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Abstract Background The efficacy of early treatment with convalescent plasma in patients with COVID-19 is debated. Nothing is known about the potential effect of other plasma components other than anti-SARS-CoV-2 antibodies. Methods To determine whether convalescent or standard plasma would improve outcomes for adults in early phase of Covid19 respiratory impairment we designed this randomized, three-arms, clinical trial (PLACO COVID) blinded on interventional arms that was conducted from June 2020 to August 2021. It was a multicentric trial at 19 Italian hospitals. We enrolled 180 hospitalized adult patients with COVID-19 pneumonia within 5 days from the onset of respiratory distress. Patients were randomly assigned in a 1:1:1 ratio to standard of care (n = 60) or standard of care + three units of standard plasma (n = 60) or standard of care + three units of high-titre convalescent plasma (n = 60) administered on days 1, 3, 5 after randomization. Primary outcome was 30-days mortality. Secondary outcomes were: incidence of mechanical ventilation or death at day 30, 6-month mortality, proportion of days with mechanical ventilation on total length of hospital stay, IgG anti-SARS-CoV-2 seroconversion, viral clearance from plasma and respiratory tract samples, and variations in Sequential Organ Failure Assessment score. The trial was analysed according to the intention-to-treat principle. Results 180 patients (133/180 [73.9%] males, mean age 66.6 years [IQR 57–73]) were enrolled a median of 8 days from onset of symptoms. At enrollment, 88.9% of patients showed moderate/severe respiratory failure. 30-days mortality was 20% in Control arm, 23% in Convalescent (risk ratio [RR] 1.13; 95% confidence interval [CI], 0.61–2.13, P = 0.694) and 25% in Standard plasma (RR 1.23; 95%CI, 0.63–2.37, P = 0.544). Time to viral clearance from respiratory tract was 21 days for Convalescent, 28 for Standard plasma and 23 in Control arm but differences were not statistically significant. No differences for other secondary endpoints were seen in the three arms. Serious adverse events were reported in 1.7%, 3.3% and 5% of patients in Control, Standard and Convalescent plasma arms respectively. Conclusions Neither high-titer Convalescent nor Standard plasma improve outcomes of COVID-19 patients with acute respiratory failure. Trial Registration Clinicaltrials.gov Identifier: NCT04428021. First posted: 11/06/2020

Abstract The aims of this study were to characterize new SARS-CoV-2 genomes sampled all over Italy and to reconstruct the origin and the evolutionary dynamics in Italy and Europe between February and June 2020. The cluster analysis showed only small clusters including

Abstract Background The objectives of the study were to investigate the organizational characteristics of acute care facilities worldwide in preventing and managing infections in surgery; assess participants’ perception regarding infection prevention and control (IPC) measures, antibiotic prescribing practices, and source control; describe awareness about the global burden of antimicrobial resistance (AMR) and IPC measures; and determine the role of the Coronavirus Disease 2019 pandemic on said awareness. Methods A cross-sectional web-based survey was conducted contacting 1432 health care workers (HCWs) belonging to a mailing list provided by the Global Alliance for Infections in Surgery. The self-administered questionnaire was developed by a multidisciplinary team. The survey was open from May 22, 2021, and June 22, 2021. Three reminders were sent, after 7, 14, and 21 days. Results Three hundred four respondents from 72 countries returned a questionnaire, with an overall response rate of 21.2%. Respectively, 90.4% and 68.8% of participants stated their hospital had a multidisciplinary IPC team or a multidisciplinary antimicrobial stewardship team. Local protocols for antimicrobial therapy of surgical infections and protocols for surgical antibiotic prophylaxis were present in 76.6% and 90.8% of hospitals, respectively. In 23.4% and 24.0% of hospitals no surveillance systems for surgical site infections and no monitoring systems of used antimicrobials were implemented. Patient and family involvement in IPC management was considered to be slightly or not important in their hospital by the majority of respondents (65.1%). Awareness of the global burden of AMR among HCWs was considered very important or important by 54.6% of participants. The COVID-19 pandemic was considered by 80.3% of respondents as a very important or important factor in raising HCWs awareness of the IPC programs in their hospital. Based on the survey results, the authors developed 15 statements for several questions regarding the prevention and management of infections in surgery. The statements may be the starting point for designing future evidence-based recommendations. Conclusion Adequacy of prevention and management of infections in acute care facilities depends on HCWs behaviours and on the organizational characteristics of acute health care facilities to support best practices and promote behavioural change. Patient involvement in the implementation of IPC is still little considered. A debate on how operationalising a fundamental change to IPC, from being solely the HCWs responsibility to one that involves a collaborative relationship between HCWs and patients, should be opened.

