Safety and effectiveness of fifth generation cephalosporins for the treatment of methicillin-resistant staphylococcus aureus bloodstream infections: a narrative review exploring past, present, and future.

INTRODUCTION: Methicillin-resistant Staphylococcus aureus (MRSA) bloodstream infection (BSI) is a major issue in healthcare, since it is often associated with endocarditis or deep site foci. Relevant morbidity and mortality associated with MRSA-BSIs forced the development of new antibiotic strategies; in particular, this review will focus the attention on fifth-generation cephalosporins (ceftaroline/ceftobiprole), that are the only ß-lactams active against MRSA. AREAS COVERED: The review discusses the available randomized controlled trials and real-world observational studies conducted on safety and effectiveness of ceftaroline/ceftobiprole for the treatment of MRSA-BSIs. Finally, a proposal of MRSA-BSI treatment flowchart, based on fifth-generation cephalosporins, is described. EXPERT OPINION: The use of anti-MRSA cephalosporins is an acceptable choice either in monotherapy or combination therapy for the treatment of MRSA-BSIs due to their relevant effectiveness and safety. Particularly, their use may be advisable in combination therapy in case of severe infections (including endocarditis or persistent bacteriemia) or in monotherapy in subjects at higher risk of drugs-induced toxicity with older regimens. On the contrary, caution should be taken in case of suspected/ascertained central nervous system infections due to inconsistent data regarding penetration of these drugs in cerebrospinal fluid and brain tissues.

An overview of SARS-CoV-2 variants circulating in the 2020-2022 period in Lombardy.

Since the beginning of the pandemic, SARS-CoV-2 has shown genetic variability. All the variants that have sustained pandemic waves have shown several mutations, especially in the Spike protein that could affect viral pathogenesis. A total of 15,729 respiratory samples, collected between December 2020 and August 2022, have been included in this study. We report the circulation of SARS-CoV-2 variants in the Lombardy region, Italy, in a 2-year study period. Alpha, Delta, and Omicron variants became predominant causing the majority of cases whereas Beta or Gamma variants mostly caused local outbreaks. Next-generation sequencing revealed several mutations and few deletions in all of the main variants. For example, 147 mutations were observed in the Spike protein of Omicron sublineages; 20% of these mutations occurred in the receptor-binding domain region.

Quantitative SARS-CoV-2 subgenomic RNA as a surrogate marker for viral infectivity: Comparison between culture isolation and direct sgRNA quantification.

Detection of subgenomic (sg) SARS-CoV-2 RNAs are frequently used as a correlate of viral infectiousness, but few data about correlation between sg load and viable virus are available. Here, we defined concordance between culture isolation and E and N sgRNA quantification by ddPCR assays in 51 nasopharyngeal swabs collected from SARS-CoV-2 positive hospitalized patients. Among the 51 samples, 14 were SARS-CoV-2 culture-positive and 37 were negative. According to culture results, the sensitivity and specificity of E and N sgRNA assays were 100% and 100%, and 84% and 86%, respectively. ROC analysis showed that the best E and N cut-offs to predict positive culture isolation were 32 and 161 copies/mL respectively, with an AUC (95% CI) of 0.96 (0.91-1.00) and 0.96 (0.92-1.00), and a diagnostic accuracy of 88% and 92%, respectively. Even if no significant correlations were observed between sgRNA amount and clinical presentation, a higher number of moderate/severe cases and lower number of days from symptoms onset characterized patients with sgRNA equal to or higher than sgRNA cut-offs. Overall, this study suggests that SARS-CoV-2 sgRNA quantification could be helpful to estimate the replicative activity of SARS-CoV-2 and can represent a valid surrogate marker to efficiently recognize patients with active infection. The inclusion of this assay in available SARS-CoV-2 diagnostics procedure might help in optimizing fragile patients monitoring and management.

What do clinicians mean by epidemics' preparedness.

