Regional spread of an atypical ESBL-producing Escherichia coli ST131H89 clone among different human and environmental reservoirs in Western Switzerland.

We describe the inter-regional spread of a novel ESBL-producing Escherichia coli subclone (ST131H89) in long-term care facility residents, general population, and environmental water sources in Western Switzerland between 2017 and 2020. The study highlights the importance of molecular surveillance for tracking emerging antibiotic-resistant pathogens in healthcare and community settings.

Evaluation of PhenoMATRIX and PhenoMATRIX PLUS for the screening of MRSA from nasal and inguinal/perineal swabs using chromogenic media.

The objective of this study was to assess the clinical performances of PhenoMATRIX and PhenoMATRIX PLUS for the screening of methicillin-resistant Staphylococcus aureus (MRSA) from nasal and inguinal/perineal ESwabs using chromogenic media. The automated performances were compared to the manual reading. Additionally, we evaluated PhenoMATRIX PLUS for the automatic release of the negative results to the Laboratory Information System (LIS) and the automatic discharge of the negative plates from the incubators. A total of 6,771 non-duplicate specimens were used by PhenoMATRIX as a machine learning model. The validation of these settings was performed on 17,223 non-duplicate specimens. The MRSA positivity rate was 5% (866/17,223). Validated settings were then used by PhenoMATRIX PLUS on another 1,409 non-duplicate specimens. The sensitivities of PhenoMATRIX and PhenoMATRIX PLUS were 99.8% [95% confidence interval (CI), 99.2%-99.9%] and 100% (95% CI, 92.1%-100%), respectively. The specificities of PhenoMATRIX and PhenoMATRIX PLUS were 99.1% (95% CI, 99.0%-99.2%) and 95.2% (95% CI, 93.8%-96.1%), respectively. All the 1,297 MRSA-negative specimens analyzed by PhenoMATRIX PLUS were automatically released and sent to the LIS immediately after availability of the culture image on the WASPLab (100% accuracy). All negative media plates were automatically discarded. PhenoMATRIX PLUS decreases the time spent by technologists on negative plates and ensures optimal usage of the incubators' capacity.

[Reading an antibiogram in the context of antimicrobial resistance]. / Lecture de l'antibiogramme dans un contexte de résistance aux antibiotiques.

Reading an antibiogram, however simple it may appear at first glance, is full of sometimes implicit but valuable information in the daily practice of the physician. A more subtle reading requires knowledge in the microbiology and pharmacology fields, even more so in the presence of resistant bacteria. The aim of this article is to provide the clinician with some keys to read and understand an antibiogram in the current context.

La lecture d'un antibiogramme, aussi simple qu'elle puisse paraître au premier coup d'œil, regorge d'informations parfois implicites qui peuvent être précieuses dans la pratique quotidienne du médecin clinicien. Une lecture plus approfondie nécessite quelques connaissances de microbiologie et de pharmacologie, ce d'autant plus lorsque la bactérie est multirésistante. Cet article a pour but d'apporter quelques clés de lecture de l'antibiogramme au médecin clinicien dans ce contexte.

Draft Genome Sequence of a Mixed-Serogroup W/Y Invasive Neisseria meningitidis Strain.

We report the genome of a Neisseria meningitidis strain (GE-156) that was isolated in Switzerland from a patient diagnosed with bacteremia. The strain belongs to a rare mixed serogroup W/Y and sequence type 11847 (clonal complex 167), as revealed by both routine laboratory examination and genomic sequencing.

Diagnostic test accuracy of an automated device for the MALDI target preparation for microbial identification.

