Microbiological diagnostics of bloodstream infections in Europe-an ESGBIES survey.

OBJECTIVES: High-quality diagnosis of bloodstream infections (BSI) is important for successful patient management. As knowledge on current practices of microbiological BSI diagnostics is limited, this project aimed to assess its current state in European microbiological laboratories. METHODS: We performed an online questionnaire-based cross-sectional survey comprising 34 questions on practices of microbiological BSI diagnostics. The ESCMID Study Group for Bloodstream Infections, Endocarditis and Sepsis (ESGBIES) was the primary platform to engage national coordinators who recruited laboratories within their countries. RESULTS: Responses were received from 209 laboratories in 25 European countries. Although 32.5% (68/209) of laboratories only used the classical processing of positive blood cultures (BC), two-thirds applied rapid technologies. Of laboratories that provided data, 42.2% (78/185) were able to start incubating BC in automated BC incubators around-the-clock, and only 13% (25/192) had established a 24-h service to start immediate processing of positive BC. Only 4.7% (9/190) of laboratories validated and transmitted the results of identification and antimicrobial susceptibility testing (AST) of BC pathogens to clinicians 24 h/day. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry from briefly incubated sub-cultures on solid media was the most commonly used approach to rapid pathogen identification from positive BC, and direct disc diffusion was the most common rapid AST method from positive BC. CONCLUSIONS: Laboratories have started to implement novel technologies for rapid identification and AST for positive BC. However, progress is severely compromised by limited operating hours such that current practice of BC diagnostics in Europe complies only partly with the requirements for optimal BSI management.

Impact of bloodstream infections caused by carbapenem-resistant Gram-negative pathogens on ICU costs, mortality and length of stay.

BACKGROUND: The aim of this study was to estimate the impact of bloodstream infections (BSIs) caused by carbapenem-resistant Gram-negative (CRGN) pathogens on hospital costs, mortality and length of stay (LOS). METHODS: All patients hospitalized for ≥3 days in the Intensive Care Unit (ICU) of a tertiary-care general hospital from 1/1/2015 to 31/12/2017 were included in the study. A retrospective case-control study was performed in order to examine the difference in medical, pharmaceutical and operating costs, LOS and in-hospital mortality between patients with BSI caused by CRGN/without BSI (cases/controls, respectively). The statistical analysis was performed using the SPSS software (v23.0). RESULTS: A total of 419 patients (67.5% males, median age 60.0 years) were included in the analysis (142 cases/277 controls); 10 patients with non-CRGN BSIs were excluded. Overall mortality was 33.7% (49.3/25.6% in cases/controls). The median LOS and total cost were 30.0 vs. 12.0 days and 20 359.1 vs. 8,509.3 €, respectively, between patients with/without CRGN BSIs. After adjusting for baseline demographics, underlying disease severity and patients' specialties, CRGN BSIs remained a significant factor in mortality (odds ratio 2.9; 95% confidence interval 1.8-4.8; p <0.001). Additionally, CRGN BSIs seem to result in significantly prolonged LOS and extra cost per infected patient (p <0.001). CONCLUSIONS: ICU patients with CRGN BSI are at increased risk for mortality and prolonged hospitalization and incur higher costs, imposing a heavy burden on healthcare system. Infection control strategies, considering also the cost-efficacy of interventions, are crucial in order to control the expansion of CRGN infections.

Changes in antimicrobial resistance of clinical isolates of Acinetobacter baumannii group isolated in Greece, 2010-2015.