Strongyloides and cytomegalovirus co-infections are rarely reported, even though they are distinguished by high morbidity and mortality, especially in immunocompromised hosts. We narratively reviewed the literature on reported cases of Strongyloides and CMV co-infections in immunosuppressed patients. Most cases occurred in males with a median age of 47 (IQR, 37–59). Strongyloides/CMV co-infections occurred among immunocompromised hosts, especially in solid organ transplants and hematological or rheumatological diseases. Most of the patients underwent a course of steroid treatment before the diagnosis of co-infections. Other common immunomodulatory agents were tacrolimus and mycophenolate. The first clinical manifestations of co-infections were mainly gastrointestinal, followed by respiratory symptoms. CMV was, in most patients, co-infected with an isolated reactivation, although Strongyloides manifested especially as hyperinfection syndrome. Ganciclovir and ivermectin are the mainstays of CMV and Strongyloides treatment. However, the treatment mortality reported in this narrative review is around 52.4%. Interestingly secondary bacterial infections are common in CMV/Strongyloides-infected patients.

To our knowledge, we have described the first case of Strongyloides/Cytomegalovirus (CMV) concomitant infection that occurred in a European country. The patient was a 76-year-old woman affected by relapsed non-Hodgkin lymphoma who presented interstitial pneumonia with a rapidly progressive worsening of respiratory insufficiency, leading to cardiac dysfunction and consequent death. CMV reactivation is a common complication in immunocompromised patients, while hyperinfection/disseminated strongyloidiasis (HS/DS) is rare in low endemic regions, but has been widely described in Southeast Asia and American countries. HS and DS are two consequences of the failure of infection control by the immune system: HS is the uncontrolled replication of the parasite within the host and DS the spreading of the L3 larvae in organs other than the usual replication sites. Only a few cases of HS/CMV infection have been reported in the literature, and only in one patient with lymphoma as an underlying disease. The clinical manifestations of these two infections overlap, usually leading to a delayed diagnosis and a consequent poor outcome.

Abstract A growing number of emerging SARS-CoV-2 variants is being identified worldwide, potentially impacting the effectiveness of current vaccines. We report the data obtained in several Italian regions involved in the SARS-CoV-2 variant monitoring from the beginning of the epidemic and spanning the period from October 2020 to March 2021.

Genomic surveillance of SARS-CoV-2 is crucial to monitor the spread of variants of concern. A new sequencing method enables cost-effective SARS-CoV-2 genomic surveillance at scale and is easily adaptable to other viruses.

Bloodstream infections (BSI) are an increasing cause of admissions to hospitals. Non-hospital-acquired BSI are defined by blood cultures that are positive less than 48 hours after admission, but a relevant difference exists between community-acquired and healthcare-associated (HCA) BSI in terms of risk of multidrug resistance (MDR). We planned a retrospective study in three different cohorts in order to develop and to temporally and spatially validate an easy and rapid prognostic model for identifying MDR non-hospital-acquired (non-HA) BSI. The pathogens most involved in BSI are Staphylococcus spp. and Escherichia coli, responsible for about 75% of all MDR isolated. The model includes age, gender, long-term care facility admission, immunocompromise, any recent invasive procedures and central line placement, recent intravenous treatment and antibiotic treatment. It shows an acceptable performance, especially for intermediate probabilities of MDR infection, with a C-index of 70%. The model was proposed in a nomogram that could allow better targeting of antibiotic therapy for non-HA BSI admitted in hospital. However, it should be further validated to determine its applicability in other populations.

BackgroundIdentifying determinants of the novel severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) transmission in settings of contagion is fundamental to inform containment strategies. We assessed SARS-CoV-2 cycle threshold value (Ct) from the first diagnostic nasal–pharyngeal swab of symptomatic index cases and which demographic or clinical characteristics among cases and contacts are associated with transmission risk within households.MethodsThis is a retrospective prevalence study on secondary SARS-CoV-2 cases (SC) among the household contacts of symptomatic adult index cases randomly sampled from all the SARS-CoV-2-positive diagnostic nasopharyngeal swabs analyzed at our regional referral hospital (Amedeo di Savoia Hospital, Turin, Italy) in March, 2020. Index cases underwent a telephone survey to collect their demographic and clinical data and all their household contacts. The Ct value of RdRp gene from the first diagnostic swab of index cases was recorded and index cases were grouped according to Ct tertiles (A < first tertile, first ≤ B ≤ second tertile, C ≥ second tertile). Post hoc analysis was performed in SC as well as contacts that did not undergo SARS-CoV-2 testing but developed compatible signs and symptoms. Non-parametric tests and generalized linear models were run.ResultsIndex (n = 72) and contact (n = 164) median age was 54 (48–63) and 32 (20–56) years, respectively. A total of 60, 50, and 54 subjects were contacts of group A, B, and C index cases, respectively; 35.9% of contacts were SC. Twenty-four further subjects (14.6%) met the criteria for symptom-based likely positive SC. The secondary attack rate was 36.0% (28.6–43.4), assuming a mean incubation period of 5 days and a maximum infectious period of 20 days. SC prevalence differed between Ct groups (53.3% A, 32.0% B, 20.4% C; p < 0.001). No difference in SC was found according to sex, presence of signs/symptoms, and COVID-19 severity of index cases, or according to contacts’ sex and number per household. The age of both index cases [aOR 4.52 (1.2–17.0) for 60 vs. ≤45 years old] and contacts [aOR 3.66 (1.3–10.6) for 60 vs. ≤45years old] and the Ct of the index [aOR 0.17 (0.07–0.4) for Ct ≥ 31.8 vs. Ct < 24.4] independently associated with SC risk. Sensitivity analysis including symptoms-based likely positive SC supported all the previous results.ConclusionIn confined transmission settings such as households, PCR Ct values may inform on the contagiousness of infected subjects and age may modulate transmission/contagion risk.

