MicroRNA Expression Levels in Peripheral Blood Mononuclear Cells from Human Immunodeficiency Virus Type 1 Positive Individuals and Relationship with Different Levels of Viral Suppression.

The persistence of low human immunodeficiency virus type 1 (HIV-1) replication in individuals undergoing antiretroviral therapy (ART) still threatens their health. Previous findings have shown that microRNAs (miRNAs) could interfere with several steps of the viral life cycle. Herein, we set out to investigate the expression of miR-150, miR-223, miR-382, miR-324-5p, miR-33a-5p, miR-34a, and miR-132 in the whole peripheral blood mononuclear cell (PBMC) population from people living with HIV-1 showing different levels of viral suppression. Levels of PBMC-associated miRNAs were analyzed in 30 individuals with undetectable viremia (target not detected) and 30 individuals with detectable low-level viremia (1-200 copies/mL). In addition, 30 samples from treatment-naive (NAIVE) individuals were investigated. Results were compared to a control group of 28 HIV-negative donors. All miRNAs analyzed were strongly downregulated in the NAIVE population, either compared to the treated group or to controls. Stratification of ART-treated donors according to the therapeutic regimen showed the downregulation of miR-33a-5p in subjects treated with non-nucleoside reverse transcriptase inhibitors compared with those treated with protease inhibitors. Collectively, the present study shows that uncontrolled viral replication leads to profound miRNA deregulation while treated individuals, irrespective of the degree of viral suppression, and even the types of antiviral drugs seem to be specifically associated with miRNA expression profiles. These evidences suggest that virological suppression could be favored by miRNA modulation.

Infections and Colon Surgery: Preliminary Results from a Surveillance Program in an Italian Hospital.

Surgical site infections (SSIs) represent a valid indicator of the healthcare quality. This study described the preliminary results of one-year active surveillance program on colon surgeries in a hospital in Molise region, central Italy. Patients who had undergone colon surgery according to National Healthcare Safety Network were included. Data on intervention, perioperative antibiotic prophylaxis, and SSIs occurrence were collected. Chi-square and Fisher's Exact test were used to evaluate any association between risk factors and SSIs. Sixty-eight patients (mean age 70.6 years) were included, and 44 (64.7%) were males. The most frequent interventions were right (n = 17, 25.0%) and left (n = 15, 22.0%) hemicolectomy. Surgical interventions were largely elective (n = 43, 63.2%) and with laparotomy (n = 56, 82.4%). During hospital stay, 10 (14.7%) SSIs were detected, including five superficial, three deep and two organ/space infections. Three (4.4%) additional SSIs were detected at post-discharge follow-up, for 13 (19.1%; CI95%: 9.7%-28.5%) total cases detected. Metronidazole plus Ceftriaxone (third generation cephalosporin) was the antibiotics combination mostly used (n = 36, 52.9%) for the perioperative antibiotic prophylaxis within 60 minutes of incision. The study underlines the need of improvements of the practices currently adopted, since SSIs could be significantly reduced through a multimodal strategy generating bundles. As third generation cephalosporins may facilitate resistant strains emergence, for perioperative prophylaxis in clean-contaminated interventions with entry into gastrointestinal tract, Cefazolin plus Metronidazole or only second generation cephalosporin are recommended. Due to the large variability of post-intervention antibiotic therapy, antimicrobial stewardship approach is strictly necessary.

Humoral and T-cell mediated response after administration of mRNA vaccine BNT162b2 in frail populations.

Patients with frailty are considered to be at greater risk to get severe infection from SARS-CoV-2. One of the most effective strategies is vaccination. In our study we evaluated both the humoral immune response elicited by the vaccination at different time points, and the T-cell response in terms of interferon (IFN)-γ production in frail patients and healthy donors. Fifty-seven patients (31 patients undergoing hemodialysis and 26 HIV positive subjects) and 39 healthcare workers were enrolled. All participants received two doses of the mRNA vaccine BNT162b2. Healthcare workers showed a significantly higher antibody titer than patients twenty-one days after the first dose (p < 0.001). From the same time point we observed for both groups a decay of the antibody levels with a steeper slope of decline in the patients group. Regarding T-cell response the only significant difference between non-reactive and reactive subjects was found in median antibody levels, higher in the responders group than in non-responders. The healthcare workers seem to better respond to the vaccination in terms of antibodies production; the lack of T-cell response in about 50% of the participants seems to suggest that in our study population both humoral and cell-mediated response decline over time remarking the importance of the booster doses, particularly for frail patients.