In recent years, hospitals in southeastern Europe have faced dramatically high rates of antibiotic-resistant Acinetobacter baumannii. We analysed the evolution of resistance among clinical isolates of A. baumannii group obtained from nine tertiary hospitals throughout Greece over 6 years (2010-2015). Identification and antimicrobial susceptibility testing were performed using Vitek 2 or Microscan walkaway automated systems. Between 2010 and 2015, resistance to ampicillin/sulbactam increased from 46.2 to 88.2 % (P=0.021), resistance to gentamicin increased from 69.3 to 86.4 % (P=0.014) and resistance to tobramycin increased from 59.8 to 76.8 % (P=0.011). Imipenem resistance rates were consistently very high, ranging from 90.3 % in 2010 to 94.5 % in 2015 (P=0.198), while meropenem resistance rates increased from 82.6 % in 2010 to 94.8 % in 2015 (P=0.006). Resistance rates to trimethoprim/sulfamethoxazole showed a remarkable decreasing trend, declining from 90.2 % in 2010 to 69.1 % in 2015 (P=0.035). These evolutions render the treatment of A. baumannii infections particularly challenging and underline the need for enhanced infection control measures.

Infection control interventions affected by resource shortages: impact on the incidence of bacteremias caused by carbapenem-resistant pathogens.

We evaluated an infection control (IC) program influenced by personnel and material resource shortages on the incidence of bloodstream infections (BSI) due to carbapenem-resistant Klebsiella pneumoniae (CRKP), Acinetobacter baumannii (CRAB), and Pseudomonas aeruginosa (CRPA) in an endemic region. Between January 2010 and December 2015, all BSI episodes caused by CRKP, CRAB, and CRPA were recorded. An IC bundle was implemented in January 2012. We evaluated the effect of the interventions on BSI rates between the pre-intervention (2010-2011) and intervention (2012-2013) periods, using an interrupted time-series model. From 2014, when interventions were still applied, BSI incidence was gradually increased. For this reason, we evaluated with a linear mixed effects model several factors possibly contributing to this increase for the years 2012-2015, which was considered as the intervention/follow-up period. During the study period, 351 patients with BSI were recorded, with a total of 538 episodes; the majority (83.6%) occurred in the intensive care unit (ICU). The BSI incidence rate per year during 2010-2015 for ICU patients was 21.03/19.63/17.32/14.45/22.85/25.02 per 1000 patient-days, respectively, with the reduction in BSI levels after the start of intervention marginal (p = 0.054). During the follow-up period (2014-2015), the most influential factors for the increased BSI incidence were the reduced participation in educational courses and compliance with hand hygiene. The implementation of IC interventions reduced the BSI incidence rates, particularly for ICU patients. However, factors possibly related to the restrictions of human and material resources apparently contributed to the observed expansion of BSI in our endemic setting.

First-year results of an antibiotic stewardship program in a Greek tertiary care hospital.

The aim of this study was to investigate the effect of the implementation of an antibiotic stewardship program (ASP) on antibiotic consumption in our 428-bed hospital. The Infection Control Committee implemented an ASP beginning in January 2016, aiming to reduce inappropriate antibiotic use through improved prescribing practices. The ASP included both pre-authorization and prospective audit and feedback strategies. We collected pharmacy and hospital data for the years 2015 (pre-intervention) and 2016 (post-intervention). Consumption data were expressed as daily defined doses (DDDs) per 100 patient-days (PD) and the significance of the differences between 2015 and 2016 was assessed by paired t-test. Antibiotic resistance rates for the most important hospital pathogens were monitored for 2015-2016. The ASP effectively reduced consumption of most antimicrobials; total antibiotic use decreased by 16.7% (from 104.3 in 2015 to 86.9 DDDs/100 patient-days in 2016, p < 0.001) owing to reduction of 19.1% for non-restricted and 13.8% for restricted antibiotics. Important restricted antimicrobials, such as colistin, carbapenems, quinolones and tigecycline showed significantly decreased usage post-intervention. Significant changes in the resistance rates were not observed, except a decreasing trend for colistin and tigecycline (Acinetobacter baumannii and Klebsiella pneumoniae) and also vancomycin (enterococci). The ASP was successful in terms of reducing the antibiotic consumption for the first year of its implementation. Interestingly, antimicrobials requiring pre-authorization exhibited a lower reduction than other antibiotics. Potential effects of the ASP in reducing resistance rates remain to be shown.