In the context of SARS-CoV-2 pandemic, rapid and easy-to-perform diagnostic methods are essential to limit the spread of the virus and for the clinical management of COVID-19 patients. Although real-time polymerase chain reaction remains the “gold standard” to diagnose acute infections, this technique is expensive, requires trained personnel, well-equipped laboratory and is time-consuming. A prospective evaluation of the Abbott ID NOW COVID-19 point-of-care testing that uses isothermal nucleic acid amplification for the qualitative detection of SARS-CoV-2 RdRp gene was run in the Emergency Department during the third wave of COVID-19 pandemic. ID-NOW significantly simplified SARS-CoV-2 identification and COVID-19 patient triaging, being highly valuable in rapidly locating febrile patients in or out of COVID-19 areas, and can be considered as a first-line diagnostic test in the Emergency Room setting.

The Usutu virus (USUV) has recently attracted the attention of scientists because of its rapid spread across Europe and its growth over the previous seasons in Italy. Here, we describe the first case of USUV infection in Asti, Piedmont region, Italy. The patient remained asymptomatic in the acute phase and during the early follow-up, despite a mild increase in liver enzymes. The prompt diagnosis in this patient was due to positive qualitative PCR for WNV blood-donor screening with negative RT-PCR of WNV and positive USUV-RNA following the confirmation test. Blood-donor screening and transmission risk monitoring are pivotal in following the spread of this Flavivirus in non-endemic countries, due to the high percentage of asymptomatic carriers.

We measured severe acute respiratory syndrome coronavirus 2 spike protein subunits S1/S2 antibodies by using capillary electrophoresis and a chemiluminescence immunoassay for 5,444 active healthcare workers in Italy. Seroprevalence was 6.9% and higher among participants having contact with patients. Seroconversion was not observed in 37/213 previously infected participants.

Background: Emerging evidence supports the “variolation hypothesis” in severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), but the derivative idea that the viral load of index cases may predict disease severity in secondary cases could be unsubstantiated. We assessed whether the prevalence of symptomatic infections, hospitalization, and deaths in household contacts of 2019 novel coronavirus disease (COVID-19) cases differed according to the SARS-CoV-2 PCR cycle threshold (Ct) from nasal-pharyngeal swab at diagnosis of linked index cases.Methods: Cross-sectional study on household contacts of COVID-19 cases randomly sampled from all the infections diagnosed in March at our Microbiology Laboratory (Amedeo di Savoia, Turin). Data were retrospectively collected by phone interviews and from the Piedmont regional platform for COVID-19 emergency. Index cases were classified as high (HVl) and low viral load (LVl) according to two exploratory cut-offs of RdRp gene Ct value. Secondary cases were defined as swab confirmed or symptom based likely when not tested but presenting compatible clinical picture.Results: One hundred thirty-two index cases of whom 87.9% symptomatic and 289 household contacts were included. The latter were male and Caucasian in 44.3 and 95.8% of cases, with a median age of 34 years (19–57). Seventy-four were swab confirmed and other 28 were symptom based likely secondary cases. Considering both, the contacts of HVl and LVl did not differ in the prevalence of symptomatic infections nor COVID-19-related hospitalization and death. No difference in median Ct of index cases between symptomatic and asymptomatic, hospitalized and not hospitalized, or deceased and survived secondary cases was found. Negative findings were confirmed after adjusting for differences in time between COVID-19 onset and swab collection of index cases (median 5 days) and after removing pediatric secondary cases.Conclusions: The amount of SARS-CoV-2 of the source at diagnosis does not predict clinical outcomes of linked secondary cases. Considering the impelling release of assays for SARS-CoV-2 RNA exact quantification, these negative findings should inform clinical and public health strategies on how to interpret and use the data.