BACKGROUND: Infectious disease pandemics and epidemics pose significant global threats, and the risk of emerging infectious diseases has increased because of factors such as international connections, travel, and population density. Despite investments in global health surveillance, much of the world remains unprepared to manage infectious disease threats. OBJECTIVES: This review article discusses the general considerations and lessons learned from the COVID-19 pandemic in terms of epidemic preparedness. SOURCES: Non-systematic search on PubMed, scientific society websites, and scientific newspapers (performed in April 2023). CONTENT: Key factors for preparedness include robust public health infrastructure, adequate allocation of resources, and effective communication between stakeholders. This narrative review emphasizes the need for timely and accurate dissemination of medical knowledge, as well as addressing the challenges of misinformation and infodemics. It also highlights the importance of quick availability of diagnostic tests and vaccines, ensuring equitable access to these technologies. The role of scientific coordination in developing treatment strategies and the safety and mental well-being of healthcare workers are discussed. Lastly, it should be emphasized the need for medical training, multidisciplinary teams, new technologies and artificial intelligence, and the active role of infectious disease physicians in epidemic preparedness efforts. IMPLICATIONS: From clinicians' perspective, healthcare authorities play a crucial role in epidemic preparedness even by providing resource management plans, ensuring availability of essential supplies and training, facilitating communication, and improving safe infection management.

Pomegranate Extract Affects Gut Biofilm Forming Bacteria and Promotes Intestinal Mucosal Healing Regulating the Crosstalk between Epithelial Cells and Intestinal Fibroblasts.

Background: Pomegranate (Punica granatum) can be used to prepare a bioactive extract exerting anti-inflammatory activities. Clinical studies demonstrated an improvement in clinical response in inflammatory bowel disease (IBD) patients when pomegranate extract (PG) was taken as a complement to standard medications. However, the molecular mechanisms underlying its beneficial effects are still scarcely investigated. This study investigates the effect of PG on bacterial biofilm formation and the promotion of mucosal wound healing. Methods: The acute colitis model was induced in C57BL/6N mice by 3% dextran sodium sulfate administration in drinking water for 5 days. During the recovery phase of colitis, mice received saline or PG (200 mg/kg body weight) by oral gavage for 11 days. Colitis was scored daily by evaluating body weight loss, bleeding, and stool consistency. In vivo intestinal permeability was evaluated by fluorescein isothiocyanate-conjugated dextran assay, bacterial translocation was assessed by fluorescence in situ hybridization on tissues, whereas epithelial and mucus integrity were monitored by immunostaining for JAM-A and MUC-2 markers. Bacterial biofilm formation was assessed using microfluidic devices for 24 or 48 h. Primary fibroblasts were isolated from healthy and inflamed areas of 8 IBD patients, and Caco-2 cells were stimulated with or without PG (5 µg/mL). Inflammatory mediators were measured at the mRNA and protein level by RT-PCR, WB, or Bio-plex multiplex immunoassay, respectively. Results: In vivo, PG boosted the recovery phase of colitis, promoting a complete restoration of the intestinal barrier with the regeneration of the mucus layer, as also demonstrated by the absence of bacterial spread into the mucosa and the enrichment of crypt-associated fibroblasts. Microfluidic experiments did not highlight a specific effect of PG on Enterobacterales biofilm formation, even though Citrobacter freundii biofilm was slightly impaired in the presence of PG. In vitro, inflamed fibroblasts responded to PG by downregulating the release of metalloproteinases, IL-6, and IL-8 and upregulating the levels of HGF. Caco-2 cells cultured in a medium supplemented with PG increased the expression of SOX-9 and CD44, whereas in the presence of HGF or plated with a fibroblast-conditioned medium, they displayed a decrease in SOX-9 and CD44 expression and an increase in AXIN2, a negative regulator of Wnt signaling. Conclusions: These data provide new insight into the manifold effects of PG on promoting mucosal homeostasis in IBD by affecting pathogen biofilm formation and favoring the regeneration of the intestinal barrier through the regulation of the crosstalk between epithelial and stromal cells.