The objective of this study was to evaluate the performance of the Copan Colibrí™ against the manual preparation of the MALDI targets. We analyzed 416 (31 different species) non-duplicate strains covering the most important species identified in clinical routine. We also assessed the intra-strain repeatability between the comparable methods. We then analyzed the performance of this new method after implementation in routine on 12,253 aerobic bacterial isolates and yeasts, encompassing a total of 42 different species. Among the 416 strains analyzed, 6.3% (26/416) and 10.8% (45/416) had a score value < 2 when processed by the Colibri™ and manual method, respectively. Only 5.9% (9/152) of the Gram positive rods and cocci had a score values < 2 by the Colibri™ versus 20.4% (31/152) by the manual method. We confirmed that this relative superiority observed for the Colibri™ was due primarily in the use of the formic acid protocol. For the Gram-negative bacteria, the results of both methods were comparable; 6.6% (17/256) and 4.7% (12/256) had a score value < 2 by the Colibri™ and the manual method, respectively. After implementation in routine, the results according to the Biotyper score cut-off values were distributed as follows: < 1.70: 2.5% (304/12,253), 1.70-1.79: 1.9% (227/12,253), 1.80-1.89: 3.1% (377/12,253), 1.90-1.99: 6.7% (825/12,253), and ≥ 2: 85.9% (10,520/12,253). The Colibrí™ coupled to MALDI-TOF/MS revealed good performances and higher intra-strain repeatability as compared to the manual preparation of the MALDI targets.

Fully Automated EUCAST Rapid Antimicrobial Susceptibility Testing (RAST) from Positive Blood Cultures: Diagnostic Accuracy and Implementation.

The objective of this study was to evaluate the accuracy and robustness of a fully automated EUCAST RAST (rapid antimicrobial susceptibility test) directly from positive blood culture and to appreciate its implementation constraints. This study was conducted in two phases: (i) spiked blood culture bottles (BCs) using 779 non-duplicate clinical isolates and (ii) a prospective clinical trial including 534 positive BCs sequentially processed in routine at the Bacteriology Laboratory of Geneva University Hospitals. The RAST results were assessed against EUCAST standardized disk diffusion testing results. Our first finding was that the results of the spiked BCs precisely predicted the clinical trial results. The overall categorical agreements for all species analyzed were greater than 95% at the different time points. RAST for Pseudomonas aeruginosa, however, raised several challenges. The categorical agreement for imipenem was lower than 95% at 6 h and was not improved with longer incubation times. Additionally, piperacillin-tazobactam, ceftazidime, and cefepime cannot be released at 6 h due to suboptimal performances, but the categorical agreement substantially improved at 8 h. Our results establish that the performance of fully automated EUCAST RAST directly from positive blood culture bottles is consistently robust, even for the detection of extended-spectrum ß-lactamase (ESBL), carbapenemase-producing bacteria, and methicillin-resistant Staphylococcus aureus (MRSA). The automation markedly enhanced the percentage of readable inhibition zones and reduced the percentage of isolates categorized in the area of technical uncertainty (ATU). In summary, a fully automated EUCAST RAST can substantially improve laboratory workflow by reducing hands-on time and removing the strong constraints linked to manual read-outs at precisely defined times.

An interventional quasi-experimental study to evaluate the impact of a rapid screening strategy in improving control of nosocomial extended-spectrum beta-lactamase-producing Enterobacterales and carbapenemase-producing organisms in critically ill patients.

INTRODUCTION: Rapid molecular tests could accelerate the control of extended-spectrum beta-lactamase-producing Enterobacterales (ESBL-PE) and carbapenemase-producing organisms (CPO) in intensive care units (ICUs). OBJECTIVE AND METHODS: This interventional 12-month cohort study compared a loop-mediated isothermal amplification (LAMP) assay performed directly on rectal swabs with culturing methods (control period, 6 months), during routine ICU screening. Contact precautions (CP) were implemented for CPO or non-E. coli ESBL-producing Enterobacterales (nEcESBL-PE) carriers. Using survival analysis, we compared the time intervals from admission to discontinuation of unnecessary preemptive CP among patients at-risk and the time intervals from screening to implementation of CP among newly identified carriers. We also compared diagnostic performances, and nEcESBL-PE/CPO acquisition rates. This study is registered, ISRCTN 23588440. RESULTS: We included 1043 patients. During the intervention and control phases, 92/147 (62.6%) and 47/86 (54.7%) of patients at-risk screened at admission were candidates for early discontinuation of preemptive CP. The LAMP assay had a positive predictive value (PPV) of 44.0% and a negative predictive value (NPV) of 99.9% for CPO, and 55.6% PPV and 98.2% NPV for nEcESBL-PE. Due to result notification and interpretation challenges, the median time from admission to discontinuation of preemptive CP increased during the interventional period from 80.5 (95% CI 71.5-132.1) to 88.3 (95% CI 57.7-103.7) hours (p = 0.47). Due to the poor PPV, we had to stop using the LAMP assay to implement CP. No difference was observed regarding the incidence of nEcESBL-PE and CPO acquisition. CONCLUSION: A rapid screening strategy with LAMP assays performed directly on rectal swabs had no benefit for infection control in a low-endemicity setting.

