Clinical Added Value of SARS-CoV-2 Antigen Detection in Blood Samples.

This study evaluated the performances of immunoassays (LFIA and ELISA) designed for SARS-CoV-2 Antigen (Ag)-detection in nasopharyngeal (NP) and serum samples in comparison to RT-PCR. NP samples from patients with respiratory symptoms (183 RT-PCR-positive and 74 RT-PCR-negative samples) were collected from March to April and November to December 2020. Seroconversion and antigen dynamics were assessed by symptom onset and day of RT-PCR diagnosis. Serum samples from 87 COVID-19 patients were used to investigate the added value of Ag quantification, at diagnosis and during follow-up. The sensitivity of COVID-VIRO-LFIA on samples with Ct ≤ 33, considered as the contagious threshold, was 86% on NPs (CI 95%: 79-90.5) and 76% on serum samples (CI 95%: 59.4-88), with a specificity of 100%. Serum N-Ag was detected during active infection as early as day two from symptom onset, with a diagnostic sensitivity of 81.5%. Within one week of symptom onset, diagnostic sensitivity and specificity reached 90.9% (95% CI, 85.1%-94.6%) and 98.3% (95% CI, 91.1%-99.9%), respectively. Serum N-Ag concentration closely correlated with disease severity. Longitudinal analysis revealed the simultaneous increase of antibodies and decrease of N-Ag. Sensitivities of COVID-VIRO-LFIA and COV-QUANTO-ELISA tests on NP and serum samples were close to 80%. They are suitable COVID-19-laboratory diagnostic tests, particularly when blood samples are available, thus reducing the requirement for NP sampling, and subsequent PCR analysis. ELISA titers may help to identify patients at risk of poor outcomes.

Outbreak of CTX-M-15 Extended-Spectrum ß-Lactamase-Producing Klebsiella pneumoniae ST394 in a French Intensive Care Unit Dedicated to COVID-19.

Infections caused by extended-spectrum ß-lactamase-producing Klebsiella pneumoniae (ESBL-KP) are constantly rising worldwide and are often reported as causative agent of outbreaks in intensive care units (ICUs). During the first wave of the COVID-19 pandemic, bacterial cross-transmission was thought unlikely to occur due to the reinforcement of hygiene measures and prevention control. However, we report here an ESBL-producing K. pneumoniae (ST394) isolate responsible for a nosocomial outbreak in an ICU dedicated to COVID-19 patients.

Usefulness of Xpert® Carba-R on enrichment broth for the early detection of carbapenemase-producing Enterobacterales.

Early detection of patients colonized with carbapenemase-producing Enterobacterales is crucial to limit their spread. Molecular biology tests are rapid but might miss low-level carriage. Culture-based methods using enrichment are cheap and detect low-level carriage but cause delay. Two clinical cases are reported, which demonstrated that molecular biology (Xpert® Carb-R) on enriched broth cumulates advantages of both strategies. In both cases, this strategy enabled early detection of patients colonized at low-level with carbapenemase-producing Enterobacterales because it saved 24 hours in detection time.

Uncovering the novel Enterobacter cloacae complex species responsible for septic shock deaths in newborns: a cohort study.