Effect of ceftriaxone on the outcome of murine pyelonephritis caused by extended-spectrum-ß-lactamase-producing Escherichia coli.

Urinary tract infections (UTIs) due to extended-spectrum-ß-lactamase (ESBL)-producing Enterobacteriaceae in children are becoming more frequent, and they are commonly treated initially with a second- or third-generation cephalosporin. We developed a murine model of ascending UTI caused by ESBL-producing Escherichia coli. Using this model, we investigated the renal bacterial burden, interleukin-6 (IL-6) expression, and histopathological alterations caused by ESBL- and non-ESBL-producing bacteria after 1, 2, or 6 days with or without ceftriaxone therapy. The renal bacterial burden, IL-6 concentration, and histological inflammatory lesions were not significantly different between mice infected with ESBL- and non-ESBL-producing bacteria without treatment at any of the time points examined. Following ceftriaxone administration, the bacterial burden was eliminated in the kidneys of mice infected with ESBL- and non-ESBL-producing bacteria on the 6th postinfection day. The histological analysis demonstrated that among mice treated with ceftriaxone, those infected with ESBL-producing bacteria had more profound renal alterations than those infected with non-ESBL-producing bacteria on the 6th day (P < 0.001). In comparison, microbiological outcomes did not differ significantly between mice infected with ESBL- and non-ESBL-producing bacteria at any of the time points examined. The effectiveness of ceftriaxone in mice with UTIs due to ESBL-producing E. coli may have therapeutic implications; it is, however, hampered by limited activity on the histopathological lesions, a finding that needs further investigation.

A large waterborne gastroenteritis outbreak in central Greece, March 2012: challenges for the investigation and management.

In March 2012, there was an unusual increase of gastroenteritis cases in a district with 37,264 inhabitants in central Greece. It was estimated that more than 3600 people developed symptoms. A 1:1 case-control study showed that consumption of tap water was a risk factor for acquiring infection [odds ratio (OR) 2.18, 95% confidence interval (CI) 1.11-4.28]. Descriptive data, low gastroenteritis incidence in adjacent areas with different water supply systems, and water-quality data further supported the hypothesis of a waterborne outbreak. Thirty-eight stool samples were positive for rotavirus. Bacterial indicators of recent faecal contamination were detected in samples from the water source and ice cubes from a local production enterprise. Molecular epidemiology of rotavirus strains, apart from the common strain, G3[P8], identified the unusual G/P combination G2P[8]. Water sanitation measures contributed to the control of the outbreak. This outbreak demonstrated the need for the cooperation of laboratories with different expertise and the importance of early notification of waterborne gastroenteritis outbreaks.

Accumulation of carbapenem resistance mechanisms in VIM-2-producing Pseudomonas aeruginosa under selective pressure.

Pseudomonas aeruginosa has the potential to achieve resistance to carbapenems via the acquisition of carbapenemase-encoding genes, the downregulation of the OprD porin, the overexpression of efflux systems and the overproduction of cephalosporinases. One hundred and fifty carbapenem-non-susceptible isolates from 2008 to 2010 were screened for carbapenemase production, OprD porin loss, efflux pumps overexpression and inducible AmpC beta-lactamase production. For comparison reasons, the presence of the same mechanisms was also assessed in a previous collection of 30 carbapenem-non-susceptible P. aeruginosa isolated between 2003 and 2005. Results showed the accumulation of various resistance mechanisms among VIM-2 producers isolated between 2008 and 2010 with a parallel considerable increase in imipenem MIC90 and the geometric mean of the MIC values of imipenem and meropenem between the two study groups. The accumulation of carbapenem resistance mechanisms highlights the potential of this formidable pathogen for evolutionary success under antibiotic selective pressure.

Restless legs syndrome in hemodialysis patients: an epidemiologic survey in Greece.

