.

Carbapenem-resistant Enterobacterales have become a major public-health issue worldwide. Here we report an outbreak caused by a clonal multidrug-resistant Proteus mirabilis strain producing VIM-4 metallo-ß-lactamase (MBL) and TEM-2 ß-lactamase in a Greek tertiary-care hospital. From July 2015 to February 2016, 27 imipenem-resistant P. mirabilis isolates were recovered from 14 patients hospitalised in two intensive care units (ICUs) and the internal medicine department in AHEPA University Hospital, Thessaloniki. The isolates were either susceptible or resistant to meropenem and were resistant to all remaining ß-lactams except aztreonam. Phenotypic and molecular analysis revealed that all of the isolates harboured a blaVIM-4 MBL gene. Resistome analysis of a representative isolate showed the presence of an IncQ1 plasmid harbouring the blaVIM-4 carbapenemase and blaTEM-2 ß-lactamase genes among resistance genes coding for resistance to ß-lactams, aminoglycosides, trimethoprim, sulfonamides and lincosamides. Genotyping by pulsed-field electrophoresis (PFGE) revealed that the isolates were epidemiologically related. After recovery of the index carbapenemase-producing P. mirabilis clinical isolate, infection control measures were intensified in the affected departments. Rectal sampling for carbapenem-resistant bacteria was initiated on a weekly basis among patients admitted to the general ICU. The outbreak was finally interrupted 6 months later in February 2016. This is the first documentation of the blaVIM-4 MBL gene in P. mirabilis as well as the first hospital outbreak caused by a MBL-producing P. mirabilis strain.

Emergence of NDM-1-producing Klebsiella pneumoniae in Greece: evidence of a widespread clonal outbreak.

OBJECTIVES: NDM-producing Enterobacteriaceae clinical isolates remain uncommon in the European region. We describe the emergence and broad dissemination of one successful NDM-1-producing Klebsiella pneumoniae clone in Greek hospitals. METHODS: During a 4 year survey (January 2013-December 2016), 480 single-patient carbapenem non-susceptible K. pneumoniae isolates, phenotypically MBL positive, were consecutively recovered in eight Greek hospitals from different locations and subjected to further investigation. Antimicrobial susceptibility testing, combined-disc test, identification of resistance genes by PCR and sequencing, molecular fingerprinting by PFGE, plasmid profiling, replicon typing, conjugation experiments and MLST were performed. RESULTS: Molecular analysis confirmed the presence of the blaNDM-1 gene in 341 (71%) K. pneumoniae isolates. A substantially increasing trend of NDM-1-producing K. pneumoniae was noticed during the survey (R2 = 0.9724). Most blaNDM-1-carrying isolates contained blaCTX-M-15, blaOXA-1, blaOXA-2 and blaTEM-1 genes. PFGE analysis clustered NDM-1 producers into five distinct clonal types, with five distinct STs related to each PFGE clone. The predominant ST11 PFGE clonal type was detected in all eight participating hospitals, despite adherence to the national infection control programme; it was identical to that observed in the original NDM-1 outbreak in Greece in 2011, as well as in a less-extensive NDM-1 outbreak in Bulgaria in 2015. The remaining four ST clonal types (ST15, ST70, ST258 and ST1883) were sporadically detected. blaNDM-1 was located in IncFII-type plasmids in all five clonal types. CONCLUSIONS: This study gives evidence of possibly the largest NDM-1-producing K. pneumoniae outbreak in Europe; it may also reinforce the hypothesis of an NDM-1 clone circulating in the Balkans.

Polyclonal predominance of concurrently producing OXA-23 and OXA-58 carbapenem-resistant Acinetobacter baumannii strains in a pediatric intensive care unit.

We report a predominance (64.7%) of polyclonal carbapenem-resistant Acinetobacter baumannii (CRAB) strains concurrently producing OXA-23 and OXA-58 carbapenemases in a pediatric intensive care unit in an endemic area. This is the first report of emergence of such double-OXA CRAB strains in a single unit worldwide.

