Simulated Pediatric Blood Cultures to Assess the Inactivation of Clinically Relevant Antimicrobial Drug Concentrations in Resin-Containing Bottles.

The bacteremia level as well as the administration of antibiotics before blood collection may significantly affect the recovery of bacterial pathogens from pediatric blood cultures in BacT/Alert Virtuo or Bactec FX BC systems, which remain the common techniques to diagnose bacteremia in pediatric patients. We simulated pediatric blood cultures with low or intermediate bacteremia level to evaluate BacT/Alert PF Plus and Bactec Peds Plus blood culture bottles for resin-based inactivation of 16 antibiotic-bacterium combinations. Overall, 105/192 (54.7%) of BacT/Alert PF Plus bottles and 69/192 (36.0%) of Bactec Peds Plus bottles allowed organisms to grow when exposed to antibiotics. In particular, both BacT/Alert PF Plus and Bactec Peds Plus bottles proved to be effective with piperacillin/tazobactam and Pseudomonas aeruginosa or with oxacillin and methicillin-susceptible Staphylococcus aureus (100% growth), whereas no effectiveness was apparent with ceftriaxone and Escherichia coli, Streptococcus agalactiae, or Streptococcus pneumoniae or with cefepime and E. coli (0% growth). In some relevant instances (e.g., with vancomycin and methicillin-resistant S. aureus or Streptococcus pneumoniae), BacT/Alert PF Plus bottles were superior to Bactec Peds Plus bottles. Together, these findings underscore the potentiality of resin-containing bottles to enhance diagnosis of bacteremia in pediatric patients on antimicrobial therapy. This is particularly true with one of the evaluated BC systems and with simulated intermediate bacteremia level only.

Evaluation of three commercial assays for SARS-CoV-2 molecular detection in upper respiratory tract samples.

The increasing COVID-19 widespread has created the necessity to assess the diagnostic accuracy of newly introduced (RT-PCR based) assays for SARS-CoV-2 RNA detection in respiratory tract samples. We compared the results of the Allplex™ 2019-nCoV assay with those of the Simplexa™ COVID-19 Direct assay and the Quanty COVID-19 assay, respectively, all performed on 125 nasal/oropharyngeal swab samples of patients with COVID-19 suspicion. Fifty-four samples were positive, and 71 were negative with the Allplex™ assay, whereas 47 of 54 samples were also positive with the Simplexa™ assay. The Quanty assay detected 55 positive samples, including the 54 positive samples with the Allplex™ assay and 1 sample that was Allplex™ negative but Simplexa™ positive. Using a consensus result criterion as the reference standard allowed to resolve the eight samples with discordant results (one Allplex™ negative and seven Simplexa™ negative) as truly false negative. Interestingly, a Spearman's negative association was found between the viral RNA loads quantified by the Quanty assay and the CT values of RT PCRs performed with either the Allplex™ assay or the Simplexa™ assay. However, the strength of this association was higher for the Allplex™ assay (N gene, ρ = - 0.92; RdRP gene, ρ = - 0.91) than for the Simplexa™ assay (ORF1ab gene, ρ = - 0.65; S gene, ρ = - 0.80). The Allplex™ 2019-nCoV, the Simplexa™ COVID-19 Direct, and the Quanty COVID-19 assays yielded comparable results. However, the role these assays might play in future clinical practice warrants larger comparison studies.

Prevalence and Clonal Distribution of Azole-Resistant Candida parapsilosis Isolates Causing Bloodstream Infections in a Large Italian Hospital.

The most prevalent cause of nosocomial bloodstream infection (BSI) among non-C. albicans Candida species, Candida parapsilosis, may not only be resistant to azole antifungal agents but also disseminate to vulnerable patients. In this survey of BSIs occurring at a large Italian hospital between May 2014 and May 2019, C. parapsilosis accounted for 28.5% (241/844) of all Candida isolates causing BSI episodes. The majority of episodes (151/844) occurred in medical wards. Across the 5 yearly periods, the rates of azole non-susceptibility were 11.8% (4/34), 17.8% (8/45), 28.6% (12/42), 32.8% (19/58), and 17.7% (11/62), respectively, using the Sensititre YeastOne® method. Among azole non-susceptible isolates (54/241; 22.4%), 49 were available for further investigation. Using the CLSI reference method, all 49 isolates were resistant to fluconazole and, except one (susceptible dose-dependent), to voriconazole. Forty (81.6%) isolates harbored the Erg11p Y132F substitution and nine (18.4%) isolates the Y132F in combination with the Erg11p R398I substitution. According to their genotypes, as defined using a microsatellite analysis based on six short tandem repeat markers, 87.7% of isolates (43/49) grouped in two major clusters (II and III), whereas 4.1% of isolates (2/49) belonged to a separate cluster (I). Interestingly, all the isolates from cluster II harbored the Y132F substitution, and those from cluster III harbored both Y132F and R398I substitutions. Of 56 non-Italian isolates included as controls, two Indian isolates with the Y132F substitution had a genotype clearly differing from that of the isolates from clusters II and I. In conclusion, these findings show the dominance of clonal Y132F isolates in our hospital and suggest detection of the Y132F substitution as helpful tool to prevent transmission among hospitalized patients at risk of BSI.

Implementation of the eazyplex® CSF direct panel assay for rapid laboratory diagnosis of bacterial meningitis: 32-month experience at a tertiary care university hospital.

