Identification of a novel intra-genotype reassortant G1P[8] rotavirus in Italy, 2021.

OBJECTIVES: Rotaviruses G1P[8] are epidemiologically relevant and are targeted by vaccines. The introduction of vaccines has altered rotavirus epidemiology. Hospital-based surveillance conducted in Sicily, Italy, showed a progressive decline in rotavirus prevalence since 2014, along with an increasing vaccine coverage (63.8% in 2020), and a marked decrease in circulation of G1P[8] strains. Surprisingly in 2021, G1P[8] viruses accounted for 90.5% (19/21) of rotavirus infections. This study aimed to understand if the increased activity of G1P[8]'s was related to virus-related peculiarities. DESIGN: In 2021, 266 patients <15 years of age were hospitalized with acute gastroenteritis (AGE) and included in rotavirus surveillance. Viral proteins (VP7 and VP4) genotyping and sequence data were generated from all rotavirus-positive samples. The genetic makeup of G1P[8] rotaviruses was investigated by full-genome sequencing. RESULTS: Peculiar G1P[8] rotaviruses, with VP7 and VP4 belonging to novel sub-lineages, circulated in 2021, accounting for 76.2% (16/21) of all rotavirus infections. On full-genome analysis, the novel G1P[8] variant displayed an intra-genotype (Wa-like) reassortant constellation, involving G12 and G1 strains, into a unique arrangement never observed before. The novel G1P[8] variant showed peculiar amino acid substitutions in 8-1 and 8-3 epitopes of the VP4 with respect to the Rotarix strain. CONCLUSIONS: Prompt identification of virus variants circulating in the human population is pivotal to understanding epidemiological trends and assessing vaccine efficacy.

Characterization of Listeria monocytogenes Strains Isolated in Palermo (Sicily and Italy) during the Years 2018-2020 from Severe Cases of Listeriosis.

Listeria monocytogenes (LM), the etiological agent of listeriosis, can cause foodborne zoonosis. In this study, we characterized 23 strains that caused human severe listeriosis in Palermo (Sicily, Italy) during the period of 2018-2020. In addition, we assessed the phenotypic susceptibility of clinical isolates to antibiotics in accordance with EUCAST guidelines. The serogroup was determined through the use of PCR, while MLST and MVLST were identified through the sequencing of housekeeping genes. Finally, susceptibility to antibiotics was assessed by means of the Phoenix automatic system. Patients hospitalized with listeriosis were predominantly males (56% vs. 44% of females). The cases not associated with pregnancy included patients >65 years of age (60%), two of whom were affected by cancer, while cases associated with pregnancy included two pregnant women and three preterm infants. The data collected showed that the main pathologies shown by patients were meningitis (60.9%) and bacteremia (39.1%). The LM strains were isolated from the blood (52%), cerebrospinal fluid (26%), cerebrospinal fluid + blood (13%), blood + a nasal swab (4%), and ascitic fluid (4%). The predominant serogroup was IVb (96%), whereas only one strain belonged to serogroup IIa (4%). Among the strains with serotypes 4b, 4d, and 4e, ST2/VT21 (92%) and ST6/VT19 (4%) were determined, while only isolates with serotypes 1/2a and3a show ST155/VT45 (CC155). This study reveals the widespread circulation of a clinical strain (ST2/VT21) associated with suspected food contamination, demonstrating the importance of carrying out molecular epidemiological surveillance. Our clinical isolates were susceptible to the beta-lactams assayed, in agreement with the literature data.

Biological Specimen Banking as a Time Capsule to Explore the Temporal Dynamics of Norovirus Epidemiology.