BACKGROUND: Enterobacter cloacae complex contains nosocomial pathogens responsible for infection outbreaks. Identification at the species level within the E cloacae complex remains difficult. Using whole genome sequencing, our aim was to decipher the transmission routes that led to the death of six of seven neonates who had bacteraemia caused by E cloacae complex isolates in a neonatal intensive care unit (NICU) over a 13 month period. METHODS: In this cohort study, E cloacae complex isolates were taken from 186 newborns in an NICU: 14 were clinical samples (eg, blood cultures), 728 rectal swab samples, and 38 environmental samples (20 from siphons, 18 from incubators, and one from a mattress). The samples were analysed to decipher the possible role of manual cross transmission or environmental source in an outbreak of fatal septic shocks related to E cloacae complex bacteraemia. After the replacement of the incubators suspected to be the reservoir of some outbreak-related isolates on Feb 1, 2018, E cloacae complex strains were screened again for 10 months (503 rectal swab samples from 163 newborns). The 71 E cloacae complex isolates recovered from screening, clinical, and environmental samples across both study periods were compared by whole genome sequencing. The pathogenicity of E cloacae complex isolates responsible for fatal septic shocks was assessed using a Galleria mellonella in-vivo model. FINDINGS: From Dec 9, 2016, to Jan 31, 2018, 249 (34%) of 728 rectal swab samples were positive for E cloacae complex, with 66 (35%) of 186 newborns colonised. E cloacae complex were also recovered from four (20%) of 20 siphons and 11 (61%) of 18 incubators. During this period, whole genome sequencing identified that the isolates responsible for the six fatal septic shocks were all Enterobacter bugandensis. A G mellonella infection model showed a higher virulence of E bugandensis. Genomic analysis highlighted the role of incubators as a long-term reservoir and source of cross contamination, leading to their replacement on Feb 1, 2018. Following incubator replacement, a 10-month follow-up investigation identified a physiological gut colonisation with polyclonal E cloacae complex in 52 (34%) of 163 neonates within a median of 9 days (5-14), but no E cloacae complex-related septic shocks. INTERPRETATION: Despite around 30% of neonates being physiologically colonised with E cloacae complex, fatal sepsis was systematically linked with bacteraemia caused by E bugandensis. Our findings highlight the need for accurate identification methods to detect the hypervirulent species within the E cloacae complex recovered in neonates. FUNDING: Assistance Publique-Hôpitaux de Paris.

Quality indicators for blood culture: 1 year of monitoring with BacT/Alert Virtuo at a French hospital.

Introduction. Blood culture (BC) remains the gold standard for the diagnosis of bloodstream infection. Clinical microbiology laboratories must ensure the quality of their BC process from receipt to definitive results.Aim. In this study, we followed the evolution of different quality indicators for BCs over the first year of implementation of the BacT/Alert Virtuo system in a French hospital.Methodology. In our laboratory, we instituted regular monitoring of several quality indicators to track (i) delays in sample registration, (ii) delays in loading BC bottles in our incubating system (BacT/Alert Virtuo) after registration, (iii) the volume of blood in bottles and (iv) the contamination rates.Results. For 53 892 BC bottles loaded in the BacT/Alert Virtuo from 23 January to 31 December 2019, the delays in sample registration, loading and unloading were respectively 3.5 h±0.016, 44 min±0.209 and 5.8 h±0.0727. Intriguingly, the automated process performed by the BacT/Alert Virtuo system to check the blood volume in bottles was only performed for 60 % of the loaded bottles. Among these, 30 % contained the recommended volume of blood (between 7 and 13 ml). Finally, the contamination rate was found to be 27.2 % for samples at our institution.Conclusions. The delays in sample registration, loading and unloading were found to be acceptable, even though they could be improved by ensuring a continuous service during the night duty period. Furthermore, the percentage of volumes measured is insufficient and must be improved and the majority of bottles do not contain the recommended blood volume.

Evaluating 10 Commercially Available SARS-CoV-2 Rapid Serological Tests by Use of the STARD (Standards for Reporting of Diagnostic Accuracy Studies) Method.

Numerous severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) rapid serological tests have been developed, but their accuracy has usually been assessed using very few samples, and rigorous comparisons between these tests are scarce. In this study, we evaluated and compared 10 commercially available SARS-CoV-2 rapid serological tests using the STARD (Standards for Reporting of Diagnostic Accuracy Studies) methodology. Two hundred fifty serum samples from 159 PCR-confirmed SARS-CoV-2 patients (collected 0 to 32 days after the onset of symptoms) were tested with rapid serological tests. Control serum samples (n = 254) were retrieved from pre-coronavirus disease (COVID) periods from patients with other coronavirus infections (n = 11), positivity for rheumatoid factors (n = 3), IgG/IgM hyperglobulinemia (n = 9), malaria (n = 5), or no documented viral infection (n = 226). All samples were tested using rapid lateral flow immunoassays (LFIAs) from 10 manufacturers. Only four tests achieved ≥98% specificity, with the specificities ranging from 75.7% to 99.2%. The sensitivities varied by the day of sample collection after the onset of symptoms, from 31.7% to 55.4% (days 0 to 9), 65.9% to 92.9% (days 10 to 14), and 81.0% to 95.2% (>14 days). Only three of the tests evaluated met French health authorities' thresholds for SARS-CoV-2 serological tests (≥90% sensitivity and ≥98% specificity). Overall, the performances varied greatly between tests, with only one-third meeting acceptable specificity and sensitivity thresholds. Knowledge of the analytical performances of these tests will allow clinicians and, most importantly, laboratorians to use them with more confidence; could help determine the general population's immunological status; and may help diagnose some patients with false-negative real-time reverse transcription-PCR (RT-PCR) results.