Molecular Epidemiology of Endemic Carbapenem-Resistant Gram-Negative Bacteria in an Intensive Care Unit.

The molecular epidemiology of endemic carbapenem-resistant Klebsiella pneumoniae (CRKP), carbapenem-resistant Pseudomonas aeruginosa (CRPA), and carbapenem-resistant Acinetobacter baumannii (CRAB) in an intensive care unit located in an endemic area with high rates of resistance was investigated. A CRPA strain producing VIM and KPC concurrently was detected for the first time in an endemic area. CRKP strains producing K. pneumoniae carbapenemase predominated and were mainly assigned to the "hyperepidemic Greek clone." Predominant OXA-23-like producing CRAB strains were assigned to multiple pulsotypes.

Hospital outbreak due to a Klebsiella pneumoniae ST147 clonal strain co-producing KPC-2 and VIM-1 carbapenemases in a tertiary teaching hospital in Northern Greece.

Here we present the characteristics of a large outbreak caused by a clonal Klebsiella pneumoniae strain producing both KPC-2 and VIM-1 carbapenemases in a tertiary teaching hospital. Between January 2013 and January 2015, 45 carbapenem-resistant K. pneumoniae isolates that gave a positive modified Hodge test and were phenotypically suspected of metallo-ß-lactamase (MBL) and K. pneumoniae carbapenemase (KPC) co-production were recovered from 25 patients hospitalised in AHEPA University Hospital (Thessaloniki, Greece). All of the patients were hospitalised in the three intensive care units of the hospital and 17 (68%) of them developed bloodstream infections; the overall mortality of the patients involved in the outbreak was 48% (12/25). Molecular testing verified that all 45 K. pneumoniae isolates co-harboured blaKPC-2 and blaVIM-1 genes and were associated with OmpK35 deficiency and OmpK36 porin loss. The blaTEM-1 gene was also present in 18 isolates. Pulsed-field gel electrophoresis (PFGE) clustered all of the isolates into a single clonal type, and multilocus sequence typing (MLST) assigned them to the emerging high-risk ST147 clonal lineage. Following recognition of the outbreak, infection control measures were implemented in the affected areas. The outbreak continued for ca. 2 years and since then only sporadic cases of K. pneumoniae harbouring both carbapenemases have been detected.

NDM-1 Hazard in the Balkan States: Evidence of the First Outbreak of NDM-1-Producing Klebsiella pneumoniae in Bulgaria.

New Delhi MBL (NDM) carbapenemase-producing Klebsiella pneumoniae has become one of the most concerning multidrug-resistant pathogens. The Balkan counties are considered a reservoir for the spread of such strains based on several reports documenting NDM infections after hospitalization in this region. Nevertheless, NDM-producing K. pneumoniae have been only occasionally documented from Balkans. The current study documents the first polyclonal outbreak caused by NDM-1-producing K. pneumoniae in Bulgaria. From July 2015 to April 2016, all 25 single-patient carbapenem-nonsusceptible K. pneumoniae isolates were collected. Phenotypic and molecular screening revealed that 17 produced NDM-1 carbapenemase. All NDM-1 producers harbored blaCTX-M-15, blaCMY-4, blaTEM-1, and blaOXA-2; five also harbored blaOXA-1. In all cases, blaNDM-1 was flanked upstream by ISAba125 element and downstream by bleMBL. Pulsed-field gel electrophoresis (PFGE) clustered NDM-1-positive isolates into four distinct clonal types, A to D. MLST assigned isolates of the dominant clonal type A (n = 14) to sequence type (ST) 11, while isolates of clonal types B, C, and D to ST16, ST15, and ST391, respectively. Of interest, ST11 isolates belonged to the same PFGE type as those of the recently described NDM-1 ST11 clonal outbreak in Greece. Traveling abroad or overseas hospitalization was not reported in any case, suggesting most likely intra- and interhospital dissemination. The study presents the first polyclonal outbreak of NDM-producing K. pneumoniae in the Balkans and underlines the need for larger epidemiological studies in the region to illustrate commonalities in the transmission of NDM clones and possible sources in the community.