Norovirus is recognised as a major cause of epidemic and sporadic acute gastroenteritis (AGE) in all age groups. Information on the genetic diversity of the noroviruses circulating in the 1980s and 1990s, before the development and adoption of dedicated molecular assays, is limited compared with the last decades. Between 1986 and 2020, uninterrupted viral surveillance was conducted in symptomatic children hospitalized with AGE in Palermo, Italy, providing a unique time capsule for exploring the epidemiological and evolutionary dynamics of enteric viruses. A total of 8433 stool samples were tested using real-time RT-PCR. All samples were stored at -20 or -80 °C until processing. In this 35-year long time span, noroviruses of genogroup II (GII) were detected in 15.6% of AGE requiring hospitalization, whilst GI noroviruses were detected in 1.4% of AGE. Overall, the predominant norovirus capsid (Cap) genotype was GII.4 (60.8%), followed by GII.3 (13.3%) and GII.2 (12.4%). Temporal replacement of the GII.4 Cap variants associated with different polymerase (Pol) types were observed over the study period. The chronology of emergence and circulation of the different GII.4 variants were consistent with data available in the literature. Also, for GII.3 and GII.2 NoVs, the circulation of different lineages/strains, differing in either the Cap or Pol genes or in both, was observed. This long-term study revealed the ability of noroviruses to continuously and rapidly modify their genomic makeup and highlights the importance of surveillance activities in vaccine design.

The Rapid Phenotypic Susceptibility Testing in Real-Life Experience: How the MIC Values Impact on Sepsis Fast Diagnostic Workflow.

The MIC value definition faithfully reflects antimicrobial sensitivity, profoundly impacting the infection's clinical outcome. Our study aimed to evaluate the Accelerate PhenoTM System in defining the importance of fast phenotypic susceptibility data. A number of 270 monomicrobial samples simultaneously underwent standard procedures and fast protocols after a contemporary Gram stain. Finally, we provided Turn-around Time (TAT) and statistical evaluations. The fast technology required a medium value of 7 h to complete ID and AST profiles. Although there were some spectrum limitations, it revealed an optimal success rate in microbial identification directly from positive blood cultures. The Gram-negative AST reached a 98.9% agreement between the Accelerate Pheno™ System and the standard method. In addition, the Gram-positive AST gathered a 98.7% agreement comparing the same systems. The chance to rapidly provide precise MIC values is one of the last frontiers in clinical microbiology, especially in high-prevalence antimicrobial resistance areas.

Elexacaftor-Tezacaftor-Ivacaftor as a Final Frontier in the Treatment of Cystic Fibrosis: Definition of the Clinical and Microbiological Implications in a Case-Control Study.

The use of modulator drugs that target the Cystic Fibrosis transmembrane conductance regulator (CFTR) is the final frontier in the treatment of Cystic Fibrosis (CF), a genetic multiorgan disease. F508del is the most common mutation causing defective formation and function of CFTR. Elexacaftor-tezacaftor-ivacaftor is the first triple combination of CFTR modulators. Herein, we report on a one-year case-control study that involved 26 patients with at least one F508del mutation. Patients were assigned to two similar groups, and patients with the worse clinical condition received treatment with the triple combination therapy. The study aimed to define the clinical and especially microbiological implications of treatment administration. The treatment provided significant clinical benefits in terms of respiratory, pancreatic, and sweat function. After one year of therapy, airway infection rates decreased and pulmonary exacerbations were dramatically reduced. Finally, treated patients reported a surprising improvement in their quality of life. The use of triple combination therapy has become essential in most CF people carrying the F508del mutation. Although the clinical and instrumental benefits of treatment are thoroughly known, further investigations are needed to properly define its microbiological respiratory implications and establish the real advantage of life-long treatment with elexacaftor-tezacaftor-ivacaftor.

Clinical and Epidemiologic Features of Viral Gastroenteritis in Hospitalized Children: An 11-Year Surveillance in Palermo (Sicily).

In order to acquire information regarding viral agents and epidemiologic features of severe paediatric Viral Acute Gastroenteritis (VAGE) across multiple seasons in the pre-rotavirus-vaccine era, the epidemiologic characteristics of VAGE were investigated among paediatric patients hospitalized in a major Sicilian paediatric hospital from 2003 to 2013. Overall, 4725 children were observed and 2355 (49.8%) were diagnosed with a viral infection: 1448 (30.6%) were found positive to rotavirus, 645 (13.7%) to norovirus, 216 (4.6%) to adenovirus, and 46 (0.97%) to astrovirus. Viral infections showed different patterns of hospitalization in terms of age at risk (younger for rotavirus and adenovirus infections), seasonality (increased risk in winter for rotavirus and norovirus), trend over time (reduced risk in 2011-2013 for norovirus and rotavirus) and major diagnostic categories (digestive diseases more frequent in adenovirus and astrovirus but not in norovirus). This study increases general knowledge of VAGE epidemiology and contributes to suggest some a priori diagnostic criteria that could help clinicians to identify and treat viral agents responsible for gastroenteritis in hospital settings.