Carbapenemase -producing Pseudomonas aeruginosa isolates from Turkey: first report of P. aeruginosa high-risk clones with VIM-5- and IMP-7-type carbapenemases in a tertiary hospital.

We investigated the presence of carbapenemases in carbapenem-resistant Pseudomonas aeruginosa isolates, which were collected over a 14-month period in a Turkish hospital, with in-depth molecular characterization of carbapenemase-producing isolates. Among 45 study isolates, 2 isolates were identified as carbapenemase producers by both Carba NP and Carbapenem Inactivation Method tests, and only 1 of them gave a positive result in polymerase chain reaction tests for a carbapenemase gene (blaVIM). Whole genome sequencing of the 2 isolates revealed the presence of blaVIM-5 gene in an ST308 isolate, while the other one expressed IMP-7 in an ST357 isolate; both STs are considered high-risk clones. The 2 carbapenemase-producing isolates were multidrug resistant, as they harbored other resistance determinants, including a variant of the recently described plasmid-encoded fluoroquinolone resistance determinant crpP gene, crpP-2. We report for the first time P. aeruginosa high-risk clones carrying VIM-5- and IMP-7-type carbapenemases with multiple resistance determinants in Turkey.

A Lateral Flow Immunoassay for the Rapid Identification of CTX-M-Producing Enterobacterales from Culture Plates and Positive Blood Cultures.

We have developed a lateral flow immunoassay (LFIA), named NG-Test CTX-M MULTI (NG-Test), to detect group 1, 2, 8, 9, 25 CTX-M producers from agar plates and from positive blood cultures in less than 15 min. The NG-Test was validated retrospectively on 113 well-characterized enterobacterial isolates, prospectively on 102 consecutively isolated ESBL-producers from the Bicêtre hospital and on 100 consecutive blood cultures positive with a gram-negative bacilli (GNB). The NG-Test was able to detect all CTX-M producers grown on the different agar plates used in clinical microbiology laboratories. No false positive nor negative results were observed. Among the 102 consecutive ESBL isolates, three hyper mucous isolates showed an incorrect migration leading to invalid results (no control band). Using an adapted protocol, the results could be validated. The NG-Test detected 99/102 ESBLs as being CTX-Ms. Three SHV producers were not detected. Among the 100 positive blood cultures with GNB tested 10/11 ESBL-producers were detected (8 CTX-M-15, 2 CTX-M-27). One SHV-2-producing-E. cloacae was missed. The NG-Test CTX-M MULTI showed 100% sensitivity and specificity with isolates cultured on agar plates and was able to detect 98% of the ESBL-producers identified in our clinical setting either from colonies or from positive blood cultures.

Rapid Determination of SARS-CoV-2 antibodies using a bedside, point-of-Care, serological test.

Background: Several serological tests for SARS-CoV-2 have been developed or use, but most have only been validated on few samples, and none provide medical practitioners with an easy-to-use, self-contained, bedside test with high accuracy. Material and methods: Two-hundred fifty-six sera from 101 patients hospitalized with SARS-CoV-2 infection (positive RT-PCR) and 50 control sera were tested for IgM/IgG using the NG-Test IgM-IgG COVID all-in-one assay. The seroconversion dynamic was assessed by symptom onset and day of RT-PCR diagnosis. Results: Among the SARS-CoV-2 infected patients, positive IgG and/or IgM result was observed for 67.3% of patients (68/101), including 17 (16.8%) already positive at the day of RT-PCR, and 51 (50.5%) with observable seroconversion, and 32.7% (33/101) remained negative as subsequent sampling was not possible (patient discharge or death). The sensitivity increased with the delay between onset of symptoms and sampling, going from 29.1%, 78.2% and 86.5% for the time periods of 0-9-, 10-14- and >14-days after the onset of symptoms, respectively. Cumulative sensitivity, specificity, Positive Predictive Value and Negative Predictive Value were 97.0%, 100%, 100% and 96.2%, respectively 15-days after the onset of symptoms. No difference in seroconversion delay was observed regardless of whether patients received ventilation. Conclusions: The NG-test is a bedside serological assay that could serve as a complementary source of diagnostic information to RT-PCR and chest imaging. It may also be useful to monitor immunological status of medical and non-medical workers during the ongoing pandemic, and the general population after social distancing measures have eased.