CMV seroprevalence and coronary CMV-DNA detection in immunocompetent patients with heart diseases.

BACKGROUND: Acute coronary syndromes (ACS) are a major cause of morbidity and mortality. As cytomegalovirus (CMV) may contribute to cardio-vascular (CV) manifestations, we sought to provide a proof-of-concept for the involvement of coronary and/or systemic CMV-reactivation as a possible ACS trigger. METHODS: We prospectively enrolled consecutive patients undergoing a coronary angiography for ACS (acute-cases, N.=136), or non-ACS reasons (chronic-cases, N.=57). Matched coronary and peripheral blood-samples were processed for quantification of CMV-DNAemia (RT-PCR), CMV-IgG/IgM, and CMV-IgG avidity (ELISA). Peripheral-blood samples from 17 healthy subjects were included as controls. RESULTS: Out of the 193 cases included, 18.1% were aged ≤55 years, 92.5% were Central-European, and 100% immunocompetent. CMV-IgG seroprevalence was 91.7% (95%CI: 87.8-95.6), significantly higher than in healthy-controls (52.9% [95%CI: 29.2-76.5]; P<0.001), yet consistent across age-groups (P=0.602), male/females (P=0.765), and acute/chronic-cases (P=0.157). Median (IQR) IgG titers were 110 (84-163) AU/mL, with 0.62 (0.52-0.72) avidity, supporting a long history of infection. No acute CMV infections were found. In 22.6% (n/N.=40/177) of the IgG-positive cases low-level coronary and/or systemic CMV-DNAemia (always <40 copies/mL) was detected. While no differences in peripheral CMV-DNAemia prevalence were observed nor among cases nor controls, coronary CMV-DNAemia was more frequent in acute-cases without modifiable CV risk-factors (n/N.=4/10; 40.0%), than in chronic-cases (n/N.=6/55, 10.9%; P=0.029), or acute-cases with risk-factors (n/N.=16/112, 14.3%; P=0.058). CONCLUSIONS: CMV-IgG seroprevalence was high in patients with heart diseases. CMV-DNAemia can be found, although uncommonly, in coronary circulation during an ACS, with increased prevalence in older subjects and in absence of CV risk-factors, identifying possible areas for novel interventions.

Increase in invasive group A streptococcal infections in Milan, Italy: a genomic and clinical characterization.

Background: Group A Streptococcus (GAS) causes multiple clinical manifestations, including invasive (iGAS) or even life-threatening (severe-iGAS) infections. After the drop in cases during COVID-19 pandemic, in 2022 a sharp increase of GAS was reported globally. Methods: GAS strains collected in 09/2022-03/2023 in two university hospitals in Milan, Italy were retrospectively analyzed. Clinical/epidemiological data were combined with whole-genome sequencing to: (i) define resistome/virulome, (ii) identify putative transmission chains, (iii) explore associations between emm-types and clinical severity. Results: Twenty-eight isolates were available, 19/28 (67.9%) from adults and 9/28 (32.1%) from pediatric population. The criteria for iGAS were met by 19/28 cases (67.9%), of which 11/19 (39.3%) met the further criteria for severe-iGAS. Pediatric cases were mainly non-invasive infections (8/9, 88.9%), adult cases were iGAS and severe-iGAS in 18/19 (94.7%) and 10/19 (52.6%), respectively. Thirteen emm-types were detected, the most prevalent being emm1 and emm12 (6/28 strains each, 21.4%). Single nucleotide polymorphism (SNP) analysis of emm1.0 and emm12.0 strains revealed pairwise SNP distance always >10, inconsistent with unique transmission chains. Emm12.0-type, found to almost exclusively carry virulence factors speH and speI, was mainly detected in children and in no-iGAS infections (55.6 vs. 5.3%, p = 0.007 and 66.7 vs. 0.0%, p < 0.001, respectively), while emm1.0-type was mainly detected in severe-iGAS (0.0 vs. 45.5%, p = 0.045). Conclusions: This study showed that multiple emm-types contributed to a 2022/2023 GAS infection increase in two hospitals in Milan, with no evidence of direct transmission chains. Specific emm-types could be associated with disease severity or invasiveness. Overall, these results support the integration of classical epidemiological studies with genomic investigation to appropriately manage severe infections and improve surveillance.