Evidence for the real impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection on preterm birth is unclear, as available series report composite pregnancy outcomes and/or do not stratify patients according to disease severity. The purpose of the research was to determine the real impact of asymptomatic/mild SARS-CoV-2 infection on preterm birth not due to maternal respiratory failure. This case-control study involved women admitted to Sant Anna Hospital, Turin, for delivery between 20 September 2020 and 9 January 2021. The cumulative incidence of Coronavirus disease-19 was compared between preterm birth (case group, n = 102) and full-term delivery (control group, n = 127). Only women with spontaneous or medically-indicated preterm birth because of placental vascular malperfusion (pregnancy-related hypertension and its complications) were included. Current or past SARS-CoV-2 infection was determined by nasopharyngeal swab testing and detection of IgM/IgG antibodies in blood samples. A significant difference in the cumulative incidence of Coronavirus disease-19 between the case (21/102, 20.5%) and the control group (32/127, 25.1%) (P= 0.50) was not observed, although the case group was burdened by a higher prevalence of three known risk factors (body mass index > 24.9, asthma, chronic hypertension) for severe Coronavirus disease-19. Logistic regression analysis showed that asymptomatic/mild SARS-CoV-2 infection was not an independent predictor of spontaneous and medically-indicated preterm birth due to pregnancy-related hypertension and its complications (0.77; 95% confidence interval, 0.41-1.43). Pregnant patients without comorbidities need to be reassured that asymptomatic/mild SARS-CoV-2 infection does not increase the risk of preterm delivery. Preterm birth and severe Coronavirus disease-19 share common risk factors (i.e., body mass index > 24.9, asthma, chronic hypertension), which may explain the high rate of indicated preterm birth due to maternal conditions reported in the literature.

Several important sex and gender differences in the clinical manifestation of diseases have been known for a long time but are still underestimated. The infectious Coronavirus 2019 disease pandemic has provided evidence of the importance of a sex and gender-based approach; it mainly affected men with worse symptomatology due to a different immune system, which is stronger in women, and to the Angiotensin-converting enzyme 2 and Transmembrane protease serine 2 roles which are differently expressed among the sexes. Additionally, women are more inclined to maintain social distance and smoke less. Analysis of data on the infectious Coronavirus 2019 disease testing from people admitted to the Amedeo di Savoia Hospital, a regional referral center for infectious diseases, has been applied to the whole of 2020 data (254,640 records). A high percentage of data in the dataset was not suitable due to a lack of information or entering errors. Among the suitable samples, records have been analyzed for positive/negative outcomes, matching records for unique subjects (N = 123,542), to evaluate individual recurrence of testing. Data are presented in age and sex-disaggregated ways. Analyses of the suitable sample also concerned the relation between testing and hospital admission motivation and symptoms. Our analysis indicated that a sex and gender-based approach is mandatory for patients and the National Health System’s sustainability.

Abstract Background Cryptococcal infections, besides being a problem for immunocompromised patients, are occasionally being a problem for immunocompetent patients. In addition, the lower susceptibility of this yeast to azoles is a growing problem in health care. To date, there are very few molecules with any activity towards Cryptococcus neoformans, leading to heightened interest in finding new alternatives or adjuvants to conventional drugs for the treatment of mycosis caused by this yeast. Since the essential oils (EOs) are considered as a potential rich source of bioactive antimicrobial compounds, we evaluated the antifungal activity of Origanum vulgare (oregano), Pinus sylvestris (pine), and Thymus vulgaris (thyme red) EOs, and their components (α-pinene, carvacrol, thymol) compared with fluconazole, itraconazole, and voriconazole, against C.neoformans clinical strains. Then, we investigated the effect of EOs and components in combination with itraconazole. Methods EO composition was analysed by Gas chromatography-mass spectrometry (GC-MS). A broth microdilution method was used to evaluate the susceptibility of C.neoformans to azoles, EOs and components. Checkerboard tests, isobolograms and time-kill assays were carried out for combination studies. Results Six C.neoformans isolates were susceptible to azoles, while one C.neoformans exhibited a reduced susceptibility to all tested azole drugs. All EOs exerted a good inhibitory activity against all C.neoformans strains. Pine EO was the most effective. Among components, thymol exerted the most remarkable activity. By checkerboard testing and isobolographic analysis, combinations of itraconazole with oregano, pine, or thyme EOs, and carvacrol were found to be synergistic (FICI≤0.5) against azole susceptible C.neoformans. Regarding the azole not susceptible C.neoformans strain, the synergistic effect with itraconazole was observed with thyme EO (chemotype: thymol 26.52%; carvacrol 7.85%), and carvacrol. Time-kill assays confirmed the synergistic effects of itraconazole and oregano or thyme EO against azole susceptible C.neoformans. Binary mixtures of itraconazole/thyme EO or carvacrol yielded additive effects on the azole not susceptible C.neoformans. Conclusions Our findings highlight the potential effectiveness of thyme, oregano EOs, and carvacrol as natural and cost-effective adjuvants when used in combination with itraconazole. Identification of EOs exerting these effects could be one of the feasible ways to overcome drug resistance, reducing drug concentration and side effects.