Staphylococcus capitis isolated from bloodstream infections: a nationwide 3-month survey in 38 neonatal intensive care units.

To increase the knowledge about S. capitis in the neonatal setting, we conducted a nationwide 3-month survey in 38 neonatal intensive care units (NICUs) covering 56.6% of French NICU beds. We demonstrated 14.2% of S. capitis BSI (S.capBSI) among nosocomial BSIs. S.capBSI incidence rate was 0.59 per 1000 patient-days. A total of 55.0% of the S.capBSIs were late onset catheter-related BSIs. The S. capitis strains infected preterm babies (median gestational age 26 weeks, median birth weight 855 g). They were resistant to methicillin and aminoglycosides and belonged to the NRCS-A clone. Evolution was favorable in all but one case, following vancomycin treatment.

A single Proteus mirabilis lineage from human and animal sources: a hidden reservoir of OXA-23 or OXA-58 carbapenemases in Enterobacterales.

In Enterobacterales, the most common carbapenemases are Ambler's class A (KPC-like), class B (NDM-, VIM- or IMP-like) or class D (OXA-48-like) enzymes. This study describes the characterization of twenty-four OXA-23 or OXA-58 producing-Proteus mirabilis isolates recovered from human and veterinary samples from France and Belgium. Twenty-two P. mirabilis isolates producing either OXA-23 (n = 21) or OXA-58 (n = 1), collected between 2013 and 2018, as well as 2 reference strains isolated in 1996 and 2015 were fully sequenced. Phylogenetic analysis revealed that 22 of the 24 isolates, including the isolate from 1996, belonged to a single lineage that has disseminated in humans and animals over a long period of time. The blaOXA-23 gene was located on the chromosome and was part of a composite transposon, Tn6703, bracketed by two copies of IS15∆II. Sequencing using Pacbio long read technology of OXA-23-producing P. mirabilis VAC allowed the assembly of a 55.5-kb structure encompassing the blaOXA-23 gene in that isolate. By contrast to the blaOXA-23 genes, the blaOXA-58 gene of P. mirabilis CNR20130297 was identified on a 6-kb plasmid. The acquisition of the blaOXA-58 gene on this plasmid involved XerC-XerD recombinases. Our results suggest that a major clone of OXA-23-producing P. mirabilis is circulating in France and Belgium since 1996.

Successful use of culture and enrichment for the detection of OXA-181-producing Escherichia coli from rectal swab samples falsely categorized as negative by Xpert® Carba-R.

An OXA-181 producing Escherichia coli isolate that went recurrently undetected directly from rectal swab sample using Xpert® Carba-R, was successfully detected using ertapenem supplemented broth enrichment followed by culture-based method. Our data suggest that implementation of culture-based methods plus enrichment might be crucial for the efficient screening of patients considered to be at "high-risk" of colonization with carbapenemase producers and who are colonized at low level.

Evaluation of the BD MAX Check-Points CPO Assay for the Detection of Carbapenemase Producers Directly from Rectal Swabs.