Performance evaluation of a new on-demand molecular test for the rapid identification of severe acute respiratory syndrome coronavirus 2 in pediatric and adult patients.

The rapid spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has increased the need to identify additional rapid diagnostic tests for an accurate and early diagnosis of infection. Here, we evaluated the diagnostic performance of the cartridge-based reverse transcription polymerase chain reaction (RT-PCR) test STANDARD M10 SARS-CoV-2 (SD Biosensor Inc., Suwon, South Korea), targeting the ORF1ab and E gene of SARS-CoV-2, and which can process up to eight samples in parallel in 60 min. From January 2022 to March 2022, STANDARD™ M10 assay performance was compared with Xpert® Xpress SARS-CoV-2 (Cepheid, Sunnyvale CA) on 616 nasopharyngeal swabs from consecutive pediatric (N = 533) and adult (N = 83) patients presenting at the "Istituto di Ricovero e Cura a Carattere Scientifico" (IRCCS) Ospedale Pediatrico Bambino Gesù, Roma. The overall performance of STANDARD M10 SARS-CoV-2 was remarkably and consistently comparable to the Xpert® Xpress SARS-CoV-2 with an overall agreement of 98% (604/616 concordant results), and negligible differences in time-to-result (60 min vs. 50 min, respectively). When the Xpert® Xpress SARS-CoV-2 results were considered as the reference, STANDARD™ M10 SARS-CoV-2 had 96.5% sensitivity and 98.4% specificity. STANDARD M10 SARS-CoV-2 can thus be safely included in diagnostic pathways because it rapidly and accurately identifies SARS-CoV-2 present in nasopharyngeal swabs.

Effective Rapid Diagnosis of Bacterial and Fungal Bloodstream Infections by T2 Magnetic Resonance Technology in the Pediatric Population.

Children are prone to bloodstream infections (BSIs), the rapid and accurate diagnosis of which is an unmet clinical need. The T2MR technology is a direct molecular assay for identification of BSI pathogens, which can help to overcome the limits of blood culture (BC) such as diagnostic accuracy, blood volumes required, and turnaround time. We analyzed results obtained with the T2Bacteria (648) and T2Candida (106) panels in pediatric patients of the Bambino Gesù Children's Hospital between May 2018 and September 2020 in order to evaluate the performance of the T2Dx instrument with respect to BC. T2Bacteria and T2Candida panels showed 84.2% and 100% sensitivity with 85.9% and 94.1% specificity, respectively. The sensitivity and specificity of the T2Bacteria panel increased to 94.9% and 98.7%, respectively, when BC was negative but other laboratory data supported the molecular result. T2Bacteria sensitivity was 100% with blood volumes <2 mL in neonates and infants. T2Bacteria and T2Candida provided definitive microorganism identification in a mean time of 4.4 and 3.7 h, respectively, versus 65.7 and 125.5 h for BCs (P < 0.001). T2 panels rapidly and accurately enable a diagnosis of a pediatric BSI, even in children under 1 year of age and for very small blood volumes. These findings support their clinical use in life-threatening pediatric infections, where the time to diagnosis is of utmost importance, in order to improve survival and minimize the long-term sequalae of sepsis. The T2 technology could be further developed to include more bacteria and fungi species that are involved in the etiology of sepsis.

Polymorphism of the HLA system and weak antibody response to BNT162b2 mRNA vaccine.