We aimed to report a 32-month laboratory experience with the eazyplex® CSF direct panel assay for the rapid diagnosis of meningitis due to six most common bacterial species (Escherichia coli, Haemophilus influenzae, Listeria monocytogenes, Neisseria meningitidis, Streptococcus agalactiae, and Streptococcus pneumoniae). We included all cerebrospinal fluid (CSF) samples from patients admitted with a clinical suspicion of meningitis/encephalitis between May 2016 and December 2018 at our hospital. In addition to the eazyplex® assay, both Gram stain microscopy and culture were performed, and results were confirmed with 16S rRNA PCR/sequencing. Patients' demographics and relevant clinical information were collected. Of 135 studied patients, 44 (32.6%) had a microbiologically documented diagnosis of meningitis. Overall, we identified 21 S. pneumoniae, 10 N. meningitidis, 6 L. monocytogenes, 3 E. coli, 2 Streptococcus pyogenes, 1 S. agalactiae, and 1 Citrobacter koseri as aetiological agents. The eazyplex® assay allowed identification in 40 (90.9%) cases, with four not identified cases due to microorganisms not included in the panel at the time of testing. Thirty-two (72.7%) cases had positive culture results, whereas 28 (63.6%) cases had positive Gram stain results. Notably, combining Gram stain and eazyplex® assay allowed identification in 100% of cases. After notification of rapid results, physicians modified the empiric antibiotic therapy, which became appropriate in three patients (all with L. monocytogenes meningitis). The eazyplex® CSF panel assay worked better than culture in detecting the most common agents of bacterial meningitis and accelerated the diagnosis leading to timely initiation or continuation of appropriate antibiotic therapy.

Incidence and antimicrobial resistance trends in bloodstream infections caused by ESKAPE and Escherichia coli at a large teaching hospital in Rome, a 9-year analysis (2007-2015).

The proportion of antimicrobial resistance (AMR) among the ESKAPE and Escherichia coli (ESKAPEEc) pathogens causing bloodstream infection (BSI) increased worldwide. We described longitudinal trends in ESKAPEEc BSI and AMR over 9 years (2007-2015) at a large teaching hospital in Italy. Of 9720 unique BSI episodes, 6002 (61.7%) were caused by ESKAPEEc pathogens. The majority of these episodes (4374; 72.9%) were hospital-onset infections. The most frequent pathogen was E. coli (32.8%), followed by Staphylococcus aureus (20.6%), Klebsiella pneumoniae (16.1%), and Pseudomonas aeruginosa (11.6%). There was a significant increase of hospital-onset K. pneumoniae (from 2.3 to 5.0 per 10,000 patient-days; P = 0.001) and community-onset E. coli (from 3.3 to 9. 1 per 10,000 emergency admissions; P = 0.04) BSIs. Among hospital-onset BSIs, increases of extended-spectrum ß-lactamase (ESBL)-producing E. coli (from 25.4 to 35.2%, P = 0.006), carbapenemase-producing K. pneumoniae (from 4.2 to 51.6%, P < 0.001), and methicillin-resistant S. aureus (from 33.9 to 44.4%, P < 0.001) BSIs were observed between the 2007-2009 and 2010-2012 study periods. In contrast, a decrease of BSIs caused by P. aeruginosa resistant to ceftazidime (from 45.5 to 28.2%, P < 0.001), ciprofloxacin (from 46 to 36.3%, P = 0.05), and meropenem (from 55 to 39.9%, P = 0.03) was observed through all 9 years of the study period. Among community-onset BSIs, increases of BSIs caused by ESBL-producing E. coli (from 28.6 to 42.2%, P = 0.002) and carbapenemase-producing K. pneumoniae (from 0 to 17.6%) were observed between the 2007-2009 and 2010-2012 study periods. Our findings show increased rates of BSI and relative AMR for specific pathogen-health care setting combinations, and call for continued active surveillance and infection control policies.

Comparison of Mycoplasma IES, Mycofast Revolution and Mycoplasma IST2 to detect genital mycoplasmas in clinical samples.

INTRODUCTION: Culture is regarded as the gold standard for the detection of genital mycoplasma in clinical samples. Commercially available diagnostic kits, based on liquid broth cultures, provide interesting alternatives to conventional culture. We assessed the laboratory performances of Mycoplasma IES (IES), the Mycofast Revolution (REV) and Mycoplasma IST 2 (IST2) compared to A7 agar plates for the detection of Ureaplasma urealyticum and Mycoplasma hominis in clinical samples. METHODOLOGY: From April to July 2013, endocervical or vaginal samples were collected from sexually active women with abnormal vaginal discharge. Each specimen was tested in parallel using the three commercial kits and the A7 agar plates. RESULTS: A total of 303 samples were included in this study, 35.6% (108/303) of which were positive on A7 plates. Sensitivities for the detection of U. urealyticum of IES, REV and IST2 were 100%, 96.2% and 95.3%, respectively while those for M. hominis were of 92.8%, 92.8% and 85.7%, respectively. Specificity was 100% for the 3 methods. Concerning antimicrobial susceptibility testing, full agreement between IES and REV was documented. CONCLUSIONS: The Mycoplasma IES kit was found to be equivalent or superior compared to other commercial culture-based assays for a rapid and accurate identification of U. urealyticum and M. hominis and detection of resistance. It might be considered a cost-effective tool for detection of these organisms, particularly attractive in developing countries.