Total Laboratory Automation for Rapid Detection and Identification of Microorganisms and Their Antimicrobial Resistance Profiles.

At a time when diagnostic bacteriological testing procedures have become more complex and their associated costs are steadily increasing, the expected benefits of Total laboratory automation (TLA) cannot just be a simple transposition of the traditional manual procedures used to process clinical specimens. In contrast, automation should drive a fundamental change in the laboratory workflow and prompt users to reconsider all the approaches currently used in the diagnostic work-up including the accurate identification of pathogens and the antimicrobial susceptibility testing methods. This review describes the impact of TLA in the laboratory efficiency improvement, as well as a new fully automated solution for AST by disk diffusion testing, and summarizes the evidence that implementing these methods can impact clinical outcomes.

Epidemiology of ESBL-producing Escherichia coli from repeated prevalence studies over 11 years in a long-term-care facility.

BACKGROUND: Escherichia coli sequence type (ST) 131 H30 is an emerging multidrug resistant subclone, known to spread and cause outbreaks in long-term care facilities (LTCFs). OBJECTIVES AND METHODS: From 2010 through 2020, we performed 11 yearly surveillance studies for determining the prevalence of digestive carriage of ESBL-producing E. coli (ESBL-EC) among residents in a university-affiliated LCTF. Sequencing and genotyping of selected isolates were performed to characterize temporal trends in the prevalence and epidemic potential of ESBL-EC subclones, and for evaluating a potential rebound effect following discontinuation of contact precautions for ESBL-EC carriers in January 2019. RESULTS: This study included 2'403 LTCF residents, with 252 (10.5%) positive for ESBL-EC. Among the 236 ESBL-EC isolates available for typing, 58.0% belonged to the ST131 lineage, including 94/137 (68.6%) ST131 H30 isolates. An increasing yearly prevalence was observed for ESBL-EC (from 4.6 to 9.4%; p = 0.11), but not for the ST131 H30 subclone, which peaked in 2015 and declined thereafter. Multiple previously unnoticed ESBL-EC outbreaks occurred in the LTCF. Since 2018, we noted the clonal expansion of a rare ST131 H89 subclone (O16:H5) harboring CTX-M-14 and CTX-M-24. No rebound effect was observed in ESBL-EC prevalence nor in the different subclones following discontinuation of contact precautions for ESBL-EC carriers since 2019. CONCLUSION: Clonal fluctuation was observed for ST131 H30 ESBL-EC with a current decline in prevalence. Surveillance should include the evolution of ST131 non-H30 subclones, which may spread in LTCFs. Our findings suggest that discontinuation of contact precautions for ESBL-EC carriers in LTCFs may be safely implemented, in support of European recommendations to limit ESBL-producing Enterobacteriaceae control measures in endemic settings to non-E. coli.

Performance of Fully Automated Antimicrobial Disk Diffusion Susceptibility Testing Using Copan WASP Colibri Coupled to the Radian In-Line Carousel and Expert System.

The purpose of the present study was to assess the agreement at the categorical level between the Vitek 2 system and the Colibri coupled to the Radian under real routine laboratory conditions. The 675 nonduplicate clinical strains included in this study (249 Enterobacterales isolates, 198 Pseudomonas aeruginosa, 107 Staphylococcus aureus, 78 coagulase-negative staphylococci, 38 Enterococcus faecalis, and 5 Enterococcus faecium) were isolated from nonconsecutive clinical samples referred to our laboratory between June and November 2020. In addition, 43 carbapenemase-producing Enterobacterales (CPE) formerly identified and stored in our laboratory were added to the panel, for a total of 718 strains. The overall categorical agreements between the two compared methods were 99.3% (4,350/4,380; 95% CI 99% to 99.5%); 98.6% (2,147/2,178; 95% CI 98.0% to 99.0%); 99.4% (1,839/1,850; 95% CI 98.9% to 99.7%); and 99.4% (342/344; 95% CI 97.9% to 99.8%) for Enterobacterales, P. aeruginosa, Staphylococcus spp., and Enterococcus spp., respectively. The most important cause of the very major errors encountered on the Vitek 2 for P. aeruginosa (62%, 13/21) was related to the presence of heteroresistant populations. Among the 43 CPE included in this study, one OXA-48-like, and one OXA-181-like were missed by the Vitek 2, even by rigorously applying the CPE screening cutoffs defined by EUCAST. The Colibri coupled to the Radian provide a fully automated solution for antimicrobial disk diffusion susceptibility testing with an accuracy that is equal to or better than that of the Vitek 2 system.