A novel real-time multiplex PCR assay, BD MAX Check-Points CPO, was evaluated to detect carbapenemase-producing organisms in clinical settings on the BD MAX system. A total of 175 well-characterized isolates (including 123 carbapenemase producers) and 128 rectal swab specimens (including 83 positives) of patients considered at high risk for carriage of carbapenemase producers were included. Bacterial suspensions were used to spike true-negative rectal swabs to mimic a clinical sample. Sample (50 µL), containing either the spiked or the patient's sample, was processed. The BD MAX Check-Points CPO assay detected carbapenemases KPC, VIM/IMP, NDM, and OXA-48-like producers with a high sensitivity and specificity of 97.1% and 98.8%, respectively. Rare variants of the IMP type (IMP-11, IMP-13, and IMP-14) and one rare and distantly related OXA-48 variant (OXA-535) remained undetected. With patients' rectal swabs, sensitivity and specificity were 92.8% and 97.8%, respectively. Failure of detection was due to weak inoculum. The time to result was short: approximately. 2.5 hours for 12 samples (including extraction and PCR). The automated sample-in results-out platform is an efficient, quick, and easy-to-use tool for the detection of the main five carbapenemases. The lack of distinction between producers of VIM and IMP may be limiting in countries where these enzymes are widespread, as in Asia, but not in France, where IMP producers are extremely rare.

Aztreonam plus Clavulanate, Tazobactam, or Avibactam for Treatment of Infections Caused by Metallo-ß-Lactamase-Producing Gram-Negative Bacteria.

Metallo-ß-lactamase (MBL)-producing Gram-negative bacteria are often extremely resistant, leading to a real therapeutic dead end. Here, we evaluated the in vitro and in vivo efficacy of aztreonam in combination with ceftazidime-avibactam, ceftolozane-tazobactam, or amoxicillin-clavulanate for the treatment of infections caused by MBL-producing Enterobacteriaceae, MBL-producing Pseudomonas aeruginosa, and extremely drug-resistant Stenotrophomonas maltophilia First, we report two clinical cases, namely, a urinary tract infection caused by an NDM-5-producing Escherichia coli isolate and a pulmonary infection caused by a S. maltophilia isolate efficiently treated with the association of aztreonam-ceftazidime-avibactam and aztreonam-amoxicillin-clavulanate, respectively. Then, a total of 50 MBL-producing Enterobacteriaceae isolates, 3 MBL-producing P. aeruginosa isolates, and 5 extremely drug-resistant S. maltophilia isolates were used to test aztreonam susceptibility in combination with ceftolozane-tazobactam, ceftazidime-avibactam, or amoxicillin-clavulanate. The Etest strip superposition method was used to determine the MICs of the aztreonam/inhibitor combinations. According to CLSI breakpoints, aztreonam susceptibility was fully restored for 86%, 20%, and 50% of the MBL-producing Enterobacteriaceae isolates when combined with ceftazidime-avibactam, ceftolozane-tazobactam, and amoxicillin-clavulanate, respectively. In P. aeruginosa, the aztreonam-ceftazidime-avibactam combination was the most potent, even though the reduction in MICs was at most 2-fold. With the 5 S. maltophilia isolates, aztreonam-ceftazidime-avibactam and aztreonam-amoxicillin-clavulanate were found to be equal (100% susceptibility). Overall, aztreonam-ceftazidime-avibactam was the most potent combination to treat infections caused by MBL producers compared with aztreonam-amoxicillin-clavulanate and aztreonam-ceftolozane-tazobactam. However, in many cases aztreonam-amoxicillin-clavulanate was found to be as efficient as aztreonam-ceftazidime-avibactam, offering the main advantage to be markedly cheaper. We also confirmed the validity of Etest superpositions as a very simple method to determine MICs of aztreonam combinations.

Germs of thrones - spontaneous decolonization of Carbapenem-Resistant Enterobacteriaceae (CRE) and Vancomycin-Resistant Enterococci (VRE) in Western Europe: is this myth or reality?

