Characteristics of the Genetic Spread of Carbapenem-Resistant Acinetobacter baumannii in a Tertiary Greek Hospital.

Acinetobacter baumannii (Ab) has increasingly been identified as a cause of hospital-acquired infections and epidemics. The rise of carbapenem-resistant Acinetobacter baumannii (CRAB) poses significant challenges in treatment. Nosocomial outbreaks linked to CRAΒ A. baumannii strains have been reported worldwide, including in Greece. This study aimed to analyze the molecular epidemiology trends of multidrug-resistant A. baumannii isolates in a tertiary hospital in Athens, Greece. A total of 43 clinical isolates of extensively drug-resistant (XDRAB), pan-drug-resistant (PDRAB), and CRAB were collected from patients suffering from blood infection, hospitalized between 2016 and 2020 at the internal medicine clinics and the ICU. A.baumannii isolates underwent testing for Ambler class B and D carbapenemases and the detection of ISAba1, and were typed, initially, using pulsed-field gel electrophoresis, and, subsequently, using sequence-based typing and multiplex PCR to determine European Clone lineages. The blaOXA-23 gene accompanied by ISAba1 was prevalent in nearly all A. baumannii isolates, except for one carrying blaOXA-58. The intrinsic blaOXA-51-like gene was found in all isolates. No Ambler class B carbapenemases (VIM, NDM) were detected. Isolates were grouped into four PF-clusters and no one-cluster spread was documented, consistent with the absence of outbreak. The study indicated that XDR/PDR-CRAB isolates predominantly produce OXA-23 carbapenemase and belong to European Clone II. Further research is needed to understand the distribution of resistant bacteria and develop effective prevention and control strategies.

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.

Emergence and persistent spread of carbapenemase-producing Klebsiella pneumoniae high-risk clones in Greek hospitals, 2013 to 2022.

BackgroundPreliminary unpublished results of the survey of carbapenem- and/or colistin-resistant Enterobacterales (CCRE survey) showed the expansion of carbapenemase-producing Klebsiella pneumoniae (CPKP) sequence type (ST) 39 in 12 of 15 participating Greek hospitals in 2019.AimWe conducted a rapid survey to determine the extent of spread of CPKP high-risk clones in Greek hospitals in 2022 and compare the distribution of circulating CPKP clones in these hospitals since 2013.MethodsWe analysed whole genome sequences and epidemiological data of 310 K. pneumoniae isolates that were carbapenem-resistant or 'susceptible, increased exposure' from Greek hospitals that participated in the European survey of carbapenemase-producing Enterobacteriaceae (EuSCAPE, 2013-2014), in the CCRE survey (2019) and in a national follow-up survey (2022) including, for the latter, an estimation of transmission events.ResultsFive K. pneumoniae STs including ST258/512 (n = 101 isolates), ST11 (n = 93), ST39 (n = 56), ST147 (n = 21) and ST323 (n = 13) accounted for more than 90% of CPKP isolates in the dataset. While ST11, ST147 and ST258/512 have been detected in participating hospitals since 2013 and 2014, KPC-2-producing ST39 and ST323 emerged in 2019 and 2022, respectively. Based on the defined genetic relatedness cut-off, 44 within-hospital transmission events were identified in the 2022 survey dataset, with 12 of 15 participating hospitals having at least one within-hospital transmission event.ConclusionThe recent emergence and rapid spread of new high-risk K. pneumoniae clones in the Greek healthcare system related to within-hospital transmission is of concern and highlights the need for molecular surveillance and enhanced infection prevention and control measures.

Antimicrobial susceptibility testing and reporting practices of public hospital microbiology laboratories in Greece, 2022: A national observational survey and call for action.

Antimicrobial resistance (AMR) in Greece is among the highest across the European Union/European Economic Area (EU/EEA), with high AMR rates even to last-line antibiotics. To better understand the clinical microbiology laboratory practices and capacities in species identification and antimicrobial susceptibility testing (AST) across public healthcare establishments in Greece, we sent a questionnaire to 98 of 128 public hospital microbiology laboratories between 1 February and 1 April 2022. Of the 73.5% (72/98) that responded, 51.4% (37/72) reported using EUCAST guidelines. Two of three laboratories used an automated instrument for species identification and AST for all laboratory samples. Broth microdilution for colistin susceptibility testing was used by 46 of the laboratories, more frequently in larger (> 400 beds) versus smaller (< 400 beds) hospitals (90.5% (19/21) vs 52.9% (27/51) respectively, p = 0.011). MALDI-TOF mass spectrometry was available in one of 10 laboratories, and more often in larger compared to smaller hospitals (p = 0.035). Although the majority of laboratories had a laboratory information system (LIS) in place, only half had the capacity to extract data directly from the LIS for the purpose of AMR surveillance; 73.6% (53/72) used restrictive antibiograms. Public microbiology laboratory AMR capacities in Greece require improvement, prioritising interventions for EUCAST guidelines implementation.

