Clinical evaluation of the BioFire Respiratory Pathogen Panel for the guidance of empirical antimicrobial therapy in critically ill patients with hospital-acquired pneumonia: A multicenter, quality improvement project.

BACKGROUND: We aimed to determine whether implementing antimicrobial stewardship based on multiplex bacterial PCR examination of respiratory fluid can enhance outcomes of critically ill patients with hospital-acquired pneumonia (HAP). METHODS: We conducted a quality improvement study in two hospitals in France. Adult patients requiring invasive mechanical ventilation with a diagnosis of HAP were included. In the pre-intervention period (August 2019 to April 2020), antimicrobial therapy followed European guidelines. In the «intervention¼ phase (June 2020 to October 2021), treatment followed a multiplex PCR-guided protocol. The primary endpoint was a composite endpoint made of mortality on day 28, clinical cure between days 7 and 10, and duration of invasive mechanical ventilation on day 28. The primary outcome was analyzed with a DOOR strategy. RESULTS: A total of 443 patients were included in 3 ICUs from 2 hospitals (220 pre-intervention; 223 intervention). No difference in the ranking of the primary composite outcome was found (DOOR: 50.3%; 95%CI, 49.9%-50.8%). The number of invasive mechanical ventilation-free days at day 28 was 10.0 [0.0; 19.0] in the baseline period and 9.0 [0.0; 20.0] days during the intervention period (p = 0.95). The time-to-efficient antimicrobial treatment was 0.43 ± 1.29 days before versus 0.55 ± 1.13 days after the intervention (p = 0.56). CONCLUSION: Implementation of Rapid Multiplex PCR to guide empirical antimicrobial therapy for critically ill patients with HAP was not associated with better outcomes. However, adherence to stewardship was low, and the study may have had limited power to detect a clinically important difference.

In Vitro Comparison of Aztreonam/Amoxicillin-Clavulanate Versus Aztreonam/Ceftazidime-Avibactam on Ceftazidime-Avibactam Resistant Stenotrophomonas maltophilia.

We investigated the in vitro susceptibility of ceftazidime-avibactam (CZA) resistant Stenotrophomonas maltophilia to the associations aztreonam/amoxicillin-clavulanate (ATM-AMC) and ATM-CZA. Forty clinical isolates of S. maltophilia recovered from sputum samples of 40 cystic fibrosis people were selected from the collection of the Nantes University Hospital, based on their resistance to CZA. Minimum inhibitory concentrations (MICs) of ATM-CZA and ATM-AMC were determined for each isolate by an Etest strip superposition method, and by Etest for each individual antibiotic. MICs of CZA, ATM, and AMC ranged from 12 to ≥256, ≥256, and 16 to ≥256 mg/L, respectively. Synergistic effects were observed with the ATM-CZA combination for all isolates (fractional inhibitory concentration index range of 0.01 to 0.27), with combination MICs ranging from 0.75 to 16 mg/L (MIC50/90 = 3/12 mg/L), corresponding to a decrease of at least 16-folds in the MIC of ATM. In 23 (57.5%) S. maltophilia isolates, the association of AMC to ATM was also synergistic and combination MICs were ≤16 mg/L (EUCAST breakpoint for ATM resistance in Pseudomonas aeruginosa). Our results show that ATM-CZA or ATM-AMC could be alternative therapeutic options against some highly resistant S. maltophilia. This encourages further experimental studies, in particular time-kill analyses, and clinical trials to delineate conditions required for use of these combinations in practice.

Doxycycline versus azithromycin for the treatment of anorectal Chlamydia trachomatis infection in women concurrent with vaginal infection (CHLAZIDOXY study): a multicentre, open-label, randomised, controlled, superiority trial.