Background: In France, Carbapenem-Resistant Enterobacteriaceae (CRE) and Vancomycin-Resistant Enterococci (VRE) are considered as Extensively Drug-Resistant (XDR) bacteria. Their management requires reinforcement of hospital's hygiene policies, and currently there is few consistent data concerning the spontaneous decolonization in XDR colonized patients. Our aim is to study the natural history of decolonization of XDR carriers over time in a hospital setting in a low prevalence country. Material and methods: Retrospective multicenter study over 2 years (2015-2016) in 2 different tertiary care hospital sites and units having an agreement for permanent cohorting of such XDR carriers. We gathered the type of microorganisms, risk factors for colonization and rectal swabs from patient's follow-up. We also evaluated patient care considering isolation precautions. Results: We included 125 patients, aged 63+/-19y, including 72.8% of CRE (n = 91), 24.8% of VRE (n = 31) and 2.4% (n = 3) co-colonized with CRE and VRE. CRE were mainly E. coli (n = 54), K. pneumoniae (n = 51) and E. cloacae (n = 6). Mechanisms of resistance were mainly OXA-48 (n = 69), NDM-1 (n = 11), OXA-232 (n = 8) and KPC (n = 3).Prior antibiotic therapy was reported in 38.4% (n = 48) of cases. Conversely, 17.6% (n = 22) received antibiotics during follow-up.Spontaneous decolonization occurred within the first 30 days in 16.4% (n = 19/116) of cases and up to 48.2% after day-90 with a median follow-up of 96 days (0-974).We estimated that XDR carriage was associated with a larger care burden in 13.6% (n = 17) of cases, especially due to a prolongation of hospitalization of 32.5 days (15-300). Conclusions: Our study shows that spontaneous decolonization is increasing over time (up to 48.2%). We can regret that only few patients underwent screening after 1 year, emphasizing the need for more monitoring and prospective studies.

Evaluation of the CRE and ESBL ELITe MGB® kits for the accurate detection of carbapenemase- or CTX-M-producing bacteria.

As carbapenemase-producing Enterobacteriaceae (CPE) and extended-spectrum ß-lactamase-producing Enterobacteriaceae (ESBL-E) are becoming a major public health issue, there is an urgent need for accurate and fast diagnostic tests. The ELITe InGenius is a fully automated sample-to-result system designed for the extraction and detection by multiplex real-time polymerase chain reaction of carbapenemases KPC, NDM, VIM, IMP, and OXA-48-like variants and CTX-M group 1 and 9-producers from diverse sample matrices such as colonies, positive blood cultures, and rectal swabs. CRE and ESBL ELITe MGB® kits were evaluated on 153 cultured colonies of enterobacterial isolates with characterized ß-lactamase content, on 30 spiked blood cultures, and the CRE kit was also evaluated on 53 clinical rectal swabs collected prospectively during a 3-month period and 10 spiked rectal swabs. CRE ELITe MGB® kit's performances reached 100% sensitivity and 100% specificity, while for the ESBL ELITe kit, 100% sensitivity and 96.6% specificity were observed, with a sample to result of less than 3 h and a total percentage of agreement with expected results of 99.6% (255/256).

Long-lasting successful dissemination of resistance to oxazolidinones in MDR Staphylococcus epidermidis clinical isolates in a tertiary care hospital in France.

OBJECTIVES: Patient- and procedure-related changes in modern medicine have turned CoNS into one of the major nosocomial pathogens. Treatments of CoNS infections are challenging owing to the large proportion of MDR strains and oxazolidinones often remain the last active antimicrobial molecules. Here, we have investigated a long-lasting outbreak (2010-13) due to methicillin- and linezolid-resistant (LR) CoNS (n = 168), involving 72 carriers and 49 infected patients. METHODS: Antimicrobial susceptibilities were tested by the disc diffusion method and MICs were determined by broth microdilution or Etest. The clonal relationship of LR Staphylococcus epidermidis (LRSE) was first determined using a semi-automated repetitive element palindromic PCR (rep-PCR) method. Then, WGS was performed on all cfr-positive LRSE (n = 30) and LRSE isolates representative of each rep-PCR-defined clone (n = 17). Self-transferability of cfr-carrying plasmids was analysed by filter-mating experiments. RESULTS: This outbreak was caused by the dissemination of three clones (ST2, ST5 and ST22) of LRSE. In these clones, linezolid resistance was caused by (i) mutations in the chromosome-located genes encoding the 23S RNA and L3 and L4 ribosomal proteins, but also by (ii) the dissemination of two different self-conjugative plasmids carrying the cfr gene encoding a 23S RNA methylase. By monitoring linezolid prescriptions in two neighbouring hospitals, we highlighted that the spread of LR-CoNS was strongly associated with linezolid use. CONCLUSIONS: Physicians should be aware that plasmid-encoded linezolid resistance has started to disseminate among CoNS and that rational use of oxazolidinones is critical to preserve these molecules as efficient treatment options for MDR Gram-positive pathogens.