Polyclonal Endemicity of Carbapenemase-Producing Klebsiella pneumoniae in ICUs of a Greek Tertiary Care Hospital.

Carbapenemase-producing Klebsiella pneumoniae (CPKP) emerged in Greece in 2002 and became endemic thereafter. Driven by a notable variability in the phenotypic testing results for carbapenemase production in K. pneumoniae isolates from the intensive care units (ICUs) of our hospital, we performed a study to assess the molecular epidemiology of CPKP isolated between 2016 and 2019 using pulse-field gel electrophoresis (PFGE) including isolates recovered from 165 single patients. We investigated the molecular relatedness among strains recovered from rectal surveillance cultures and from respective subsequent infections due to CPKP in the same individual (48/165 cases). For the optimal interpretation of our findings, we carried out a systematic review regarding the clonality of CPKP isolated from clinical samples in ICUs in Europe. In our study, we identified 128 distinguishable pulsotypes and 17 clusters that indicated extended dissemination of CPKP within the hospital ICU setting throughout the study period. Among the clinical isolates, 122 harbored KPC genes (74%), 2 harbored KPC+NDM (1.2%), 38 harbored NDM (23%), 1 harbored NDM+OXA-48 (0.6%), 1 harbored NDM+VIM (0.6%) and 1 harbored the VIM (0.6%) gene. Multiple CPKP strains in our hospital have achieved sustained transmission. The polyclonal endemicity of CPKP presents a further threat for the selection of pathogens resistant to last-resort antimicrobial agents.

COVID-19 and Antimicrobial Resistance: Data from the Greek Electronic System for the Surveillance of Antimicrobial Resistance-WHONET-Greece (January 2018-March 2021).

Changes in hospitals' daily practice due to COVID-19 pandemic may have an impact on antimicrobial resistance (AMR). We aimed to assess this possible impact as captured by the Greek Electronic System for the Surveillance of Antimicrobial Resistance (WHONET-Greece). Routine susceptibility data of 17,837 Gram-negative and Gram-positive bacterial isolates from blood and respiratory specimens of hospitalized patients in nine COVID-19 tertiary hospitals were used in order to identify potential differences in AMR trends in the last three years, divided into two periods, January 2018-March 2020 and April 2020-March 2021. Interrupted time-series analysis was used to evaluate differences in the trends of non-susceptibility before and after the changes due to COVID-19. We found significant differences in the slope of non-susceptibility trends of Acinetobacter baumannii blood and respiratory isolates to amikacin, tigecycline and colistin; of Klebsiella pneumoniae blood and respiratory isolates to meropenem and tigecycline; and of Pseudomonas aeruginosa respiratory isolates to imipenem, meropenem and levofloxacin. Additionally, we found significant differences in the slope of non-susceptibility trends of Staphylococcus aureus isolates to oxacillin and of Enterococcus faecium isolates to glycopeptides. Assessing in this early stage, through surveillance of routine laboratory data, the way a new global threat like COVID-19 could affect an already ongoing pandemic like AMR provides useful information for prompt action.

The Rising Burden of Salmonellosis Caused by Monophasic Salmonella Typhimurium (1,4,[5],12:i:-) in Greece and New Food Vehicles.