Comparison of FTD SARS-CoV-2 Assay and RealStar RT-PCR kit 1.0 for the detection of SARS-CoV-2.

The aim of the study was to evaluate the clinical performance of FTD SARS-CoV-2 compared to the RealStar RT-PCR kit 1.0. The analysis of 100 nasopharyngeal swabs showed an overall agreement of 88 %. The positive percentage agreement was 85.6 % and the negative percentage agreement was 91 %. In conclusion we observed a substantial agreement among the two methods, with discrepancies mainly observed in specimens with relatively low amount of viral RNA.

KI and WU Polyomavirus in Respiratory Samples of SARS-CoV-2 Infected Patients.

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) has been declared a global pandemic. Our goal was to determine whether co-infections with respiratory polyomaviruses, such as Karolinska Institutet polyomavirus (KIPyV) and Washington University polyomavirus (WUPyV) occur in SARS-CoV-2 infected patients. Oropharyngeal swabs from 150 individuals, 112 symptomatic COVID-19 patients and 38 healthcare workers not infected by SARS-CoV-2, were collected from March 2020 through May 2020 and tested for KIPyV and WUPyV DNA presence. Of the 112 SARS-CoV-2 positive patients, 27 (24.1%) were co-infected with KIPyV, 5 (4.5%) were positive for WUPyV, and 3 (2.7%) were infected simultaneously by KIPyV and WUPyV. Neither KIPyV nor WUPyV DNA was detected in samples of healthcare workers. Significant correlations were found in patients co-infected with SARS-CoV-2 and KIPyV (p < 0.05) and between SARS-CoV-2 cycle threshold values and KIPyV, WUPyV and KIPyV and WUPyV concurrently detected (p < 0.05). These results suggest that KIPyV and WUPyV may behave as opportunistic respiratory pathogens. Additional investigations are needed to understand the epidemiology and the prevalence of respiratory polyomavirus in COVID-19 patients and whether KIPyV and WUPyV could potentially drive viral interference or influence disease outcomes by upregulating SARS-CoV-2 replicative potential.

SARS-CoV-2 diagnostics in the virology laboratory of a University Hospital in Rome during the lockdown period.

Italy was one of the most affected nations by coronavirus disease 2019 outside China. The infections, initially limited to Northern Italy, spread to all other Italian regions. This study aims to provide a snapshot of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) epidemiology based on a single-center laboratory experience in Rome. The study retrospectively included 6565 subjects tested for SARS-CoV-2 at the Laboratory of Virology of Sapienza University Hospital in Rome from 6 March to 4 May. A total of 9995 clinical specimens were analyzed, including nasopharyngeal swabs, bronchoalveolar lavage fluids, gargle lavages, stools, pleural fluids, and cerebrospinal fluids. Positivity to SARS-CoV-2 was detected in 8% (527/6565) of individuals, increased with age, and was higher in male patients (P < .001). The number of new confirmed cases reached a peak on 18 March and then decreased. The virus was detected in respiratory samples, in stool and in pleural fluids, while none of gargle lavage or cerebrospinal fluid samples gave a positive result. This analysis allowed to gather comprehensive information on SARS-CoV-2 epidemiology in our area, highlighting positivity variations over time and in different sex and age group and the need for a continuous surveillance of the infection, mostly because the pandemic evolution remains unknown.

Molecular Epidemiological Insights into Colistin-Resistant and Carbapenemases-Producing Clinical Klebsiella pneumoniae Isolates.