BACKGROUND: Anorectal infections with Chlamydia trachomatis are commonly found in women. Although the efficacy of doxycycline and azithromycin is comparable in the treatment of urogenital infection, their efficacies toward anorectal infection remain unclear. We therefore aimed to compare a single dose of azithromycin with a 7-day course of doxycycline for the treatment of anorectal C trachomatis infection in women with concurrent vaginal infection. METHODS: We did a multicentre, open-label, randomised, controlled, superiority trial involving four sexually transmitted infection screening centres and three pregnancy termination centres in France. We included sexually active adult women (≥18 years) with a positive C trachomatis vaginal swab who agreed to provide self-collected anorectal swabs for C trachomatis detection. Participants were randomly assigned (1:1), using block sizes of six and eight and stratification by each investigating centre, to orally receive either azithromycin (a single 1-g dose, with or without food) or doxycycline (100 mg in the morning and evening at mealtimes for 7 days [ie, 100 mg of doxycycline twice per day for 7 days]). All laboratory staff who did the bacteriological analyses, but not the participants and the investigators, were masked to the treatment groups. The primary outcome was the microbiological anorectal cure rate defined as a C trachomatis-negative nucleic acid amplification test (NAAT) result in anorectal specimens 6 weeks after treatment initiation among women who had a baseline C trachomatis-positive anorectal NAAT result. The primary analysis was done in the modified intention-to-treat population, with multiple imputation, which included all women who underwent randomisation and had a C trachomatis-positive vaginal and anorectal NAAT result at baseline. Adverse events were reported in all women who underwent randomisation. This study is registered with ClinicalTrials.gov, number NCT03532464. FINDINGS: Between Oct 19, 2018, and April 17, 2020, we randomly assigned a total of 460 participants to either the doxycycline group (n=230) or the azithromycin group (n=230). Four (1%) of 460 participants were excluded because they refused to take doxycycline or were found to be ineligible after randomisation. Among the 456 participants, 357 (78%) had a concurrent C trachomatis-positive anorectal NAAT result at baseline; 184 (52%) of 357 were in the doxycycline group and 173 (48%) were in the azithromycin group (ie, the modified intention-to-treat population). Microbiological anorectal cure occurred in 147 (94%) of 156 participants in the doxycycline group (28 missing values) versus 120 (85%) of 142 in the azithromycin group (31 missing values; adjusted odds ratio with imputation of missing values 0·43 [95% CI 0·21-0·91]; p=0·0274). Reported adverse events possibly related to treatment were notified in 53 (12%) of 456 women: 24 (11%) of 228 in the doxycycline group and 29 (13%) of 228 in the azithromycin group. Gastrointestinal disorders were the most frequently occurring, in 43 (9%) of 456 women: 17 (8%) of 228 in the doxycycline group and 26 (11%) of 228 in the azithromycin group. INTERPRETATION: The microbiological anorectal cure rate was significantly lower among women who received a single dose of azithromycin than among those who received a 1-week course of doxycycline. This finding suggests that doxycycline should be the first-line therapy for C trachomatis infection in women. FUNDING: French Ministry of Health. TRANSLATION: For the French translation of the abstract see Supplementary Materials section.

Potential Impact of Rapid Multiplex PCR on Antimicrobial Therapy Guidance for Ventilated Hospital-Acquired Pneumonia in Critically Ill Patients, A Prospective Observational Clinical and Economic Study.

Objectives: To investigate the potential impact of the syndromic multiplex FilmArray® Pneumonia plus Panel (FAPP) on the antimicrobial treatment guidance of patients with ventilated hospital-acquired pneumonia (VHAP). Methods: Respiratory fluids from 100 adult patients with VHAP, receiving invasive mechanical ventilation in three intensive care units from one French university hospital, were tested prospectively using FAPP. Conventional cultures were performed in parallel as routine practice. Clinicians were left blinded to the FAPP results. Antimicrobial therapies based on FAPP results were simulated by independent blinded experts according to a predefined algorithm and compared to 1) those prescribed in practice according to local guidelines (real-life), and 2) those that complied with the international ERS/ESICM/ESCMID/ALAT recommendations. The primary endpoint was the number of days of broad-spectrum antimicrobial therapy. Secondary endpoints were the rates of microbiological treatment failure and cost-effectiveness ratio. Results: The predicted median duration of broad-spectrum antibiotics was 0 [0-1.25] day in the FAPP-based simulation, versus 2 [0-6] days in real-life (p<0.0001) and 2 [2-3.25] days in the recommendations-based simulation (p<0.0001). Treatment failure was predicted in 3% of cases with FAPP results versus observed in 11% in real-life (p=0.08) and 6% with recommendations-based simulation (p=0.37). The incremental cost-effectiveness ratio was 1 121 € [-7021; 6794] to avoid one day of non-optimized antimicrobial therapy. Conclusions: Our results suggest that using FAPP in patients with VHAP has the potential to reduce the use of broad-spectrum antimicrobial therapy without increasing the risk of microbial treatment failure.

Assessment of a Multiplex LAMP Assay (Eazyplex® CSF Direct M) for Rapid Molecular Diagnosis of Bacterial Meningitis: Accuracy and Pitfalls.