Monophasic Salmonella typhimurium is of increasing importance worldwide. Here we present the available data regarding monophasic S. typhimurium from 2007 to 2019 in Greece, in order to assess its public health impact. Surveillance data, data on antimicrobial resistance, molecular typing by pulsed-field gel electrophoresis (PFGE), and results of the investigation of monophasic S. typhimurium outbreaks were analyzed. Overall, 403 cases were identified; 329 (81.6%) sporadic and 74 (18.4%) related to two community outbreaks in 2017. A total of 305 isolates from sporadic cases tested for antimicrobial resistance revealed resistance to ampicillin, streptomycin, sulphamethoxazole, and tetracycline (41.3%). Some 23.3% were further resistant to trimethoprim and 5.2% were also resistant to chloramphenicol. Outbreak 1 in 2017 with 37 identified cases was attributed to the consumption of raw milk from a vending machine and isolates were resistant to ampicillin, streptomycin, sulphamethoxazole, tetracycline, and trimethoprim. Outbreak 2 also with 37 cases was attributed to the consumption of pork and isolates were resistant to the five above mentioned antibiotics plus chloramphenicol. The number of human monophasic S. typhimurium isolates is low; however, since 2009, it has been among the five most frequently identified serotypes in Greece. Investigation of the outbreaks revealed that other vehicles apart from pork may be implicated in the occurrence of outbreaks.

Eight-year trends in the relative isolation frequency and antimicrobial susceptibility among bloodstream isolates from Greek hospitals: data from the Greek Electronic System for the Surveillance of Antimicrobial Resistance - WHONET-Greece, 2010 to 2017.

BackgroundAntimicrobial resistance (AMR) changes over time and continuous monitoring provides insight on trends to inform both empirical treatment and public health action.AimsTo survey trends in relative isolation frequency (RIF) and AMR among key bloodstream pathogens using data from the Greek Electronic System for the Surveillance of AMR (WHONET-Greece).MethodsThis observational study looked into routine susceptibility data of 50,488 blood culture isolates from hospitalised patients in 25 tertiary hospitals, participating in the WHONET-Greece for trends over time between January 2010 and December 2017. Only the first isolate per species from each patient was included. Hospital wards and intensive care units (ICUs) were analysed separately.ResultsDuring the study, the RIF of Acinetobacter baumannii increased in wards, as did the proportion of A. baumannii isolates, which were non-susceptibleto most antibiotics in both wards and ICUs. Coincidently, Klebsiella pneumoniae RIF declined while the respective rates of non-susceptible isolates to carbapenems and gentamicin increased. Pseudomonas aeruginosa RIF remained stable but decreasing proportions of non-susceptible isolates to all studied antibiotics, except imipenem were observed. Escherichia coli RIF increased as did the proportion of isolates non-susceptible to third-generation cephalosporins, carbapenems and fluoroquinolones. Concerning Staphylococcus aureus, a decline in the percentage of meticillin resistant isolates in ICUs was found, while the percentages of Enterococcus faecium isolates with non-susceptibility to vancomycin stayed stable.ConclusionsRecognising these trends over time is important, since the epidemiology of AMR is complex, involving different 'bug and drug' combinations. This should be taken into consideration to control AMR.

An outbreak of a possibly new Salmonella enterica subspecies enterica serovar with the antigenic formula 11:z41:e,n,z15, Greece, March to May 2016: preliminary results.

Eleven Salmonella spp. isolates with the antigenic type 11:z41:e,n,z15 - not referred to in the 9th edition of the White-Kauffman-Le Minor Scheme - were identified at the National Reference Laboratory for Salmonella in Greece. Their pulsed-field gel electrophoresis profiles were indistinguishable. No apparent epidemiological link has yet been identified; the results of a case-case study are pending.

Circulation of a multiresistant, conjugative, IncA/C plasmid within the nosocomial Providencia stuartii population in the Athens area.

The objective of the study is to report a multidrug-resistant outbreak of Providencia stuartii that occurred in inpatients in the Athens area in 2012 resulting from a very successful transmissible A/C multidrug-resistant plasmid. Thirteen multidrug-resistant P. stuartii clinical isolates from 5 hospitals were studied. Molecular typing was performed by pulsed-field gel electrophoresis. Antibiotic resistance genes and their genetic surround were detected by PCR and sequencing. Plasmid analysis included conjugation experiments using liquid cultures, sizing by S1 digestion, and incompatibility replicon typing by PCR. Isolates were grouped into 2 distinct clonal types A and B, exhibiting similarity less than 70%. Isolates of type A were recovered from patients hospitalized in 4 different hospitals with no obvious epidemiological linkage, while isolates of type B were recovered from patients treated in a single hospital. Both clonal types harbored a conjugative plasmid of 130 bp and IncA/C replicon type carrying 5 ß-lactamase genes bla(SHV-5), bla(VEB-1), bla(VIM-1), bla(OXA-10), and bla(TEM-1) and aminoglycosides resistant determinants. All ß-lactamase genes were included in stable structures as IS26, IS1999, and In-e541. The current plasmid seemed to have many common determinants with previously reported plasmids derived from P. stuartii and Proteus mirabilis clinical isolates and exhibited the ability to circulate in nosocomial bacterial populations.