Cluster-distinguishing genotypic and phenotypic diversity of carbapenem-resistant Gram-negative bacteria in solid-organ transplantation patients: a comparative study.

Purpose. Solid-organ transplant recipients may display high rates of colonization and/or infection by multidrug-resistant bacteria. We analysed and compared the phenotypic and genotypic diversity of carbapenem-resistant (CR) strains of Klebsiella pneumoniae, Pseudomonas aeruginosa and Acinetobacter baumannii isolated from patients in the Solid Organ Transplantation department of our hospital.Methodology. Between March 2012 and August 2013, 56 CR strains from various biological fluids underwent antimicrobial susceptibility testing with VITEK 2, molecular analysis by PCR amplification and genotypic analysis with pulsed-field gel electrophoresis (PFGE). They were clustered according to antimicrobial drug susceptibility and genotypic profiles. Diversity analyses were performed by calculating Simpson's diversity index and applying computed rarefaction curves.Results/Key findings. Among K. pneumoniae, KP-producers predominated (57.1 %). VIM and OXA-23 carbapenemases prevailed among P. aeruginosa and A. baumannii (89.4 and 88.9 %, respectively). KPC-producing K. pneumoniae and OXA-23 A. baumannii were assigned in single PFGE pulsotypes. VIM-producing P. aeruginosa generated multiple pulsotypes. CR K. pneumoniae strains displayed phenotypic diversity in tigecycline, colistin (CS), amikacin (AMK), gentamicin (GEN) and co-trimoxazole (SXT) (16 clusters); P. aeruginosa displayed phenotypic diversity in cefepime (FEP), ceftazidime, aztreonam, piperacillin, piperacillin-tazobactam, AMK, GEN and CS (9 clusters); and A. baumannii displayed phenotypic diversity in AMK, GEN, SXT, FEP, tobramycin and rifampicin (8 clusters). The Simpson diversity indices for the interpretative phenotype and PFGE analysis were 0.89 and 0.6, respectively, for K. pneumoniae strains (P<0.001); 0.77 and 0.6 for P. aeruginosa (P=0.22); and 0.86 and 0.19 for A. baumannii (P=0.004).Conclusion. The presence of different antimicrobial susceptibility profiles does not preclude the possibility that two CR K. pneumoniae or A. baumannii isolates are clonally related.

Colistin Resistance in KPC-2- and SHV-5-Producing Klebsiella pneumoniae Clinical Isolates in Bulgaria.

BACKGROUND/AIMS: Colistin resistance is increasingly recognized among carbapenemase-producing Klebsiella pneumoniae isolates in several European regions. The current study documents the appearance of colistin resistance among KPC-2 and SHV-5-produning K. pneumoniae strains in Bulgaria. METHODS: Four colistin-resistant K. pneumoniae isolates were recovered from 2 patients hospitalized in the anesthesiology and resuscitation clinic of a tertiary care university hospital in Sofia, Bulgaria. Microbial identification and antimicrobial susceptibility testing was performed by Vitek 2 (Biomerieux, France). ß-Lactamase genes were amplified using a panel of primers for detection of all MBL-types, KPCs, plasmid-mediated AmpCs in single PCR reactions, OXA-type carbapenemases, extended-spectrum ß-lactamases (ESBLs) and TEM enzymes. The colistin-resistant mcr-1 gene was also investigated using previously described primers and conditions. Pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) were used to investigate clonality. RESULTS: The 4 K. pneumoniae isolates exhibited colistin MICs >16 mg/L and showed multidrug-resistant phenotypes, remaining intermediately susceptible only to gentamicin. They were clustered into a single PFGE clonal type and MLST assigned them to sequence type 258. All isolates possessed KPC-2 carbapenemase and SHV-5 ESBL. They were negative for the plasmid-mediated colistin-resistant mcr-1 gene, possibly implying an intrinsic mechanism of resistance. CONCLUSIONS: Although colistin use in Bulgaria only started moderately during 2014, the findings of the current study notify the appearance of colistin resistance among carbapenemase-producing Klebsiella species in another European region.