The polymorphism of the HLA system has been extensively studied in COVID-19 infection, however there are no data about the role of HLA on vaccine response. We report here the HLA-A, -B, -C, and DRB1 allelic frequencies of n = 111 individuals after BNT162b2 mRNA vaccine, selected on the basis of lower antibody levels (<5% percentile) after the second dose among a total of n = 2569 vaccinees, and compare them with the frequencies of a reference population. We found that differences in the frequencies of the alleles HLA-A*03:01, A*33:03, B*58:01 and at least one haplotype (HLA-A*24:02~C*07:01~B*18:01~DRB1*11:04) are associated with a weaker antibody response after vaccination, together with the age of vaccinees. Our results might suggest a role played by some HLA alleles or haplotypes in antibody production after the BNT162b2 mRNA vaccine, giving insights into the tracking of potentially susceptible individuals across populations. Further studies are needed to better define our exploratory findings and dissect the role of HLA polymorphism on response to anti-COVID-19 vaccines.

Viral Respiratory Infections: New Tools for a Rapid Diagnosis.

Respiratory tract infection is one of the most common diseases in human worldwide. Many viruses are implicated in these infections, including emerging viruses, such as the novel coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Identification of the causative viral pathogens of respiratory tract infections is important to select a correct management of patients, choose an appropriate treatment, and avoid unnecessary antibiotics use. Different diagnostic approaches present variable performance in terms of accuracy, sensitivity, specificity, and time-to-result, that have to be acknowledged to be able to choose the right diagnostic test at the right time, in the right patient. This review describes currently available rapid diagnostic strategies and syndromic approaches for the detection of viruses commonly responsible for respiratory diseases.

Results of the RENAISSANCE Study: REsponse to BNT162b2 COVID-19 vacciNe-short- And long-term Immune reSponSe evAluatioN in health Care workErs.

OBJECTIVE: To evaluate the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) anti-spike (S) IgG antibody production after vaccination with BNT162b2 and the protection from symptomatic breakthrough infections in health care workers. METHODS: This prospective observational study (RENAISSANCE) had as a primary end point the evaluation of serologic response to BNT162b2 14 days after a second dose. SARS-CoV-2 anti-S IgG antibodies were evaluated with LIAISON SARS-CoV-2 TrimericS IgG assay (DiaSorin S.p.A.), which is able to detect the presence of both binding and neutralizing antibodies for trimeric spike glycoprotein. Participants were recruited from February 1, 2021, to February 22, 2021. Occurrence of vaccine breakthrough infections was assessed by reverse transcription-polymerase chain reaction on symptomatic and contact cases up to June 6, 2021. RESULTS: Of 2569 staff evaluated, only 4 were nonresponders (0.16%; 95% CI, 0.04% to 0.41%). All 4 nonresponders were severely immunosuppressed and receiving treatment with mycophenolate mofetil or mycophenolic acid. At 14 days after the second dose, 67.5% (1733) of staff had anti-S IgG titers of 2000 BAU/mL or higher; 19.2% (494), between 1500 and 2000 BAU/mL; 9.8% (251), between 1000 and 1500 BAU/mL; and 3.4% (87), 1000 BAU/mL or lower. Women had a higher probability of having higher titers than men (64.5% [1044/1618] vs 58.3% [410/703]; P=.005). This was confirmed after adjustment for age group (odds ratio, 1.275; 95% CI, 1.062 to 1.531; P=.009). Four months after the end of the vaccination program, only 13 participants (0.26%) had experienced a breakthrough SARS-CoV-2 infection, including 1 nonresponder. This was the only participant requiring hospitalization for severe COVID-19. CONCLUSION: The vaccination campaign among health care workers at the ASST GOM Niguarda has resulted in a marked serologic response and reduction of incident COVID-19 cases. Yet, the lack of protection should not be overlooked in immunocompromised individuals.

Sex differences in electrolyte imbalances caused by SARS-CoV-2: A cross-sectional study.