We evaluated the clinical protection of BNT162b2 mRNA vaccine in healthcare workers (HCWs) and how COVID-19 manifestations and contagiousness change as the time since first dose increases. A matched (1:2 ratio) parallel cohort study was performed. During the first three months of vaccination campaign, HCWs of the entire health district ASL Città di Torino (Turin, Italy) were classified according to SARS-CoV-2-positivity in respect of the vaccination schedule: post-first-dose (fHCWs, n = 110), no difference was observed in fHCWs, while PHCWs and THCWs showed higher prevalence of asymptomatic infections, fewer signs/symptoms with a milder systemic involvement, and significantly higher Ct values (PHCWs 30.3 (24.1–35.5) vs. 22.3 (19.6–30.6), p = 0.023; THCWs 35.0 (31.3–35.9) vs. 22.5 (18.2–30.6), p = 0.024). Duration of symptoms was also shorter in THCWs (5 days (3–6) vs. 9 (7–14), p = 0.028). A linear increase of 3.81 points in Ct values was observed across the groups by vaccination status (p = 0.001) after adjusting for age, sex, comorbidities, and time between COVID-19 onset and swab collection. BNT162b2 decreased the risk of PCR-confirmed infections and severe disease, and was associated with a virologic picture of lesser epidemiologic concern as soon as 12 days after the first vaccine dose.

The epidemiology of yeast infections and resistance to available antifungal drugs are rapidly increasing, and non-albicans Candida species and rare yeast species are increasingly emerging as major opportunistic pathogens. In order to identify new strategies to counter the threat of antimicrobial resistant microorganisms, essential oils (EOs) have become an important potential in the treatment of fungal infections. EOs and their bioactive pure compounds have been found to exhibit a wide range of remarkable biological activities. We investigated the in vitro antifungal activity of nine commercial EOs such as Thymus vulgaris (thyme red), Origanum vulgare (oregano), Lavandula vera (lavender), Pinus sylvestris (pine), Foeniculum vulgare (fennel), Melissa officinalis (lemon balm), Salvia officinalis (sage), Eugenia caryophyllata (clove) and Pelargonium asperum (geranium), and some of their main components (α-pinene, carvacrol, citronellal, eugenol, γ-terpinene, linalool, linalylacetate, terpinen-4-ol, thymol) against non-albicans Candida strains and uncommon yeasts. The EOs were analyzed by GC-MS, and their antifungal properties were evaluated by minimum inhibitory concentration and minimum fungicidal concentration parameters, in accordance with CLSI guidelines, with some modifications for EOs. Pine exhibited strong antifungal activity against the selected non-albicans Candida isolates and uncommon yeasts. In addition, lemon balm EOs and α-pinene exhibited strong antifungal activity against the selected non-albicans Candida yeasts. Thymol inhibited the growth of all uncommon yeasts. These data showed a promising potential application of EOs as natural adjuvant for management of infections by emerging non-albicans Candida species and uncommon pathogenic yeasts.

Despite the substantial changes resulting from the introduction of combination antiretroviral therapy (cART), the prevalence of HIV-associated neurocognitive disorders (HAND) remains substantial. Blood–brain barrier impairment (BBBi) is a frequent feature in people living with HIV (PLWH) and it may persist despite effective antiretroviral treatment. A cross-sectional study was performed in PLWH who underwent lumbar puncture for clinical reasons or research protocols and several cerebrospinal fluid biomarkers were studied. BBBi was defined as cerebrospinal fluid-to-serum albumin ratio (CSAR) >6.5 (8 (>40 years). We included 464 participants: 147 cART-naïve and 317 on cART. Male sex was prevalent in both groups (72.1% and 72.2% respectively); median age was 44 (38–52) years in naïve and 49 (43–57) years in treated subjects. BBBi was observed in 35.4% naïve and in 22.7% treated participants; the use of integrase inhibitors was associated with a lower prevalence (18.3 vs. 30.9%, p = 0.050). At multivariate binary logistic regression (including age and sex) nadir CD4 cell count (p = 0.034), presence of central nervous system (CNS) opportunistic infections (p = 0.024) and cerebrospinal fluid (CSF) HIV RNA (p = 0.002) in naïve participants and male sex (p = 0.021), a history of CNS opportunistic infections (p = 0.001) and CSF HIV RNA (p = 0.034) in treated patients were independently associated with BBBi. CSF cells and neopterin were significantly higher in participants with BBBi. BBBi was prevalent in naïve and treated PLWH and it was associated with CSF HIV RNA and neopterin. Systemic control of viral replication seems to be essential for BBB integrity while sex and treatment influence need further studies.