Invasive cryptococcal disease in COVID-19: systematic review of the literature and analysis.

During the Coronavirus Disease 2019 (COVID-19) pandemic, an increasing number of fungal infections associated with SARS-CoV-2 infection have been reported. Among them, cryptococcosis could be a life-threatening disease. We performed a Systematic Review (PRISMA Statement) of cryptococcosis and COVID-19 co-infection, case report/series were included: a total of 34 cases were found, then we added our case report. We collected patients' data and performed a statistical analysis comparing two groups of patients sorted by outcome: "dead" and "alive". Three cases were excluded for lack of information. To compare categorical data, we used a Fisher-exact test (α=0.05). To compare quantitative variables a U Mann-Whitney test was used (α=0.05), with a 95% Confidence Interval. A total of 32 co-infected patients were included in the statistical analysis. Mortality rate was 17/32 (53.1%): these patients were included in "dead" group, and 15/32 (46.9%) patients survived and were included in "alive" group. Overall, males were 25/32 (78.1%), the median age was 60 years (IQR 53-70) with non-statistically significant difference between groups (p=0.149 and p=0.911, respectively). Three variables were associated with mortality: ARDS, ICU admission and inadequate treatment. Overall, 21 out of 24 (87.5%) patients were in ARDS with a statistically significant difference among two groups (p=0.028). ICU admission for COVID-19 was observed in 18/26 (69.2%), more frequently among dead group (p=0.034). Finally, 15/32 (46.9%) patients had adequate treatment (amphotericin B + flucytosine for invasive cryptococcosis) mostly among alive patients (p=0.039). In conclusion, mortality due to cryptococcal infection among COVID-19 patients remains high but an early diagnosis and appropriate treatment could reduce mortality.

Evaluation of the diagnostic performances of two commercially available assays for the detection of enteric adenovirus antigens.

The performance of 2 antigenic commercial assays for enteric adenovirus (AdV) infection, bioNexia Rota-Adeno ImmunoChromatographic Tests (ICT) and LIAISON® Adenovirus ChemiLuminescence Immuno Assays (CLIA), was evaluated on 321 stools from children hospitalized for acute gastroenteritis in Palermo, Italy, using a Real time-PCR (Rt-PCR) as reference method. The CLIA showed higher sensitivity (77% vs 60%), accuracy (94.4 vs 90.9) and concordance (k: 0.81 vs 0.67) with respect to ICT, despite equivalent specificity (98.8%). Using the Ct values of the Rt-PCR as a proxy of the fecal viral load, similar Ct values (mean 9.32 vs 9.89) were observed among the true positive samples, whilst a significant difference (P < 0.05) was observed in false negative samples of CLIA (mean Ct 25.68) and ICT (mean Ct 19.87). Cross-reactivity with other enteric viruses was not observed. These results indicate that both the assays tested are suitable for diagnosis of AdV gastroenteritis.

Immune Response Failure in Paucisymptomatic Long-Standing SARS-CoV-2 Spreaders.

The coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has rapidly spread throughout the world. This disease has a spectrum of different clinical pictures with different outcomes. Herein, we report all the data from three paucisymptomatic patients during a hospital stay that might represent a paradigmatic example of the method by which SARS-CoV-2 is shed. We demonstrated the lack of an adequate qualitative and quantitative immune response by multiparametric flow cytometry analysis. Our data can provide a new perspective about the method by which SARS-CoV-2 is shed and the clinical weight of viral persistence. In all three cases, the long persistence of the virus and the consistent reduction in both innate and adaptative immune cells are not associated with greater disease severity. These patients might represent at least part of the population. In particular, one patient oscillated between positive and negative swab tests several times without presenting any immune response. In all three cases, the immune response failure was not associated with a clinically significant involvement, indicating that it is not the virus's ability to impair the immune system, as well as its presence and persistence the fundamental mechanism that might causally lead to death. Finally, this kind of immune response in paucisymptomatic patients could pose a considerable danger to public health that questions the quarantine period. It is urgent to quantify the phenomenon with a large sample study.