BACKGROUND: Restless legs syndrome (RLS) is a sensorimotor disorder characterized by an uncontrolled need to move extremities accompanied by unpleasant sensations, which frequently leads to sleep disturbances. In hemodialysis (HD) patients, the previously reported RLS prevalence varied enormously, between 6% and 60%. In our study, we investigated the RLS prevalence in HD patients for the first time in Greece. METHODS: A continuous sample of HD patients was studied between January and September of 2010 in six dialysis units in Greece. RLS diagnosis was based on the essential clinical criteria of the International RLS Study Group (IRLSSG). The standardized incidence ratio (SIR) for RLS in HD patients was calculated in comparison to data from a recent survey of the general population in Greece. RESULTS: In our study of 579 HD patients in Greece (236 women; mean age, 65±13years), the prevalence of RLS was elevated in comparison to the general population (26.6% vs 3.9%), with an SIR of 5.4 (95% confidence interval [CI], 4.6-6.3). In the fully adjusted model, the risk for RLS in HD patients was reduced in older age (odds ratio [OR], 0.98 [95% CI, 0.96-0.99]) and increased in women (OR, 1.60 [95% CI, 1.05-2.43]) in cases with elevated levels of ß2 microglobulin (OR, 1.15 [95% CI, 1.01-1.32]) and intact parathormone (iPTH) (OR, 1.30 [95% CI, 1.08-1.56]). CONCLUSION: A high RLS prevalence was recorded in a large HD population in Greece, clearly suggesting the need for enhanced awareness of RLS in nephrology. The RLS risk was increased in women and in younger HD patients as well as in those with elevated ß2 microglobulin and iPTH levels.

Shiga toxin-producing Escherichia coli in Central Greece: prevalence and virulence genes of O157:H7 and non-O157 in animal feces, vegetables, and humans.

In Greece, Shiga toxin-producing Escherichia coli (STEC) have only been sporadically reported. The objective of this study was to estimate the prevalence of STEC and Escherichia coli O157:H7 in farm animals, vegetables, and humans in Greece. A total number of 1,010 fecal samples were collected from farm animals (sheep, goats, cattle, chickens, pigs), 667 diarrheal samples from humans, and 60 from vegetables, which were cultured in specific media for STEC isolates. Enzyme-linked immunosorbent assay (ELISA) was used to detect toxin-producing colonies, which, subsequently, were subjected to a multiplex polymerase chain reaction (PCR) for stx1, stx2, eae, rfbE O157, and fliC h7 genes. Eighty isolates (7.9 %) from animal samples were found to produce Shiga toxin by ELISA, while by PCR, O157 STEC isolates were detected from 8 (0.8 %) samples and non-O157 STEC isolates from 43 (4.2 %) samples. STEC isolates were recovered mainly from sheep and goats, rarely from cattle, and not from pigs and chickens, suggesting that small ruminants constitute a potential risk for human infections. However, only three human specimens (0.4 %) were positive for the detection of Shiga toxins and all were PCR-negative. Similarly, all 60 vegetable samples were negative for toxin production and for toxin genes, but three samples (two roman rockets and one spinach) were positive by PCR for rfbE O157 and fliC h7 genes. These findings indicate that sheep, goats, cattle, and leafy vegetables can be a reservoir of STEC and Escherichia coli O157:H7 isolates in Greece, which are still rarely detected among humans.

Diversity of vanA-type vancomycin-resistant Enterococcus faecium isolated from broilers, poultry slaughterers and hospitalized humans in Greece.