OXA-244-Producing Escherichia coli Isolates, a Challenge for Clinical Microbiology Laboratories.

OXA-244 is a single-point-mutant derivative of OXA-48 displaying reduced carbapenemase activity. Here, we report the microbiological features of seven OXA-244-producing Escherichia coli isolates. Only one isolate grew on ChromID Carba Smart medium (bioMérieux), but six of the seven isolates grew on ChromID extended-spectrum-ß-lactamase (ESBL) medium (bioMérieux), as they coproduced an ESBL and/or a plasmid-encoded cephalosporinase. The production of a carbapenemase was detected in 57.1%, 71.4%, 71.4%, and 100% of the E. coli isolates using the Carba NP test, the Rapidec Carba NP test (bioMérieux), a matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) hydrolysis assay (Bruker), and the OXA-48 K-SeT assay (Coris BioConcept), respectively. Our results indicate that OXA-244-producing E. coli isolates are difficult to detect, which may lead to their silent spread.

Performance of the Xpert® Carba-R v2 in the daily workflow of a hygiene unit in a country with a low prevalence of carbapenemase-producing Enterobacteriaceae.

Early detection of patients colonised with carbapenemase-producing Enterobacteriaceae (CPE) is crucial for implementing proper infection control measures. Here we evaluated the biological performance of the Xpert® Carba-R v2 (Cepheid) in the daily workflow of a hygiene unit in a country with a low CPE prevalence. Patients repatriated from countries known for high CPE prevalence or contact patients of a known CPE carrier were targeted as being 'high-risk patients' for CPE carriage. Between September 2015 and March 2016, 241 'high-risk patients' for CPE carriage were screened using the Xpert® Carba-R v2 and by plating on chromID® CARBA Smart medium (bioMérieux) with and without ertapenem-containing enrichment culture for 24 h. Of these patients, 81.7% were previously hospitalised abroad and 18.3% were contact patients of known CPE carriers. The Xpert® Carba-R v2 was able to detect 12 OXA-48-like, 1 KPC and 1 OXA-48-like/NDM carriers. For 2 of the 14 Xpert® Carba-R v2-positive samples, cultures remained negative even on two additional screenings (performed at Days 4 and 7). The Xpert® Carba-R v2 presents 100% sensitivity, 99.13% specificity, 85.71% positive predictive value and 100% negative predictive value. This study demonstrated that the Xpert® Carba-R v2 kit is well adapted for rapid screening of high-risk patients even in low prevalence regions (in <1 h versus 24/48 h for culture). This assay may guide infection control programmes to limit the spread of CPE.

First Occurrence of OXA-72-Producing Acinetobacter baumannii in Serbia.

Here, we characterized the first OXA-72-producing Acinetobacter baumannii isolate (designated MAL) recovered from a urine sample from a Serbian patient. Antimicrobial susceptibility testing, plasmid analysis, and whole-genome sequencing (WGS) were performed to fully characterize the resistome of the A. baumannii MAL clinical isolate. The isolate was multidrug resistant and remained susceptible only to colistin and tigecycline. PCR analysis revealed the presence of the carbapenemase OXA-72, an OXA-40 variant. Extraction by the Kieser method revealed the presence of two plasmids, and one of these, a ca. 10-kb plasmid, harbored the blaOXA-72 gene. WGS revealed 206 contigs corresponding to a genome of 3.9 Mbp in size with a G+C content of 38.8%. The isolate belonged to sequence type 492 and to worldwide clone II (WWCII). Naturally occurring ß-lactamase-encoding genes (blaADC-25 and blaOXA-66) were also identified. Aminoglycoside resistance genes encoding one aminoglycoside adenyltransferase (aadA2), three aminoglycoside phosphatases (strA, strB, aphA6), and one 16S RNA methylase (armA) conferring resistance to all aminoglycosides were identified. Resistance to fluoroquinolones was likely due to mutations in gyrA, parC, and parE Of note, the resistome matched perfectly with the antibiotic susceptibility testing results.