Performances of automated digital imaging of Gram-stained slides with on-screen reading against manual microscopy.

The objective of this study was to evaluate the performances of the automated digital imaging of Gram-stained slides against manual microscopy. Four hundred forty-three identified Gram-stained slides were included in this study. When both methods agreed, we considered the results as correct, and no further examination was carried out. Whenever the methods gave discrepant results, we reviewed the digital images and the glass slides by manual microscopy to avoid incorrectly read smears. The final result was a consensus of multiple independent reader interpretations. Among the 443 slides analyzed in this study, 101 (22.8%) showed discrepant results between the compared methods. The rates of discrepant results according to the specimen types were 5.7% (9/157) for positive blood cultures, 42% (60/142) for respiratory tract specimens, and 22% (32/144) for sterile site specimens. After a subsequent review of the discrepant slides, the final rate of discrepancies dropped to 7.0% (31/443). The overall agreement between the compared methods and the culture results reached 78% (345/443) and 79% (349/443) for manual microscopy and automated digital imaging, respectively. According to culture results, the specificity for automated digital imaging and manual microscopy were 90.8% and 87.7% respectively. In contrast, sensitivity was 84.1% for the two compared methods. The discrepant results were mostly encountered with microorganism morphologies of rare occurrence. The results reported in this study emphasize that on-screen reading is challenging, since the recognition of morphologies on-screen can appear different as compared to routine manual microscopy. Monitoring of Gram stain errors, which is facilitated by automated digital imaging, remains crucial for the quality control of reported Gram stain results.

Unexpectedly High False-Positive Rates for Haemophilus influenzae Using a Meningoencephalitis Syndromic PCR Panel in Two Tertiary Centers.

False-positive results in the diagnostic of meningitis and encephalitis pose important challenges. This study aimed to determine false-positive rates for Haemophilus influenzae in cerebrospinal fluids evaluated by the BioFire FilmArray® Meningitis/Encephalitis Panel. We conducted a retrospective study of all H. influenzae-positive FilmArray®. Meningitis/Encephalitis Panel results from June 2016 to October 2019 in two Swiss university hospitals. Cases were classified as true positive, likely true-positive, and likely false-positive results according to cerebrospinal fluid culture, H. influenzae-specific quantitative real-time PCR (qPCR), and Gram staining, as well as culture of other materials. We performed 3,082 panels corresponding to 2,895 patients: 0.6% of the samples (18/3,082) were positive for H. influenzae. Culture and H. influenzae-specific qPCR were performed on 17/18 (94.4%) and 3/18 (16.7%) cerebrospinal fluid samples, respectively; qPCR was negative in all cases. Among 17 samples sent for culture, 10 concerned patients were not treated with antibiotics prior to lumbar puncture. Only 1/17 revealed growth of H. influenzae and was classified as a true positive. We further classified 3/18 (16.7%) cases with the identification of Gram-negative rods in the cerebrospinal fluid or positive blood cultures for H. influenzae as likely true-positive and 14/18 (77.8%) cases as likely false-positive. Diagnostic results should always be interpreted together with the clinical presentation, cerebrospinal fluid analysis, and other available microbiological results. All H. influenzae-positive results should be viewed with special caution and a H. influenzae-specific qPCR should be systematically considered.

Household acquisition and transmission of extended-spectrum ß-lactamase (ESBL) -producing Enterobacteriaceae after hospital discharge of ESBL-positive index patients.