To date, there is no severe acute respiratory syndrome coronavirus 2-(SARS-CoV-2)-specific prognostic biomarker available. We assessed whether SARS-CoV-2 cycle threshold (Ct) value at diagnosis could predict novel CoronaVirus Disease 2019 (COVID-19) severity, clinical manifestations, and six-month sequelae. Hospitalized and outpatient cases were randomly sampled from the diagnoses of March 2020 and data collected at 6 months by interview and from the regional database for COVID-19 emergency. Patients were stratified according to their RNA-dependent-RNA-polymerase Ct in the nasopharyngeal swab at diagnosis as follows: Group A ≤ 20.0, 20.0 < group B ≤ 28.0, and Group C > 28.0. Disease severity was classified according to a composite scale evaluating hospital admission, worst oxygen support required, and survival. Two hundred patients were included, 27.5% in Groups A and B both, 45.0% in Group C; 90% of patients were symptomatic and 63.7% were hospitalized. The median time from COVID-19 onset to swab collection was five days. Lethality, disease severity, type, and number of signs and symptoms, as well as six-month sequelae distributed inversely among the groups with respect to SARS-CoV-2 Ct. After controlling for confounding, SARS-CoV-2 Ct at diagnosis was still associated with COVID-19-related death (p = 0.023), disease severity (p = 0.023), number of signs and symptoms (p < 0.01), and presence of six-month sequelae (p < 0.01). Early quantification of SARS-CoV-2 may be a useful predictive marker to inform differential strategies of clinical management and resource allocation.

This prospective observational study aimed to evaluate whether women with SARS-CoV-2 infection during the first trimester of pregnancy are at higher risk of noninvasive prenatal screening test alterations and/or of congenital fetal anomalies at the second-trimester fetal anatomy scan. Maternal symptoms were secondly investigated. The study was carried out on 12-week pregnant women admitted for noninvasive prenatal testing (16 April and 22 June 2020). The cohort had seromolecular tests for SARS-CoV-2, after which they were divided into a positive case group and a negative control group. Both groups had 20-week ultrasound screening. Seventeen out of the 164 women tested positive for SARS-CoV-2 (10.3%). There were no significant differences in mean nuchal translucency thickness or biochemical markers (pregnancy-associated plasma protein A, alpha-fetoprotein, human chorionic gonadotropin, unconjugated estriol) between cases and controls (p = 0.77, 0.63, 0.30, 0.40, 0.28) or in the fetal incidence of structural anomalies at the second-trimester fetal anatomy scan (p = 0.21). No pneumonia or hospital admission due to COVID-19-related symptoms were observed. Asymptomatic or mildly symptomatic SARS-CoV-2 infection during the first trimester of pregnancy did not predispose affected women to more fetal anomalies than unaffected women. COVID-19 had a favorable maternal course at the beginning of pregnancy in our healthy cohort.

During June 9–September 30, 2015, five cases of louseborne relapsing fever were identified in Turin, Italy. All 5 cases were in young refugees from Somalia, 2 of whom had lived in Italy since 2011. Our report seems to confirm the possibility of local transmission of louse-borne relapsing fever.

Primary study outcome was absence of treatment failure (virological failure, VF, or treatment interruption) per protocol at week 48.Patients on 3-drug ART with stable HIV-1 RNA

Neisseria gonorrhoeae (NG) antimicrobial susceptibility trends to azithromycin, cefixime and ceftriaxone were analyzed, from 2009 to 2016, to monitor changing antimicrobial susceptibility concomitant with the change in prescribing practice in 2012 from cefixime, or ceftriaxone, to ceftriaxone plus azithromycin. Patient characteristics predictive to be infected by antibiotic resistant N. gonorrhoeae were estimated. Finally, the protocol for the treatment of gonorrhoea, in comparison with the international guidelines, was also evaluated.Data on NG antimicrobial resistance were obtained from a network of sexually transmitted diseases clinics and other laboratories in 12 cities in Italy. We tested the 1,433 gonococci for antimicrobial susceptibility to azithromycin, cefixime and ceftriaxone using a gradient diffusion method. Logistic-regression methods with cluster robust standard errors were used to investigate the association of resistance categories with demographic and clinical patient characteristics and to assess changes in prescribing practices. To minimize bias due to missing data, all statistical models were fitted to data with forty rounds of multiple imputation, using chained equations.The percentage of isolates resistant to cefixime was 17.10% in 2009 and declined up to 1.39% in 2016; at the same time, those resistant to azithromycin was 23.68% in 2009 and 3.00% in 2012. Starting from 2013, azithromycin resistant gonococci tended to increase up to 7.44% in 2016. No ceftriaxone resistant isolates were observed. By multivariate analysis, the men who have sex with women (MSW) and women had a proportional adjusted OR of resistance of 1.25 (95%CI: 0.90; 1.73) and 1.67 (95%CI: 1.16; 2.40), respectively, in comparison with men who have sex with men (MSM). An aOR of resistance of 0.48 (95%CI: 0.21; 1.12) among NG isolated in the pharynx, compared with those isolated in genital sites, was calculated. The proportional aOR of resistance was 0.58 (95%CI: 0.38; 0.89) for presence vs absence of co-infection and 2.00 (95%CI: 1.36; 2.96) for past history vs no history of gonorrhoea.Finally, at least for the period 2013-2016, the older, subjects with anorectal or pharyngeal gonorrhoea infection, subjects with a co-infection, subjects with a previous gonorrhoea infection were not always correctly treated.Overall, our findings suggest the shifts in N. gonorrhoeae susceptibility to cefixime and azithromycin in the time frame period. First of all, the increasing rate of azithromycin resistance in 2015-2016 in NG isolated in the country need to be monitor in the future. Finally, extensive information on treatment regimens may be useful to asses treatment adherence particularly for the older subjects, subjects with an anorectal or pharyngeal infection, subjects with a co-infection and subjects with a previous history of gonorrhoea. Gonorrhoea treatment strategy should be based on the evidence obtained by the local antimicrobial surveillance system and data about treatment failures.