Immunological Characteristics of Non-Intensive Care Hospitalized COVID-19 Patients: A Preliminary Report.

The outbreak of coronavirus disease 2019 (COVID-19) is posing a threat to global health. This disease has different clinical manifestations and different outcomes. The immune response to the novel 2019 coronavirus is complex and involves both innate and adaptive immunity. In this context, cell-mediated immunity plays a vital role in effective immunity against SARS-CoV-2. Significant differences have been observed when comparing severe and non-severe patients. Since these immunological characteristics have not been fully elucidated, we aimed to use cluster analysis to investigate the immune cell patterns in patients with COVID-19 who required hospitalization but not intensive care. We identified four clusters of different immunological patterns, the worst being characterized by total lymphocytes, T helper lymphocytes CD4+ (CD4+), T cytotoxic lymphocytes CD8+ (CD8+) and natural killer (NK) cells below the normal range, together with natural killer lymphocyte granzyme < 50% (NK granzyme+) and antibody-secreting plasma cells (ASCs) equal to 0 with fatal outcomes. In the worst group, 50% of patients died in the intensive care unit. Moreover, a negative trend was found among four groups regarding total lymphocytes, CD4+, CD8+ and B lymphocytes (p < 0.001, p < 0.005, p < 0.000, p < 0.044, respectively). This detailed analysis of immune changes may have prognostic value. It may provide a new perspective for identifying subsets of COVID-19 patients and selecting novel prospective treatment strategies. Notwithstanding these results, this is a preliminary report with a small sample size, and our data may not be generalizable. Further cohort studies with larger samples are necessary to quantify the prognostic value's weight, according to immunological changes in COVID-19 patients, for predicting prognoses and realizing improvements in clinical conditions.

Multidrug-resistant Gram-negative post-neurosurgical meningitis and the role of intraventricular colistin: a case series.

The aim of this study was to report the clinical experience of intraventricular colistin for the treatment of multi-resistant Gram-negative post-surgical meningitis in a tertiary hospital. Post-neurosurgical meningitis (PNM) is one of the life-threatening complications of neurosurgical procedures, and is frequently sustained by Acinetobacter baumannii and Klebsiella pneumoniae. Here we describe our experience of five cases of PNM caused by gram-negative multi-drug resistant (MDR) bacteria, treated with intraventricular (IVT) colistin, admitted to the Neurosurgery Unit of A.R.N.A.S. Civico of Palermo, Italy, from January 2016 to June 2020. In four patients the cerebrospinal fluid (CSF) culture was positive for A. baumannii, while in one patient it was positive for K. pneumoniae. IVT colistin therapy was administered for a median time of 18 days (range 7-29). The median time to CSF negativization was seven days (range 5-29). IVT colistin administration was associated with intravenous administration of meropenem and colistin in all patients. As regards clinical outcome, four patients were successfully treated and were subsequently discharged, while one patient died following respiratory complications and subsequent brain death. IVT colistin administration is an effective therapy for MDR post-neurosurgical meningitis and its administration is also prescribed by guidelines. However, IVT therapy for Gram-negative ventriculitis is mostly understudied. Our paper adds evidence for such treatment that can actually be considered life-saving.

Urinary schistosomiasis in migrant population: a case series from a single centre in southern Italy.

PURPOSE: To understand the frequency of urinary schistosomiasis, in migrants in clinical follow-up at the infectious disease outpatient clinic of ARNAS Civico Hospital in Palermo Italy, to raise awareness on this neglected tropical disease. METHODS: A retrospective analysis of migrant patients in clinical care in our centre during the triennium 2015-2017. RESULTS: 2639 migrants have been in clinical care during the triennium 2015-2017, 72% are male and 28% are female. 214 patients were tested for the presence of Schistosoma eggs in urine, these patients are all male. All the patients tested, reported macroscopic haematuria and the 54% had an increase in the peripheral blood eosinophil count. Ninety subjects had a positive microscopic examination for Schistosoma haematobium eggs. Patients were treated with a standard dose of praziquantel (40 mg/kg), and tested for Schistosoma 1 month after the end of therapy. All the subjects fully recovered. CONCLUSIONS: Considering the migration phenomenon, the observation of these tropical diseases in European hospitals is becoming more and more common and an increasing number of health care professionals will be dealing with migrants. Searching for haematuria and eosinophilia and then testing for Schistosoma in this specific population will increase the number of diagnosis and correct treatment of urinary schistosomiasis, improving the patients' quality of life and preventing severe complications of the disease.