OBJECTIVES: This study aimed to determine rates and risk factors of extended-spectrum ß-lactamase-producing Enterobacteriaceae (ESBL-PE) acquisition and transmission within households after hospital discharge of an ESBL-PE-positive index patient. METHODS: Two-year prospective cohort study in five European cities. Patients colonized with ESBL-producing Escherichia coli (ESBL-Ec) or Klebsiella pneumoniae (ESBL-Kp), and their household contacts were followed up for 4 months after hospital discharge of the index case. At each follow up, participants provided a faecal sample and personal information. ESBL-PE whole-genome sequences were compared using pairwise single nucleotide polymorphism-based analysis. RESULTS: We enrolled 71 index patients carrying ESBL-Ec (n = 45), ESBL-Kp (n = 20) or both (n = 6), and 102 household contacts. The incidence of any ESBL-PE acquisition among household members initially free of ESBL-PE was 1.9/100 participant-weeks at risk. Nineteen clonally related household transmissions occurred (case to contact: 13; contact to case: 6), with an overall rate of 1.18 transmissions/100 participant-weeks at risk. Most of the acquisition and transmission events occurred within the first 2 months after discharge. The rate of ESBL-Kp household transmission (1.16/100 participant-weeks) was higher than of ESBL-Ec (0.93/100 participant-weeks), whereas more acquisitions were noted for ESBL-Ec (1.06/100 participant-weeks) compared with ESBL-Kp (0.65/100 participant-weeks). Providing assistance for urinary and faecal excretion to the index case by household members increased the risk of ESBL-PE transmission (adjusted prevalence ratio 4.3; 95% CI 1.3-14.1). CONCLUSIONS: ESBL-PE cases discharged from the hospital are an important source of ESBL-PE transmission within households. Most acquisition and transmission events occurred during the first 2 months after hospital discharge and were causally related to care activities at home, highlighting the importance of hygiene measures in community settings. CLINICAL STUDY REGISTRATION: German Clinical Trials Register, DRKS-ID: DRKS00013250.

Mini Review: Clinical Routine Microbiology in the Era of Automation and Digital Health.

Clinical microbiology laboratories are the first line to combat and handle infectious diseases and antibiotic resistance, including newly emerging ones. Although most clinical laboratories still rely on conventional methods, a cascade of technological changes, driven by digital imaging and high-throughput sequencing, will revolutionize the management of clinical diagnostics for direct detection of bacteria and swift antimicrobial susceptibility testing. Importantly, such technological advancements occur in the golden age of machine learning where computers are no longer acting passively in data mining, but once trained, can also help physicians in making decisions for diagnostics and optimal treatment administration. The further potential of physically integrating new technologies in an automation chain, combined to machine-learning-based software for data analyses, is seducing and would indeed lead to a faster management in infectious diseases. However, if, from one side, technological advancement would achieve a better performance than conventional methods, on the other side, this evolution challenges clinicians in terms of data interpretation and impacts the entire hospital personnel organization and management. In this mini review, we discuss such technological achievements offering practical examples of their operability but also their limitations and potential issues that their implementation could rise in clinical microbiology laboratories.

Impact of Total Laboratory Automation on Turnaround Times for Urine Cultures and Screening Specimens for MRSA, ESBL, and VRE Carriage: Retrospective Comparison With Manual Workflow.

Using computerized time-stamps, we compared the turnaround-times (TAT) for urine samples and screening ESwabs of MRSA, VRE, and ESBL carriage in the bacteriology laboratory of Geneva University Hospitals between January and December 2017 (period preceding the implementation of the WASPLabTM) with the same specimen types analyzed between January and December 2019 (period after the implementation of the automation). During both 1-year periods, a total of 98'380 specimens were analyzed (48'158 in 2017 vs. 50'222 in 2019). On the WASPLabTM, all culture plates were imaged at defined intervals each day of incubation, but the processing of the cultures (i.e., pathogen identification and antimicrobial susceptibility testing) was only performed during day shift hours (~8:00 A.M. to 4:30 P.M.). The median TAT for negative reports decreased by almost half for urine samples from 52.1 (2017) to 28.3 h (2019) (p < 0.001), and for MRSA screening specimens from 50.7 to 26.3 h (p < 0.001). The difference in median TAT for negative reports was less pronounced for screening of ESBL (50.2 vs. 43.0 h) (p < 0.001) and VRE (50.6 vs. 45.7 h) (p < 0.001). Despite a trend toward shorter result delivery for positive samples, there was no significant change in the median TAT. These results suggest that TAT for negative samples immediately benefit from automation, whereas TAT for positive samples also depend on the laboratory hours of operation and daily human resource management.