BACKGROUND: Since SARS-CoV-2 spread, evidence regarding sex differences in progression and prognosis of COVID-19 have emerged. Besides this, studies on patients' clinical characteristics have described electrolyte imbalances as one of the recurrent features of COVID-19. METHODS: We performed a cross-sectional study on all patients admitted to the emergency department (ED) from 1 March to 31 May 2020 who had undergone a blood gas analysis and a nasopharyngeal swab test for SARS-CoV-2 by rtPCR. We defined positive patients as cases (n = 710) and negatives as controls (n = 619), for a total number of patients of 1.329. The study was approved by the local ethics committee Area 3 Milan. Data were automatically extracted from the hospital laboratory SQL-based repository in anonymised form. We considered as outcomes potassium (K+ ), sodium (Na+ ), chlorine (Cl- ) and calcium (Ca++ ) as continuous and as categorical variables, in their relation with age, sex and SARS-CoV-2 infection status. RESULTS: We observed a higher prevalence of hypokalaemia among patients positive for SARS-CoV-2 (13.7% vs 6% of negative subjects). Positive patients had a higher probability to be admitted to the ED with hypokalaemia (OR 2.75, 95% CI 1.8-4.1, P < .0001) and women were twice as likely to be affected than men (OR 2.43, 95% CI 1.67-3.54, P < .001). Odds ratios for positive patients to manifest with an alteration in serum Na+ was (OR 1.6, 95% CI 1.17-2.35, P < .001) and serum chlorine (OR 1.6, 95% CI 1.03-2.69, P < .001). Notably, OR for positive patients to be hypocalcaemic was 7.2 (95% CI 4.8-10.6, P < .0001) with a low probability for women to be hypocalcaemic (OR 0.63, 95% CI 0.4-0.8, P = .005). CONCLUSIONS: SARS-CoV-2 infection is associated with a higher prevalence of hypokalaemia, hypocalcaemia, hypochloraemia and sodium alterations. Hypokalaemia is more frequent among women and hypocalcaemia among men.

Rapid Detection and Quantification of Mycobacterium tuberculosis DNA in Paraffinized Samples by Droplet Digital PCR: A Preliminary Study.

Background: Rapid and reliable diagnosis of tuberculosis (TB) represents a diagnostic challenge in compartmentalized extrapulmonary TB infection because of the small number of mycobacteria (MTB) and the frequent lack of fresh samples to perform culture. Here, we estimate the performances of homemade droplet digital PCR (ddPCR)-based assays against culture in 89 biopsies, for those fresh and formalin-fixed and paraffin-embedded (FFPE) subsamples were available. Methods: MTB diagnosis in fresh subsamples was performed by culture. Fresh subsamples were also analyzed for acid-fast bacilli smear-microscopy (AFB) and Xpert® MTB/RIF (Xpert). MTB examination was repeated in blind in the 89 FFPE subsamples by in-house ddPCR assays targeting the IS6110 and rpoB. Analytical sensitivity of ddPCR assays was evaluated using serial dilution of H37Rv strain. Limit of detection (LOD) was calculated by probit analysis. Results were expressed in copies/106 cells. Results: IS6110 and rpoB ddPCR assays showed a good linear correlation between expected and observed values (R 2: 0.9907 and 0.9743, respectively). Probit analyses predicted a LOD of 17 and 40 copies/106 cells of MTB DNA for IS6110 and rpoB, respectively. Of the 89 biopsies, 68 were culture positive and 21 were culture negative. Considering mycobacterial culture as reference method, IS6110 assay yielded positive results in 67/68 culture-positive samples with a median interquartile range (IQR) of 1,680 (550-8,444) copies/106 cells (sensitivity: 98.5%; accuracy: 98.9). These performances were superior to those reported by the rpoB assay in FFPE subsamples (sensitivity: 66.20%; accuracy: 74.1) and even superior to those reported by Xpert and AFB in fresh subsamples (sensitivity: 79.4 and 33.8%, respectively; accuracy: 84.3 and 49.4, respectively). When Xpert and AFB results were stratified according to mycobacterial load detected by rpoB and IS6110 ddPCR, bacterial load was lower in Xpert and AFB negative with respect to Xpert and AFB-positive samples (p = 0.003 and 0.01 for rpoB and p = 0.01 and 0.11 for IS6110), confirming the poor sensitivity of these methods in paucibacillary disease. Conclusion: ddPCR provides highly sensitive, accurate, and rapid MTB diagnosis in FFPE samples, as defined by the high concordance between IS6110 assay and culture results. This approach can be safely introduced in clinical routine to accelerate MTB diagnosis mainly when culture results remain unavailable.