Background: Automated molecular panels are attractive tools for improving early meningitis diagnosis. This study assessed the Eazyplex® CSF direct M panel (EP), a multiplex real-time Loop-Mediated Isothermal Amplification assay. Methods: From December 2016 to December 2019, cerebrospinal fluid (CSF) samples were routinely tested with the EP V1.0. CSF parameters and microbiological and clinical data were retrospectively collected. Results: Out of 230 CSF samples, the EP yielded positive, negative, and invalid results for 32 (13.9%) (16 N. meningitidis, nine S. pneumoniae, two S. agalactiae, two E. coli, two H. influenzae, one L. monocytogenes), 182 (79.1%), and 16 (7%) samples, respectively. Among the positive samples, 14 (44%) remained negative in culture (antibiotic therapy before lumbar puncture (n = 11), meningococcal meningitis (n = 3)). High CSF protein concentrations and cellularity were associated with LAMP inhibition, counteracted by centrifugation. The automated software yielded 13 false positive and five false negative results. Amplification curve analysis was necessary and enabled the attainment of positive (PPA) and negative percentage agreement and positive and negative predictive values of 91.4%, 100%, 100%, and 98.3%. Three false negative results remained (two E. coli and one N. meningitidis). E. coli presented the poorest PPA (50%). Conclusion: This work confirms the strong performance of the EP, of particular interest in cases of antibiotic therapy before lumbar puncture.

Evaluation of the FilmArray® Pneumonia Plus Panel for Rapid Diagnosis of Hospital-Acquired Pneumonia in Intensive Care Unit Patients.

The FilmArray® Pneumonia plus Panel (FAPP) is a new multiplex molecular test for hospital-acquired pneumonia (HAP), which can rapidly detect 18 bacteria, 9 viruses, and 7 resistance genes. We aimed to compare the diagnosis performance of FAPP with conventional testing in 100 intensive care unit (ICU) patients who required mechanical ventilation, with clinically suspected HAP. A total of 237 samples [76 bronchoalveolar lavages (BALDS) and 82 endotracheal aspirates (ETADS) obtained at HAP diagnosis, and 79 ETA obtained during follow-up (ETATT)], were analyzed independently by routine microbiology testing and FAPP. 58 patients had paired BALDS and ETADS. The positivity thresholds of semi-quantified bacteria were 103-104 CFUs/mL or 104 copies/mL for BAL, and 105 CFUs/mL or copies/mL for ETA. Respiratory commensals (H. influenzae, S. aureus, E. coli, S. pneumoniae) were the most common pathogens. Discordant results for bacterial identification were observed in 33/76 (43.4%) BALDS and 36/82 (43.9%) ETADS, and in most cases, FAPP identified one supplemental bacteria (23/33 BALDS and 21/36 ETADS). An absence of growth, or polybacterial cultures, explained almost equally the majority of the non-detections in culture. No linear relationship was observed between bin and CFUs/mL variables. Concordant results between paired BALDS and ETADS were obtained in 46/58 (79.3%) patients with FAPP. One of the 17 resistance genes detected with FAPP (mecA/C and MREJ) was not confirmed by conventional testing. Overall, FAPP enhanced the positivity rate of diagnostic testing, with increased recognition of coinfections. Implementing this strategy may allow clinicians to make more timely and informed decisions.

Listeria Endophthalmitis Cured With Linezolid in an Immunocompetent Farmer Woman: Hazard of a Sweep of a Cow's Tail.

We report the first case of an unexpected exogenous Listeria monocytogenes endophthalmitis in a previously healthy woman after a cow's tail's sweep, successfully treated with surgery and linezolid. It is the first case carried out with linezolid to treat Listeria endophthalmitis. Therefore, it may challenge the requirement for intravenous antibiotics for long-term treatment.

First experience of a multicenter external quality assessment of molecular 16S rRNA gene detection in bone and joint infections.

