Legionella pneumophila is ubiquitous in aquatic environments and responsible for severe pneumonia in humans through inhalation of aerosol containing Legionella spp. Macrolides and fluoroquinolones are frequently used antimicrobials, but treatment failures are increasingly being reported. As susceptibility testing is not routinely performed, this study aimed to determine the minimum inhibitory concentrations (MICs) on 58 environmental Legionella pneumophila strains (24 of serogroup 1 and 34 of non-serogroup 1) isolated in Northern Italy. MICs of azithromycin, erythromycin, ciprofloxacin, levofloxacin, and rifampicin were determined by the microdilution method using buffered yeast extract broth supplemented with α-ketoglutarate (BYEα). Seventy-five percent of Legionella pneumophila isolates showed MIC values below the tentative highest MICs indicated by the European Committee on Antimicrobial Susceptibility Testing (EUCAST); rifampicin was the most active agent with MIC90 values below 0.008 mg/L. Interestingly, one isolate was tested and found to be PCR-positive for the azithromycin LpeAB active efflux system, further confirmed by the reserpine/resazurin microtiter assay. In conclusion, this study has provided additional susceptibility data for environmental Legionella pneumophila isolates from Northern Italy demonstrating, in general, low MICs values for the tested antimicrobials, although one strain tested was shown to possess the LpeAB resistance determinant, indicating that future surveillance studies are warranted.

The accuracy of available HPV molecular assays on self-samples needs to be evaluated as compared to clinician-collected samples. This pilot study aimed to investigate the BD Onclarity™ HPV assay on vaginal and first-void urine samples. Sixty-four women referred to colposcopy for cervical dysplasia performed a vaginal self-collection and provided a first-void urine sample, after informed consent. A cervical specimen was collected during the clinician examination. All samples were tested using BD Onclarity™ HPV assay on the BD Viper™ LT System. Overall positive agreement (OPA) between cervical and self-sample results was evaluated using Cohen’s kappa value (κ). Using a clinical cut-off of 38.3 Ct for HPV 16 and 34.2 Ct for other HR genotypes, compared to cervical sample, the self-collected vaginal sample OPA was 85.9%, and κ = 0.699. Without a clinical cut-off, the OPA was 95.3%, and the κ = 0.890. Data obtained comparing cervical and urine samples showed an OPA of 87.5% with a κ = 0.79 using a clinical cut-off, and an OPA of 90.6% with a κ = 0.776 without a clinical cut-off. Data showed a substantial agreement between both self-collected and clinician-collected samples. A specific clinical cut-off analysis should be considered based on type of sample analysed.

Human Papillomavirus (HPV) testing on self-collected samples allows for improved coverage rates of cervical cancer (CC) screening programs. ThinPrep®PreservCyt® (HOLOGIC®, USA) medium is widely used for the suspension of cervical and vaginal self-samples. However, this medium is costly, toxic, and flammable, involving special handling procedures which make its use difficult in screening programs, particularly in low- and middle-income countries. This pilot study aimed to evaluate the analytical performance of eNat® (Copan SpA), an alternative non-alcohol-based suspension medium, compared to ThinPrep®PreservCyt® (HOLOGIC®) for high-risk HPV (hrHPV) detection in vaginal self-collected swabs using three different real-time polymerase chain reaction (RT-PCR) HPV assays: Anyplex™II HPV28 (Seegene, Korea), Papilloplex® High Risk HPV (GeneFirst, UK), and HPV OncoPredict (Hiantis, Italy). 30 women, referred to colposcopy, were enrolled in this observational, prospective pilot study and asked to collect two vaginal self-taken samples, which were suspended in 5 mL of ThinPrep®PreservCyt® or eNat®. Nucleic acids were extracted from 200 μL using Microlab Nimbus platform (Seegene, Korea) and tested with the three different RT-PCR full-genotyping high-risk HPV assays. The HPV results of vaginal samples resuspended in the two different media were compared to those obtained from the reference clinician-collected cervical sample from the same woman. hrHPV detection in vaginal self-samples suspended in both media demonstrated a substantial agreement with cervical samples with the three assays under-investigation (0.667<k<0.796). Moreover, the discordances between vaginal self-samples collected from the same woman were found only in cases of normal cytology or low-grade cytological lesions and were generally related to low hrHPV viral loads as indicated by the quantitative HPV OncoPredict assay (6.24E+02 copies/10,000 cells). The study's preliminary findings demonstrated a very good agreement between cervical and vaginal self-collected samples suspended in ThinPrep®PreservCyt® and eNat®, suggesting that the latter could represent a good alternative medium in HPV screening programs based on self-collection.

The collection and storage of water-related matrices such as biofilm from collection to processing are critical for the detection of Legionella pneumophila by cultural and molecular tests. SRK™ is a liquid medium that acts both as an antimicrobial neutralizing agent and a transport medium for bacterial culture enumeration and is useful to maintain the stability of the sample from collection to analysis. The aims of this study were to evaluate Legionella pneumophila viability and bacterial nucleic acids’ stability in SRK™ medium over time at different storage conditions. Artificial bacterial inoculates with an approximate concentration of 104, 103 and 102 CFU/mL were made using Legionella pneumophila certified reference material suspended in SRK™ medium. Bacteria recovery was analyzed by cultural and molecular methods at time 0, 24 and 48 h at room temperature and at 0, 24, 48 and 72 h at 2–8 °C, respectively. SRK™ medium supported Legionella pneumophila culture viability with CFU counts within the expected range. The recovery after 72 h at 2–8 °C was 83–100% and 75–95% after 48 h at room temperature. Real-time PCR appropriately detected Legionella pneumophila DNA at each temperature condition, dilution and time point. Results demonstrated a good performance of SRK™ medium for the reliable recovery of environmental Legionella.

Human papillomavirus (HPV) assays used in cervical cancer screening should be clinically validated according to international criteria. OncoPredict HPV® Screening (SCR) is a partial genotyping multiplex real‐time PCR assay targeting E6/E7 genes of 13 high‐risk (hr) HPVs. OncoPredict HPV® SCR (index assay) identifies HPV‐16 and HPV‐18 separately, 11 other hrHPV in aggregate and includes quality controls for sample adequacy, DNA extraction efficiency and PCR inhibition. 1300 VALGENT‐2 study samples (from women aged 20–60 attending the Scottish cervical cancer screening program) were tested with the index assay and the GP5+/6+ PCR enzyme immunoassay (standard comparator assay). Non‐inferior accuracy detecting cervical intraepithelial neoplasia of grade 2 or worse (CIN2+) of the index versus comparator was verified. Intra‐ and interlaboratory reproducibility of the index was evaluated by overall concordance and Cohen's kappa, using a sub‐population (n = 526). Relative sensitivity and specificity for CIN2+ of the index versus comparator were 1.01 (95% confidence interval [CI]: 0.99–1.03) and 1.02 (95% CI: 1.0–1.04), respectively. Noninferiority p values were all ≤0.05, except for CIN3+ in patients ≥30 years. Excellent intra‐ and interlaboratory reproducibility was shown with concordance >98% and kappas >0.95. OncoPredict HPV® SCR fulfills the three international validation criteria for hrHPV DNA tests in cervical cancer screening.