Norovirus GII.17 as Major Epidemic Strain in Italy, 2015-16.

In winter 2015-16, norovirus GII.17 Kawasaki 2014 emerged as a cause of sporadic gastroenteritis in children in Italy. Median patient age was higher for those with GII.17 than GII.4 infection (55 vs. 24 months), suggesting limited cross-protection for older children.

Imported and Indigenous cases of Invasive Meningocococcal Disease W:P1.5,2:F1-1: ST-11 in migrants' reception centers. Italy, June-November 2014.

We report about three unliked cases of meningococcal meningitis caused by the ST-11/ET-37 strain of Neisseria meningitidis serogroup W. Two of the three cases, detected in Sicily on June and July 2014, were migrants from Mali and Eritrea. The third case was a fatal meningitis occurred on November 2014 in a 37 years old man, working in an immigrant center in Calabria. This report suggests that tetravalent conjugate vaccines (ACYW) should be actively offered to the staff of migrants' reception centers.

An Update of the Evolving Epidemic of blaKPC Carrying Klebsiella pneumoniae in Sicily, Italy, 2014: Emergence of Multiple Non-ST258 Clones.

BACKGROUND: In Italy, Klebsiella pneumoniae carbapenemase producing K. pneumoniae (KPC-Kp) strains are highly endemic and KPC producing CC258 is reported as the widely predominating clone. In Palermo, Italy, previous reports have confirmed this pattern. However, recent preliminary findings suggest that an epidemiological change is likely ongoing towards a polyclonal KPC-Kp spread. Here we present the results of molecular typing of 94 carbapenem non susceptible K. pneumoniae isolates detected during 2014 in the three different hospitals in Palermo, Italy. METHODS AND RESULTS: Ninety-four consecutive, non replicate carbapenem non susceptible isolates were identified in the three largest acute general hospitals in Palermo, Italy, in the six-month period March-August 2014. They were characterized by PCR for ß-lactam, aminoglycoside and plasmid mediated fluoroquinolone resistance genetic determinants. The mgrB gene of the colistin resistant isolates was amplified and sequenced. Clonality was assessed by pulsed field gel electrophoresis and multilocus sequence typing. Eight non-CC258 sequence types (STs) were identified accounting for 60% of isolates. In particular, ST307 and ST273 accounted for 29% and 18% of isolates. CC258 isolates were more frequently susceptible to gentamicin and non-CC258 isolates to amikacin. Colistin non susceptibility was found in 42% of isolates. Modifications of mgrB were found in 32 isolates. CONCLUSIONS: Concurrent clonal expansion of some STs and lateral transmission of genetic resistance determinants are likely producing a thorough change of the KPC-Kp epidemiology in Palermo, Italy. In our setting mgrB inactivation proved to substantially contribute to colistin resistance. Our findings suggest the need to continuously monitor the KPC-Kp epidemiology and to assess by a nationwide survey the possible shifting towards a polyclonal epidemic.

Data mining from a 27-years rotavirus surveillance in Palermo, Italy.

Uninterrupted surveillance conducted in Palermo, Sicily, for 27 years (1985-2012) detected rotavirus infection in 32.7% of 6522 children <5 years of age, hospitalised at the "G. Di Cristina" Children's Hospital of Palermo. Increased rotavirus activity usually occurred from the beginning of winter to mid-spring. G1P[8] rotaviruses were the prevalent strains in most of the years and were only occasionally overcome by G9P[8], G4P[8] or G2P[4]. The circulation of non-G1P[8] strains was discontinuous and fluctuating. Phylogenetic analyses revealed an heterogeneous population of viruses within each genotype, with different lineages and sublineages emerging over the time. Amino acid substitutions in both VP7 and VP8(∗) antigenic epitopes were generally associated with different lineages/sublineages, emerging sequentially and replacing partially or completely the former strains. The present study summarises one of the longest surveillance activities conducted in the European continent, offering a useful temporal observatory of rotavirus epidemiology and strains variation and evolution in a settled population.