OBJECTIVES: This study investigated the prevalence of vancomycin-resistant enterococci (VRE) in the broiler production environment after the avoparcin ban and their epidemiological relationship with human clinical VRE from the same geographical regions in Greece. METHODS: Caecal contents from broilers (n = 500) from eight livestock farms and faecal samples from poultry slaughterers (n = 50), all collected in two slaughterhouses during 2005-08, were analysed for species and vancomycin resistance gene identification using multiplex PCR. Sixty-three human clinical vancomycin-resistant Enterococcus faecium (VREF) isolates, obtained during 2006-09, were also examined. Discriminant analysis (DA) was used to establish the relationship of antimicrobial resistance profiles (ARPs) among broiler, poultry slaughterer and human clinical VREF. PFGE was conducted to study the genetic relatedness among VREF from the different sources. RESULTS: A total of 120 VRE were recovered from 113 (22.6%) broiler samples. VREF carrying the vanA gene were predominant, being recovered from 72 (14.4%) samples from five (62.5%) broiler farms. Concerning poultry slaughterers, VREF were recovered from 10 (20%) samples. Susceptibility testing revealed that broiler VREF were consistently resistant to tetracycline, whereas 93.7% of clinical VREF were resistant to ampicillin. Furthermore, 92.1% of clinical VREF compared with 54.4% of broiler VREF were multiresistant (resistant to at least five antimicrobial classes). DA classified broiler and human clinical VREF into their corresponding source with high classification rates (100% and 85.7%, respectively), while the classification rate of poultry slaughterer VREF was relatively low (50%), with 40% of them classified closely to broiler VREF. PFGE patterns were clearly related to the source of the VREF, with broiler isolates being clustered distinctly from all human isolates. CONCLUSIONS: A remarkable persistence of VREF was observed in the broiler production environment even >10 years after the avoparcin ban. Human and broiler VREF belonged to clearly unrelated populations, strongly indicating no clonal spread of VREF among the different sources, even between broilers and poultry slaughterers, despite them sharing common ARPs, as also supported by DA.

Predictors of mortality in patients with bloodstream infections caused by KPC-producing Klebsiella pneumoniae and impact of appropriate antimicrobial treatment.

Bloodstream infections (BSIs) caused by Klebsiella pneumoniae carbapenemases (KPC)-producing K. pneumoniae (KPC-KP) are associated with high mortality rates. We investigated outcomes, risk factors for mortality and impact of appropriate antimicrobial treatment in patients with BSIs caused by molecularly confirmed KPC-KP. All consecutive patients with KPC-KP BSIs between May 2008 and May 2010 were included in the study and followed-up until their discharge or death. Potential risk factors for infection mortality were examined by a case-control study. Case-patients were those who died from the BSI and control-patients those who survived. Appropriate antimicrobial therapy was defined as treatment with in vitro active antimicrobials for at least 48 h. A total of 53 patients were identified. Overall mortality was 52.8% and infection mortality was 34%. Appropriate antimicrobial therapy was administered to 35 patients; mortality due to infection occurred in 20%. All 20 patients that received combination schemes had favourable infection outcome; in contrast, seven of 15 patients given appropriate monotherapy died (p 0.001). In univariate analysis, risk factors for mortality were age (p <0.001), APACHE II score at admission and infection onset (p <0.001) and severe sepsis (p <0.001), while appropriate antimicrobial treatment (p 0.003), combinations of active antimicrobials (p 0.001), catheter-related bacteraemia (p 0.04), prior surgery (p 0.014) and other therapeutic interventions (p 0.015) were significantly associated with survival. Independent predictors of mortality were age, APACHE II score at infection onset and inappropriate antimicrobial treatment. Among them, appropriate treatment is the only modifiable independent predictor of infection outcome.

Identification of the plasmid-encoded qacA efflux pump gene in meticillin-resistant Staphylococcus aureus (MRSA) strain HPV107, a representative of the MRSA Iberian clone.

Meticillin-resistant Staphylococcus aureus (MRSA) is a major nosocomial bacterium for which prevention and control measures consist mainly of the application of biocides with antiseptic and disinfectant activity. In this study, we demonstrated the presence of the plasmid-located efflux pump gene qacA in MRSA strain HPV107, a clinical isolate representative of the MRSA Iberian clone. The existence of efflux activity in strain HPV107 due to the QacA pump was also established and this QacA efflux activity was linked with a phenotype of reduced susceptibility towards several biocide compounds. No association could be made with antibiotic resistance. This work emphasises the potential of QacA pump activity in the maintenance and dissemination of important MRSA strains in the hospital setting and, increasingly, in the community.

Trends in antimicrobial resistance of clinical isolates of Enterococcus faecalis and Enterococcus faecium in Greece between 2002 and 2007.