Strain coverage of Bexsero vaccine assessed by whole-genome sequencing over a cohort of invasive meningococci of serogroups B and W isolated in Switzerland.

Invasive meningococcal disease (IMD), caused by Neisseria meningitidis (Nm) strains, is a life-threatening but vaccine-preventable condition. Bexsero is a four-component vaccine that offers broad protection against Nm of serogroup B (NmB), particularly common in Europe. In Switzerland, Bexsero has not yet been licensed and no information is available concerning the predicted vaccine coverage on isolates of circulating Nm. We performed genotyping of Bexsero antigen loci by whole-genome sequencing (WGS) on 104 NmB collected in Switzerland in the 2010-2015 period. We searched for antigen variants previously defined as predictors of strain coverage and estimated that 50% of IMD NmB strains were potentially covered by the vaccine. Clonal complexes (cc) 32, 41/44 and 269, considered the best covered lineages, were further sub-typed according to Bexsero Antigen Sequence Type (BAST) scheme. We also genotyped by WGS 40 Nm of serogroup W (NmW) collected in the country between 2010 and 2016. NmW cc22 isolates appeared to be covered by the vaccine, which was not the case for cc11 isolates, whose incidence has recently increased in Switzerland and all over Europe. Our work underlines the benefit of using WGS for surveillance of vaccine antigen variant distribution in local Nm population and taking proper measures to prevent the spread of NmB.

Septic shock caused by Capnocytophaga canis after a cat scratch.

Capnocytophaga canis is an uncommon cause of septic shock. Only three cases have been previously reported in the literature. In this article, we describe the case of a 70-year-old male admitted to the intensive care unit for septic shock of unknown origin. On day 2, one anaerobic bottle out of the two sets taken at admission turned positive with Gram-negative bacilli. The pathogen was identified by 16S rRNA gene as C. canis. The strain was characterized and compared with other clinical isolates of Capnocytophaga spp.

Implementation of the WASPLab™ and first year achievements within a university hospital.

In essence, automation can be driven by several of the following incentives: increased processing capacity of the laboratory, better costs control through processes standardization, optimized traceability, or improved workflows to reduce turnaround times (TAT). This project aims at presenting an overview of the project management and change management with a focus on the major challenges addressed by lab staff and laboratory leadership during the different phases of the implementation of the WASPLab™ in a routine clinical bacteriology laboratory. This paper reports our experience and reviews changes in the bacteriology laboratory at Geneva University Hospitals when shifting to the WASPLab™. Practically, the whole automation process was segmented into different packages (specimen type-based segmentation) allowing sequential validation, staff training, and routine implementation. Such process allowed reaching 90% of the identified "automatable" samples within 1 year, including personal training, documentation for accreditation supported by publications, without interrupting routine operations. In addition, we implemented a validated automated solution for antimicrobial disk diffusion susceptibility testing. Structured supervision and accurate monitoring of all the activities related to the automation project including key partners such as IT support, technical committee, and after-sales service guaranteed a swift and timely achievement of the project allowing the improvement of the workflow in routine bacteriology within 1 year.

Second Periprosthetic Joint Infection Caused by Streptococcus dysgalactiae: How Genomic Sequencing Can Help Defining the Best Therapeutic Strategy.

Primary and revision arthroplasties are increasing worldwide, as are periprosthetic joint infections (PJI). The management of PJI requires surgery, the strategy of which is dictated by the acute or chronic nature of the infection, with an exchange of the implant in the event of a chronic PJI or in the case of recurrence with the same pathogen. We report the case of a 63-year-old man with two episodes of Streptococcus dysgalactiae subsp. equisimilis PJI within 9 months. Based on clinical suspicion of an haematogenous PJI, the patient was treated by DAIR (debridement, antibiotics, implant retention), while genomic sequencing revealed two different strains, confirming our hypothesis that no additional surgery was needed. Hence, we report a case where genomic analysis was decisive for the decision of the best therapeutic strategy.