Epidemiological surveillance of multidrug-resistant gram-negative bacteria in a solid organ transplantation department.

BACKGROUND: We assessed the impact of intensified infection control measures (ICM) on colonization and infection caused by carbapenem-resistant (CR) Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter baumannii in a solid organ transplantation (SOT) department. METHODS: A quasi-experimental methodology was followed. The study was divided into three periods: pre-intervention, intervention with implementation of an ICM bundle including active surveillance program (ASP) and gradually enhanced measures, and post-ASP without ASP. The bundle included active surveillance cultures, contact precautions, hand hygiene, education of health care workers (HCWs), monitoring of compliance, and environmental cleaning. Incidence of colonization and infection caused by CR gram-negative bacteria was recorded. Molecular analysis of CR bacteria was performed for a certain period. RESULTS: During the intervention, incidence of colonization reduced from 19% to 9% (P<.001). The compliance of HCWs with contact precautions and hand hygiene also improved. Monthly incidence of infections caused by these CR bacteria increased from 2.8 to 6.9/1000 bed-days (P<.001). However, this increase did not have such a strong trend after the intervention. Most K. pneumoniae isolates, the commonest pathogen, carried the blaKPC gene. Colonization and infection rates by CR K. pneumoniae, P. aeruginosa, and A. baumannii were high among SOT recipients. CONCLUSION: In settings where CR gram-negative bacteria are endemic, colonization and infection rates by these bacteria are high among SOT recipients. Implementation of enhanced ICM in all related units of a hospital, although challenging, reduces colonization rates by CR gram-negative bacteria.

In Vitro Bactericidal Activity of Trimethoprim-Sulfamethoxazole Alone and in Combination with Colistin against Carbapenem-Resistant Acinetobacter baumannii Clinical Isolates.

Trimethoprim-sulfamethoxazole alone and combined with colistin was tested in vitro against six carbapenem-resistant Acinetobacter baumannii (CRAB) clinical strains. After 24 h, at achievable serum concentrations, trimethoprim-sulfamethoxazole effectively killed all strains, while colistin killed only one strain. Trimethoprim-sulfamethoxazole plus colistin rapidly killed all strains after 6 h and for up to 24 h. Trimethoprim-sulfamethoxazole, one of the few remaining antimicrobials that still has a degree of activity, particularly combined with colistin, might represent an effective therapy for severe CRAB infections.

Absence of human rhinovirus and respiratory syncytial virus from bronchoalveolar lavage and bronchial biopsies of selected patients with stable chronic obstructive pulmonary disease.

Previous studies have reported very different rates of human rhinovirus (HRV) and respiratory syncytial virus (RSV) genome detection in nasal and sputum samples, but not in bronchoalveolar lavage (BAL) and bronchial biopsy samples. Our study aimed to investigate the presence of HRV and RSV in the lungs of 31 consecutive patients with stable COPD (11 GOLD stage I, 11 II, and 9 III) and 22 control subjects (12 current or past smokers, and 10 non-smokers), who underwent diagnostic (e.g., lung cancer) and/or therapeutic (e.g., hemoptysis) fibreoptic bronchoscopy in a university hospital in Athens, Greece. Viral RNA of HRV and RSV were not detected in any of the samples of COPD patients or control subjects after being processed with real-time PCR.

Emergence of OXA-162 Carbapenemase- and DHA-1 AmpC Cephalosporinase-Producing Sequence Type 11 Klebsiella pneumoniae Causing Community-Onset Infection in Greece.

OXA-48-like carbapenemases have only recently emerged in Europe. OXA-162 is a rare OXA-48 variant usually coexpressed with extended-spectrum ß-lactamases. Here, we report the identification of the first OXA-162 carbapenemase-producing Klebsiella pneumoniae isolates, which coexpressed an AmpC cephalosporinase (DHA-1), retrieved from a patient in Greece. They belonged to a single sequence type (ST11) and caused the first documented community-onset urinary tract infections attributable to an OXA-48-like-producing Enterobacteriaceae strain.