The objective of this study was to assess the performance of seven French laboratories for 16S rRNA gene detection by real-time PCR in the diagnosis of bone and joint infection (BJI) to validate a large multicenter study. External quality control (QC) was required owing to the differences in extraction procedures and the molecular equipment used in the different laboratories. Three proficiency sets were organized, including four bacterial DNA extracts and four bead mill-pretreated osteoarticular specimens. Extraction volumes, 16S rRNA gene primers, and sequencing interpretation rules were standardized. In order to assess each laboratory's ability to achieve the best results, scores were assigned, and each QC series was classified as optimal, acceptable, or to be improved. A total of 168 QCs were sent, and 160 responses were analyzed. The expected results were obtained for 93.8%, with the same proportion for extracts (75/80) and clinical specimens (75/80). For the specimens, there was no significant difference between manual and automated extraction. This QC demonstrated the ability to achieve good and homogeneous results using the same 16S rRNA gene PCR with different equipment and validates the possibility of high-quality multicenter studies using molecular diagnosis for BJI.

Evaluation of 16S rRNA gene PCR sensitivity and specificity for diagnosis of prosthetic joint infection: a prospective multicenter cross-sectional study.

There is no standard method for the diagnosis of prosthetic joint infection (PJI). The contribution of 16S rRNA gene PCR sequencing on a routine basis remains to be defined. We performed a prospective multicenter study to assess the contributions of 16S rRNA gene assays in PJI diagnosis. Over a 2-year period, all patients suspected to have PJIs and a few uninfected patients undergoing primary arthroplasty (control group) were included. Five perioperative samples per patient were collected for culture and 16S rRNA gene PCR sequencing and one for histological examination. Three multicenter quality control assays were performed with both DNA extracts and crushed samples. The diagnosis of PJI was based on clinical, bacteriological, and histological criteria, according to Infectious Diseases Society of America guidelines. A molecular diagnosis was modeled on the bacteriological criterion (≥ 1 positive sample for strict pathogens and ≥ 2 for commensal skin flora). Molecular data were analyzed according to the diagnosis of PJI. Between December 2010 and March 2012, 264 suspected cases of PJI and 35 control cases were included. PJI was confirmed in 215/264 suspected cases, 192 (89%) with a bacteriological criterion. The PJIs were monomicrobial (163 cases [85%]; staphylococci, n = 108; streptococci, n = 22; Gram-negative bacilli, n = 16; anaerobes, n = 13; others, n = 4) or polymicrobial (29 cases [15%]). The molecular diagnosis was positive in 151/215 confirmed cases of PJI (143 cases with bacteriological PJI documentation and 8 treated cases without bacteriological documentation) and in 2/49 cases without confirmed PJI (sensitivity, 73.3%; specificity, 95.5%). The 16S rRNA gene PCR assay showed a lack of sensitivity in the diagnosis of PJI on a multicenter routine basis.

Significant finding of Bordetella holmesii DNA in nasopharyngeal samples from French patients with suspected pertussis.

Pertussis is routinely diagnosed with real-time PCR based on insertion sequence IS481, which is not specific for Bordetella pertussis. We conducted a retrospective study using real-time PCRs specific for Bordetella pertussis and for Bordetella holmesii on 177 samples positive for IS481 PCR. Bordetella holmesii DNA was detected in 20.3% samples collected from adolescents and adults.

Proficiency program for real-time PCR diagnosis of Bordetella pertussis infections in French hospital laboratories and at the French National Reference Center for Whooping Cough and other Bordetelloses.

With the support of a ministerial program for innovative and expensive technologies, dedicated to the economic evaluation of laboratory diagnosis of pertussis by real-time PCR, external quality assessment for real-time IS481 PCR was carried out. Coordinated by the National Centre of Reference of Pertussis and other Bordetelloses (NCR), this study aimed to harmonize and to assess the performances of eight participating microbiology hospital laboratories throughout the French territory. Between January 2006 and February 2007, 10 proficiency panels were sent by the NCR (ascending proficiency program), representing a total of 49 samples and including eight panels to analyze and evaluate the global sensitivity and specificity of real-time PCR, one to assess the limit of detection, and one to evaluate nucleic acid extraction methods. As part of the descending proficiency program, extracted DNA from clinical samples was sent by the eight participating laboratories in different panels and analyzed by the NCR. In the ascending proficiency analysis, the sensitivity and specificity of the real-time PCR methods were 92.2% and 94.3%, respectively. The limit of detection of the different methods ranged between 0.1 and 1 fg/microl (0.2 to 2 CFU/microl). The nucleic acid extraction methods showed similar performances. During the descending proficiency analysis, performed with 126 samples, the result of the NCR for 15 samples (11.9%) was discordant with the result obtained by the source laboratory. Despite several initial differences, harmonization was easy and performances were homogeneous. However, the risk of false-positive results remains quite high, and we strongly recommend establishment of uniform quality control procedures performed regularly.