Extended-spectrum beta-lactamase-producing and carbapenemase-producing Enterobacter cloacae ventriculitis successfully treated with intraventricular colistin.

We present a case of post-neurosurgical ventriculitis caused by carbapenemase-producing Enterobacter cloacae successfully treated with intraventricular colistin. Enterobacter spp are intrinsically resistant to aminopenicillins, cefazolin, and cefoxitin due to the production of constitutive chromosomal AmpC beta-lactamases. Moreover, extended-spectrum beta-lactamase-producing Enterobacter spp have been identified in the USA and Europe, and carbapenems are considered the drug of choice in these cases. Our isolate was sensitive only to fosfomycin, tigecycline, and colistin, and 6 days of intravenous colistin had failed to eradicate the infection. This case provides clinical evidence to support the administration of intraventricular colistin in such patients.

Co-colonization with carbapenem-resistant Klebsiella pneumoniae and Acinetobacter baumannii in intensive care unit patients.

OBJECTIVES: This investigation was conducted to study co-colonization by carbapenem-resistant Klebsiella pneumoniae carbapenemase-producing K. pneumoniae (KPC-Kp) and Acinetobacter baumannii (CRAB) in intensive care unit (ICU) patients in Palermo, Sicily, a geographic area where both organisms are endemic in the healthcare setting. Risk factors at admission and during ICU stay and outcomes were also evaluated. METHODS: All patients colonized by KPC-Kp, or CRAB, or both in 2 ICUs of a large general hospital during the period October 2011-March 2012 were enrolled. Demographics and clinical data were collected. Resistance determinants and clonality of the 2 organisms were characterized by molecular methods. RESULTS: Seventy-five of 391 patients (19.2%) proved to be colonized by KPC-Kp, CRAB, or both: 30 (40%) were co-colonized and 44 (58.7%) were mono-colonized by CRAB and 1 by KPC-Kp. Younger age, major trauma, and length of stay were positively associated with co-colonization. However, no significant differences were detected between co-colonized and non co-colonized patients in infection and ICU mortality rates and length of stay after the first isolation. Both organisms proved to be circulating in a clonal way. CONCLUSIONS: In our setting, co-colonization by KPC-Kp and CRAB disproportionately affected young trauma patients with those with a prolonged ICU stay.

[Campylobacter enteritis in Western Sicily. Remarks on 35 cases]. / Il ruolo di Campylobacter spp quale agente di enterite in Sicilia Occidentale. Considerazioni su 35 casi.

INTRODUCTION: Campylobacter spp is the main cause of bacterial gastroenteritis in the developed countries, resulting mainly from the contamination of poultry and animal products. Pathogenesis remains unclear. Various clinical features ranging from watery diarrhoea in apyrexia to dysentery in hyperpyrexia may be the result of differences between strains in the expression of different pathogenetic factors. There are many difficulties in isolating the bacteria in question. METHODS: We processed 540 faecal specimens of children with gastroenteritis. We detected enteropathogen bacteria including Campylobacter spp as well as Rotavirus, Adenovirus and protozoans. For the isolation of Campylobacter we employed the filter membrane technique. The filtrate was plated onto non-selective agar (blood-agar) in microaerophilic atmosphere. RESULTS: The rank order of isolation frequency was: Rotavirus (30.0%), Salmonella (18.5%), Adenovirus (12.6%), Campylobacter (6.5%), Shigella (1.1%), Giardia (0.6%). The Campylobacter strains belonged to C.jejuni in 31 cases and C. coli in 4 cases. The clinical features were: fever (80%), vomitus (31,4%) and diarrhoea in all cases (watery diarrhoea (25.7%), stools more compact with mucus and blood (57.1%) or with mucus alone (17.1%)). CONCLUSIONS: In Western Sicily Campylobacter spp is the second most common agent of bacterial enteritis. We believe that some procedural errors, such as the employment of excessively selective media, could be the cause of the past failure to isolate Campylobacter.