We analysed trends in antimicrobial resistance of Enterococcus faecalis (N=1498) and E. faecium (N=625) recovered from clinical infections during 2002-2007 in a Greek tertiary care hospital. Molecular assays were used to confirm speciation and genotype of vancomycin-resistant enterococci (VRE). The incidence of infections per 1000 admissions caused by E. faecalis and E. faecium increased during the study period (chi(2) for trend=25.5 and 13.3, respectively; P<0.0001). Resistance to ampicillin, high level gentamicin and streptomycin, vancomycin, teicoplanin and linezolid was found in E. faecalis/E. faecium at rates of 1.3/82.4%, 45.6/51.2%, 48.9/69.1%, 0.5/9.6%, 0.1/8.2% and 0.3/1.6%, respectively. The vanA gene was identified in 79.1% of the VRE isolates, with vanB found in the remaining 20.1%. Analysis of antimicrobial resistance trends showed consistently high rates of ampicillin resistance among E. faecium isolates. For both enterococcal species, high level resistance to gentamicin and streptomycin were noted to have increased significantly (P<0.0001). Regardless of these alarming trends, strains exhibiting resistance to oxazolidinones seem to be only sporadic in our region and a trend toward increasing resistance rates to glycopeptides was not detected.

The effect of estrogens on plasma ghrelin concentrations in women.

BACKGROUND: Data regarding the possible effects of estrogen on ghrelin secretion in humans are limited and contradictory. AIM: To investigate the effect of estradiol (E2) on ghrelin levels in normal pre- and post-menopausal women. SUBJECTS AND METHODS: A total of 21 women divided into 3 groups, i.e.13 normally cycling women (no.=7, group 1 and no.=6, group 2) and 8 post-menopausal women (group 3). Women of group 1 received increasing doses of E2 through skin patches from cycle days 3 to 5. Women of group 2, underwent total abdominal hysterectomy plus bilateral salpingo-oophorectomy (TAH+BSO) on cycle day 3. Women of group 3 received po increasing doses of E2 valerate for 15 days. Acylated ghrelin and E2 were measured in all blood samples. RESULTS: In group 1, plasma ghrelin levels did not show any significant changes for the week following cycle day 3. In group 2, ghrelin levels were similar before and after TAH+BSO and remained stable during the first 7 post-operative days. In group 3, no significant changes in plasma ghrelin levels were seen during the 15 days of E2 administration. CONCLUSIONS: The present study demonstrates for the first time that ghrelin values were not affected either by exogenous short-term estrogen administration to pre- and post-menopausal women or following ovariectomy in pre-menopausal women. It is suggested that ovarian hormones are not involved in the regulation of ghrelin secretion in women.

Acquired carbapenemases in Gram-negative bacterial pathogens: detection and surveillance issues.

Acquired carbapenemases are emerging resistance determinants in Gram-negative pathogens, including Enterobacteriaceae, Pseudomonas aeruginosa and other Gram-negative non-fermenters. A consistent number of acquired carbapenemases have been identified during the past few years, belonging to either molecular class B (metallo-beta-lactamases) or molecular classes A and D (serine carbapenemases), and genes encoding these enzymes are associated with mobile genetic elements that allow their rapid dissemination in the clinical setting. Therefore, detection and surveillance of carbapenemase-producing organisms have become matters of major importance for the selection of appropriate therapeutic schemes and the implementation of infection control measures. As carbapenemase production cannot be simply inferred from the resistance profile, criteria must be established for which isolates should be suspected and screened for carbapenemase production, and for which tests (phenotypic and/or genotypic) should be adopted for confirmation of the resistance mechanism. Moreover, strategies should be devised for surveillance of carbapenemase producers in order to enable the implementation of effective surveillance programmes. The above issues are addressed in this article, as a follow-up to an expert meeting on acquired carbapenemases that was recently organized by the ESCMID Study Group for Antibiotic Resistance Surveillance.