Frontline Screening for SARS-CoV-2 Infection at Emergency Department Admission by Third Generation Rapid Antigen Test: Can We Spare RT-qPCR?

To complement RT-qPCR testing for diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections, many countries have introduced the use of rapid antigen tests. As they generally display lower real-life performances than expected, their correct positioning as frontline screening is still controversial. Despite the lack of data from daily clinical use, third generation microfluidic assays (such as the LumiraDx SARS-CoV-2 Ag test) have recently been suggested to have similar performances to RT-qPCR and have been proposed as alternative diagnostic tools. By analyzing 960 nasopharyngeal swabs from 960 subjects at the emergency department admissions of a tertiary COVID-19 hospital, LumiraDx assay demonstrated a specificity of 97% (95% CI: 96-98), and a sensitivity of 85% (95% CI: 82-89) in comparison with RT-qPCR, which increases to 91% (95% CI: 86-95) for samples with a cycle threshold ≤ 29. Fifty false-negative LumiraDx-results were confirmed by direct quantification of genomic SARS-CoV-2 RNA through droplet-digital PCR (median (IQR) load = 5880 (1657-41,440) copies/mL). Subgenomic N and E RNAs were detected in 52% (n = 26) and 56% (n = 28) of them, respectively, supporting the presence of active viral replication. Overall, the LumiraDx test complies with the minimum performance requirements of the WHO. Yet, the risk of a misrecognition of patients with active COVID-19 persists, and the need for confirmatory RT-qPCR should not be amended.

Dolutegravir Plus Lamivudine Two-Drug Regimen: Safety, Efficacy and Diagnostic Considerations for Its Use in Real-Life Clinical Practice-A Refined Approach in the COVID-19 Era.

The diagnostic and therapeutic management of the Coronavirus Disease 2019 (COVID-19) pandemic in the HIV population brought some known criticalities (and opportunities) to the forefront, for both those who are facing their first therapeutic line today, and for those already well viro-suppressed. The clinical, socioeconomic, and psychological impact of the COVID-19 pandemic should not affect the long-term care of people living with HIV, which creates an urgent need to optimize the diagnostic and treatment approach to the first-line or switch regimens. The use of dolutegravir plus a lamivudine two-drug regimen is one of the most promising solutions to ease the management of HIV treatment in this difficult period. In this review, we report the most salient features related to the use of this regimen from real-life cohorts, meta-analyses, randomized clinical trials, and studies presented at international conferences up to March 2021. We focused on the diagnostic and clinical-management implications of its use in real life, and how these comply with the contingent historical situation. The issue of the timing and type of diagnostic procedures and the relevance of classical diagnostic tests (such as genotype for resistance detection) is also discussed. According to the currently available results, dolutegravir plus a lamivudine two-drug regimen represents an outstanding tool, whose expected advantages fulfill the current requirements for optimal daily care of our HIV patients.

Genetic Determinants in a Critical Domain of NS5A Correlate with Hepatocellular Carcinoma in Cirrhotic Patients Infected with HCV Genotype 1b.