In high risk patients such as in eldery, newborns and immunosuppressed subjects, a timely diagnosis of influenza is required for the most appropriate antiviral strategy in order to avoid severe secondary respiratory complications and viral spreading. Influenza is preventable by vaccination and chemoprophylaxis and is treatable by specific antiviral indications. The need for a timely diagnosis has led to the introduction of numerous rapid diagnostic tests.These are mostly antigen detection test giving results within 30 minutes, a clinically relevant time-frame to complement with the use of antiviral medications or chemoprophylaxis strategy. When evaluating performances of rapid test for influenza viruses, it is important to consider the type and quality of specimen to be tested, as well as sensitivity and specificity of the assays. Nasal/nasopharyngeal swabs are the most frequently submitted specimens, but nasal/nasopharingeal aspirates and washs can improve the diagnostic sensitivity of the test. Only some rapid assays can be successful used with broncoalveolar washings. In this review,we evaluated the sensitivity, specificity, reproducibility and feasibility of the most currently licensed rapid tests for influenza virus A and B. A flow-chart for the laboratory diagnosis of influenza with rapid test in combination with confirmatory test is proposed.

Nucleic acid detection by amplification (NAT) assays began to be used in diagnostic microbiology about twenty years ago. Since then, the progress of molecular methods has been continuous and rapid, aimed to obtain more and more sensitive and specific results and automation.To actually achieve such objectives, microbiology laboratories have to use suited quality controls and to participate to external quality control programs. Quality controls for NAT assays need to be adjusted to follow the evolution of the molecular methods.The need of standardization and suited controls for NAT assays became evident approximately by the middle of ’90 years.Thus International Standards (IS) were established by the World Health Organization, for the main blood borne viruses. Such preparations had a well known concentration expressed as International Units, a new measure unit for nucleic acid quantitation. Then, IS were used in order to calibrate working reagents and synthetic calibrators. Another tool for standardization are multicentre studies aimed to verify the performances of participating laboratories using different molecular methods, and allow the comparison of inter-laboratory results.The CoSBio of AMCLI, as other European organizations, in the last years promoted, some multicentre studies on NAT assays. The results of such studies demonstrated a good level of diagnostic performances of participating laboratories. However, with the introduction of the real time PCR (RQ-PCR) a new problem emerged: the variability of results obtained by molecular methods due to different coefficient of linear amplification. Solutions suggested by Literature to this and similar problems are examined, mainly focusing on the assessment of the Calibration curves in the RQ-PCR. Finally, the Authors signal some important fields in the molecular diagnostics, such as the detection of HPV and the detection of BKV, still lacking of sufficient standardization.

Success of antiviral therapy for chronic hepatitis B is supported by highly sensitive PCR-based assays for Hepatitis B virus (HBV) DNA. Nucleic acid extraction from biologic specimens is technically demanding and reliable PCR results depend it. Performances of the fully automatic system COBAS AmpliPrep™/COBAS TaqMan™ 48 (CAP/CTM) (Roche, Branchburg, NJ) for HBV DNA extraction and real -time PCR quantification were assessed and compared with the end-point PCR COBAS AMPLICOR HBV Monitor (CAHBM, Roche). Analytical evaluation with a proficiency panel showed that CAP/CTM quantitated HBV DNA levels in one single run over a wide dynamic range (7 logs) with a close correlation between expected and observed values (r=0.976, interassay variability below 5%). Clinical evaluation as tested with samples from 92 HBsAg-positive patients, demonstrated excellent correlation with CAHBM (r=0.966, mean difference in quantitation: 0.36 log10 IU/ml). CAP/CTM detected 10% more viremic patients and longer period of residual viremia in those on therapy. In lamivudine (LAM)-resistant patients, reduction of HBV DNA after 12 months of Adefovir (ADF) was higher in the combination (LAM+ADF) schedule than in ADF monotherapy (5.1 vs. 3.5 logs) suggesting a benefit in continuing LAM. In conclusion,CAP/CTM can improve the management of HBV infection, the assessment of antiviral therapy and drug resistance, supporting further insights in the emerging area of drug resistance.