The acute effect of smoking in healthy and asthmatic smokers.

BACKGROUND: Acute exposure to cigarette smoke is related to airway and systemic inflammation and oxidative stress. Little is known about the acute effect of cigarette smoking in smoking asthmatics. The aim of this study was to evaluate the acute effect of smoking in airway and systemic inflammation and oxidative stress in normal smokers and patients with properly treated well-controlled persistent asthma. MATERIALS AND METHODS: Ten normal smokers and 10 smokers with moderate persistent asthma controlled with LABA and ICS were recruited. Subjects refrained from smoking for at least 12 h prior to their inclusion. We compared the effects of smoking of two cigarettes on airway obstruction, airway inflammation and oxidative stress [by measuring fraction of exhaled nitric oxide (FeNO), plus pH and 8-isoprostane in exhaled breath condensate (EBC)] before and 30, 90 and 180 min after smoking. Furthermore, we evaluated systemic oxidative stress, C-reactive protein (CRP) and serum amyloid A (SAA) and urine leukotriene E(4) (LTE(4)) before and 180 min after smoking. RESULTS: No differences were observed in EBC pH and 8-isoprostane, FeNO and systemic oxidative stress between the groups at baseline. In asthmatics, EBC pH decreased 30 min and EBC 8-isoprostane increased 90 min after smoking (P = 0.039 and P = 0.029 respectively), which was not evident in smoking controls. Serum oxidative stress increased only in asthmatic smokers at 180 min (P = 0.001). No differences were observed in SAA, CRP and urine LTE(4) levels before and after smoking. CONCLUSION: Acute smoking has more deleterious effects in well-controlled properly treated asthmatic smokers compared with matched normal smokers.

Clusters of imipenem-resistant Acinetobacter baumannii clones producing different carbapenemases in an intensive care unit.

During a 2-year period (April 2005-March 2007), 31 intensive care unit (ICU) patients in a Greek hospital were infected or colonised with imipenem-resistant isolates of Acinetobacter baumannii. Twelve patients died, with imipenem-resistant A. baumannii infection contributing to the death of seven patients. The 31 representative A. baumannii isolates were multidrug-resistant and clustered in four distinct clones, each of which contained different carbapenemase genes: clone I was predominant and contained bla(VIM-1), bla(OXA-58) and the intrinsic bla(OXA-66) gene; clone II contained bla(VIM-4), bla(OXA-58) and the intrinsic bla(OXA-69) gene; clone III contained bla(OXA-58) and the intrinsic bla(OXA-69) gene; and clone IV contained only the intrinsic bla(OXA-66) gene. ISAba1 was not associated with the intrinsic bla(OXA-51-like) alleles, whereas ISAba3 was found upstream and downstream of bla(OXA-58) in isolates of clone I, and upstream of bla(OXA-58) in isolates of clone III, but was not detected in isolates of clone II. PCR, curing and hybridisation experiments indicated that the bla(VIM) alleles were chromosomally located, whereas the bla(OXA-58) alleles were plasmid-located. This study provides the first description of the clonal spread of multidrug-resistant A. baumannii isolates carrying bla(VIM-1) and bla(VIM-4) metallo-beta-lactamase genes, and revealed that distinct carbapenem-resistant A. baumannii clusters bearing different carbapenemase genes may emerge and cause severe infections, even in a well-defined regional hospital setting.

Use of real-time PCR to detect human papillomavirus-16 viral loads in vaginal and urine self-sampled specimens.

Increasing the accuracy of self-sampling methods to detect oncogenic human papillomavirus (HPV) infection would contribute to the wider application of these approaches. In this study, 120 women were tested for HPV-16 by conventional and quantitative real-time PCR (QRT-PCR) in cervical and self-sampled vaginal and urine specimens. QRT-PCR had a higher detection rate, and the HPV viral load in all three sampling sites correlated with the severity of disease, as determined by histology. The vaginal and urine viral loads correlated with HPV-16 positivity according to both conventional and QRT-PCR, and were proportional to the cervical viral load.