Carbapenem-resistant Klebsiella pneumoniae infections in a Greek intensive care unit: Molecular characterisation and treatment challenges.

Acquisition of carbapenemase-producing Klebsiella pneumoniae (CP-Kp) strains poses a major threat to critically ill patients. The objectives of this study were to describe the epidemiology of CP-Kp isolates as well as the clinical outcome associated with the corresponding infections and to identify risk factors for mortality of intensive care unit (ICU) patients in a Greek hospital. A prospective, observational study was conducted in a nine-bed general ICU over a 2-year period (April 2010-March 2012). Imipenem-resistant K. pneumoniae isolates recovered from clinical samples of ICU patients were prospectively collected and studied for the presence of carbapenemases. Isolates were submitted to molecular typing using pulsed-field gel electrophoresis (PFGE). In total, 61 CP-Kp isolates (48 KPC-producers and 13 VIM-producers) were recovered from 58 ICU patients. The majority of KPC-producers were classified into a single PFGE type, indicating potent clonal dissemination. Among the 32 infected patients, bacteraemia was diagnosed in 16. Tigecycline+colistin was the most common combination antimicrobial regimen. Infection-attributable mortality was 43.8%. Regarding mortality risk factors, non-survivors were older (P=0.080), all of them presented with septic shock (P=0.010) and they had higher Sepsis-related Organ Failure Assessment (SOFA) scores at infection onset (P=0.004) compared with survivors. Appropriate definitive treatment and combination regimens were not associated with patient survival. In conclusion, CP-Kp infections are associated with limited treatment options and high in-hospital mortality. Effective measures for preventing dissemination of respective isolates in the hospital setting are required.

Outbreak of pan-susceptible Klebsiella pneumoniae in a neonatal intensive care unit.

We describe the outbreak of a pan-susceptible Klebsiella pneumoniae strain in a neonatal intensive care unit. A total of 7 neonates developed bacteraemia (37% attack rate), of whom 3 died (43% case fatality rate). A birth weight < 1500 g was the only statistically significant risk factor. Despite an extensive environmental investigation, the source was not identified.

An update of the evolving epidemic of blaKPC-2-carrying Klebsiella pneumoniae in Greece (2009-10).

OBJECTIVES: To follow the epidemic of KPC-2-producing Klebsiella pneumoniae in Greece. METHODS: KPC-2-producing isolates (n = 378) were collected during January 2009-April 2010 in 40 Greek hospitals. bla(KPC) and bla(VIM) were detected by PCR. Carbapenemase production was confirmed by spectrophotometry. Sequences flanking bla(KPC-2) and their plasmid carriers were studied. Isolates were typed by PFGE and multilocus sequence typing (MLST). RESULTS: All 378 isolates were bla(KPC-2) positive; 18 also carried bla(VIM-1/VIM-4). Higher isolation frequencies were observed in Athens and Crete. Isolates were classified into 13 PFGE types and 11 sequence types (STs). ST258 was predominant (n = 322), followed by ST147 (n = 20), ST383 (n = 9), ST133 (n = 6), ST274 (n = 4) and ST323 (n = 3). Of the remaining isolates, seven were distributed into five STs (11, 17, 340 and the novel 494 and 495) and seven were not typed. bla(KPC-2) could not be transferred from ST258 isolates, in contrast to isolates of ST17, ST133, ST147, ST274, ST494 and ST495. All bla(KPC-2)-encoding plasmids were of similar size (∼100 kb) and showed indistinguishable restriction fragment length polymorphism (RFLP) patterns except those from the ST340 isolates. Sequences flanking bla(KPC-2) revealed that the Tn4401a isoform was present in plasmids from all STs except ST340 containing Tn4401b. Co-production of VIM enzymes was observed in isolates of ST147, ST323 and ST383. CONCLUSIONS: Apart from the epidemic of KPC-2-producing K. pneumoniae belonging to ST258 in Greece, diffusion of bla(KPC-2) to at least 10 additional STs has taken place. Notably, strains from three of the latter STs (147, 323 and 383) were found to carry both bla(KPC-2) and bla(VIM).