PURPOSE: Carbapenemases-producing Klebsiella pneumoniae are challenging antimicrobial therapy of hospitalised patients, which is further complicated by colistin resistance. This study describes molecular epidemiological insights into colistin-resistant and carbapenemases-producing clinical K. pneumoniae. PATIENTS AND METHODS: Cultures collected from 26 hospitalised patients during 2014-2017 in the main hospital in Molise Region, central Italy, were characterized. The minimum inhibitory concentration for 19 antibiotics was determined, including carbapenems and colistin. Prevalence of resistance-associated genes was investigated through PCR, detecting bla KPC, bla GES, bla VIM, bla IMP, bla NDM, bla OXA-48, bla CTX-M, bla TEM, bla SHV, and mcr-1,2,3,4,5,6,7,8. The mgrB gene was also analysed in colistin-resistant strains by PCR and sequencing assays. K. pneumoniae were typed by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). RESULTS: Twenty out of 26 K. pneumoniae were phenotypically resistant to carbapenems and 19 were resistant to colistin. All isolates harbored bla KPC, and bla SHV, bla TEM and bla VIM were further the most common resistance-associated genes. In colistin-resistant strains, mcr-1,2,3,4,5,6,7,8 variants were not detected, while mutations and insertion elements in mgrB were observed in 68.4% (n=13) in 31.6% (n=6) isolates, respectively. PFGE revealed 12 clusters and 18 pulsotypes at 85% and 95% cut-off, while the Sequence Types ST512 (n=13, 50%), ST101 (n=10, 38.5%), ST307 (n=2, 7.7%) plus a novel ST were detected using MLST. CONCLUSION: All K. pneumoniae showed a multidrug-resistant phenotype, particularly to carbapenems and colistin. According to national data, bla KPC was the prevailing carbapenemase, followed by bla VIM, while bla TEM and bla SHV were among the most frequent beta-lactamases. Consistent with previous reports in Italy, ST512 was the most common clone, particularly during 2014-15, whilst ST101 became dominant in 2016-17. Colistin resistance was mainly associated with deleterious mutations and transposon in the mgrB gene. Improvements of surveillance, compliance with infection prevention procedures and antimicrobial stewardship are essential to limit the spread of resistant K. pneumoniae.

Tracking Multidrug-Resistant Klebsiella pneumoniae from an Italian Hospital: Molecular Epidemiology and Surveillance by PFGE, RAPD and PCR-Based Resistance Genes Prevalence.

Antimicrobial-resistant Klebsiella pneumoniae represent a global public health concern. K. pneumoniae strains isolated during 2010 and 2014-2016 within a single hospital of Molise Region, Central Italy, were analyzed testing antimicrobial susceptibility, clonality by pulsed-field gel electrophoresis (PFGE) and random amplified polymorphic DNA (RAPD)-PCR, and prevalence of carbapenem resistance genes by PCR. Forty isolates (23 wild-type in 2010 and 17 non-wild-type in 2014-2016) were collected from hospitalized patients (65.2 ± 18.1 years old, 75% male, 80% from intensive care unit-ICU). K. pneumoniae showed multidrug-resistant profiles and 15 resistotypes were identified (discriminatory power D = 0.88). The 69.6 and 17.4% of isolates in 2010 resulted intermediate and resistant to imipenem, respectively, and 91.3% was sensitive to meropenem, while 88.2% of isolates of 2014-2016 were resistant to both antibiotics. PFGE identified 16 clusters versus 23 by RAPD, 26 pulsotypes versus 33 RAPD patterns (D ≥ 0.97). PFGE separated strains according to isolation period and identified an outbreak occurred in the ICU during December 2014 and January 2015. No strains harbored blaGES, blaIMP, blaNDM-1, and blaOXA-48 genes, as well as AmpC plasmid-mediated beta-lactamases genes. Only K. pneumoniae isolated during 2014-2016 were blaKPC positive. Prevalence of blaVIM was 87 and 76.5% during 2010 and 2014-2016, respectively. No strains colistin-resistant harbored mcr-1 plasmid-mediated resistance gene. The study findings underline an increased circulation of multidrug-resistant K. pneumoniae within the hospital, and the acquisition of carbapenem resistance mechanism. The implementation of surveillance and molecular characterization of isolates are needed to identify outbreaks, reduce the spread of resistance, and guide empirical therapy.