Background/Aims: Genotyping and subtyping of hepatitis C virus (HCV) is epidemiologically and clinically relevant to prognosis and therapeutical management of HCV infection.Aim was to study the feasibility of an “in house” direct sequencing method for the genotyping and subtyping of HCV strains compared with a commercially available genotyping assay (Inno-LiPA,VERSANT® HCV Genotype 2.0 Assay, Bayer). Methods: 74 clinical plasma samples cross-representing HCV genotypes 1 to 5 typed with the line probe assay Inno-LiPA (LiPA) were subjected to a laboratory-developed 5’UTR direct sequencing protocol (5’UTR Seq).A NS5B direct sequencing protocol (NS5B Seq) was apply to 23/74 samples (17 negative by the 5’UTR).Two libraries of 5’UTR and NS5B HCV prototypes were constructed; BioEdit 7.0.0 and ClustalW were used for alignments and phylogenetic analysis. Results: 5’UTR Seq typed 47/74 LiPA positive samples (64%). Concordance between the two assays was 91% (43/47) and 72% (31/43) for HCV genotyping and subtyping, respectively. LiPA and 5’UTR Seq discordant samples were typed by NS5B Seq. NS5B Seq typed 13/17 samples negative with 5’UTR Seq.All genotypes and subtypes detected with NS5B Seq were concordant with LiPA. 5’UTR and NS5B direct sequencing pointed out a wider HCV subtype distribution than LiPA for genotype 1, 2 and 4. By the combination of 5’UTR and NS5B direct sequencing, 81% (60/74) of LiPA positive samples were typed. Conclusion: HCV typing and subtyping with the line probe assay Inno-LiPA is highly recommended for clinical purposes.A more detailed HCV typing for epidemiologic purposes can by achieved by the direct sequencing of a region with a moderate degree of genetic variability such as NS5B. HCV NS5B analysis is more efficient in resolving viral strain genetic variability than direct sequencing of a highly conserved region such as 5’UTR.

Quantitave Polymerase Chain Reaction (PCR) for Cytomegalovirus (CMV) DNA provides highly sensitive and specific data for detecting CMV as well as monitoring the infection and determining the appropriate antiviral strategy.To determine the clinical application of a recently introduced real-time (RT) PCR assay for CMV DNA quantitation in peripheral blood leukocytes (PBLs) and defining its correlation with the commercial quantitative end-point (EP) PCR method COBAS AMPLICOR CMV Monitor and pp65 antigen test. Sequential PBL samples (n=158) from 32 liver transplanted patients with CMV asymptomatic infection and positive for CMV DNA by EP-PCR were retrospectively analysed with RT-PCR and studied according to pp65 antigen levels. A good correlation was found between RT-PCR and pp65 antigen test (r=0.691) and between the two PCR assays (r=0.761). RT-PCR data were significantly higher in pre-emptive treated patients (those with >20 pp65+positive cells, median value: 3.8 log10 copies/500,000 PBLs) than in not-treated ones (2.9 logs).According to pp65 levels of 0, 1-10, 11-20, 21-50, 51-100 and >100 positive cells/200,000 PBLs, median CMV DNA load by RT-PCR was 2.6, 3.0, 3.6, 4.0. 4.2 and 4.8, log10 copies/ 500,000 PBLs, respectively (EP-PCR CMV DNA levels: 2. 8, 2.9, 3.8, 3.7, 3.9 and 4.0 logs). For samples with >20 pp65+cells, that is above the level at which pre-emptive therapy was started, RT-PCR values were significantly higher than in groups with less than 20 pp65+cells, whereas EP-PCR values did not significantly differ and showed a slower progression rate. Dilutions of DNA from CMV AD169 strain were used to probe RT-PCR reproducibility (between and intra-assay variability < 2%) and sensitivity (100% detection rate at 10 copies/reaction, 28.5% with EP-PCR). A significant improvement is coming from the introduction of RT-PCR to the study of CMV DNA dynamics in differently CMV infected patients due to a more reliable quantitation of CMV DNA for moderate and high DNA level compared to EP-PCR with better sensitivity and specificity. RTPCR gives more precise informations on viral load kinetics for evaluating the infection progress and assessing antiviral response, significantly simplifying and accelerating the process of producing a reliable quantification of CMV DNA for clinical purposes.