HCV is an important cause of hepatocellular carcinoma (HCC). HCV NS5A domain-1 interacts with cellular proteins inducing pro-oncogenic pathways. Thus, we explore genetic variations in NS5A domain-1 and their association with HCC, by analyzing 188 NS5A sequences from HCV genotype-1b infected DAA-naïve cirrhotic patients: 34 with HCC and 154 without HCC. Specific NS5A mutations significantly correlate with HCC: S3T (8.8% vs. 1.3%, p = 0.01), T122M (8.8% vs. 0.0%, p < 0.001), M133I (20.6% vs. 3.9%, p < 0.001), and Q181E (11.8% vs. 0.6%, p < 0.001). By multivariable analysis, the presence of >1 of them independently correlates with HCC (OR (95%CI): 21.8 (5.7-82.3); p < 0.001). Focusing on HCC-group, the presence of these mutations correlates with higher viremia (median (IQR): 5.7 (5.4-6.2) log IU/mL vs. 5.3 (4.4-5.6) log IU/mL, p = 0.02) and lower ALT (35 (30-71) vs. 83 (48-108) U/L, p = 0.004), suggesting a role in enhancing viral fitness without affecting necroinflammation. Notably, these mutations reside in NS5A regions known to interact with cellular proteins crucial for cell-cycle regulation (p53, p85-PIK3, and ß-catenin), and introduce additional phosphorylation sites, a phenomenon known to ameliorate NS5A interaction with cellular proteins. Overall, these results provide a focus for further investigations on molecular bases of HCV-mediated oncogenesis. The role of theseNS5A domain-1 mutations in triggering pro-oncogenic stimuli that can persist also despite achievement of sustained virological response deserves further investigation.

SARS-CoV-2 RNA in plasma samples of COVID-19 affected individuals: a cross-sectional proof-of-concept study.

BACKGROUND: Recent studies showed that plasma SARS-CoV-2 RNA seems to be associated with worse COVID-19 outcome. However, whether specific population can be at higher risk of viremia are to date unexplored. METHODS: This cross-sectional proof-of-concept study included 41 SARS-CoV-2-positive adult individuals (six affected by haematological malignancies) hospitalized at two major hospital in Milan, for those demographic, clinical and laboratory data were available. SARS-CoV-2 load was quantified by ddPCR in paired plasma and respiratory samples. To assess significant differences between patients with and patients without viremia, Fisher exact test and Wilcoxon test were used for categorical and continuous variables, respectively. RESULTS: Plasma SARS-CoV-2 RNA was found in 8 patients (19.5%), with a median (IQR) value of 694 (209-1023) copies/mL. Viremic patients were characterized by an higher mortality rate (50.0% vs 9.1%; p = 0.018) respect to patients without viremia. Viremic patients were more frequently affected by haematological malignancies (62.5% vs. 3.0%; p < 0.001), and had higher viral load in respiratory samples (9,404,000 [586,060-10,000,000] vs 1560 [312-25,160] copies/mL; p = 0.002). CONCLUSIONS: Even if based on a small sample population, this proof-of-concept study poses the basis for an early identification of patients at higher risk of SARS-CoV-2 viremia, and therefore likely to develop severe COVID-19, and supports the need of a quantitative viral load determination in blood and respiratory samples of haematologic patients with COVID-19 in order to predict prognosis and consequently to help their further management.

Genomic epidemiology of SARS-CoV-2 reveals multiple lineages and early spread of SARS-CoV-2 infections in Lombardy, Italy.

From February to April 2020, Lombardy (Italy) reported the highest numbers of SARS-CoV-2 cases worldwide. By analyzing 346 whole SARS-CoV-2 genomes, we demonstrate the presence of seven viral lineages in Lombardy, frequently sustained by local transmission chains and at least two likely to have originated in Italy. Six single nucleotide polymorphisms (five of them non-synonymous) characterized the SARS-CoV-2 sequences, none of them affecting N-glycosylation sites. The seven lineages, and the presence of local transmission clusters within three of them, revealed that sustained community transmission was underway before the first COVID-19 case had been detected in Lombardy.