Epstein-Barr virus reactivation in pediatric allogeneic stem cell transplant recipients: an 11-year experience on viral load and B lymphocyte monitoring strategy.

Background: Epstein-Barr virus (EBV) reactivation represents a frequent condition after allogeneic hematopoietic stem cell transplantation (allo-HCT) and can cause the development of a severe complication: post-transplant lymphoproliferative disease (PTLD). This retrospective study aims at investigating the incidence of EBV reactivations and analyzing the potential impact of recipient/donor-related transplant-related factors in pediatric patients. The secondary objective was to study the consequences of the approach used at our center regarding the initiation of pre-emptive therapy.

Monitoring and Management of Cytomegalovirus Reactivations After Allogeneic Hematopoietic Stem Cell Transplantation in Children: Experience from a Single Pediatric Center.

Background: CMV reactivation represents a frequent complication after HSCT. The aim of this study was to describe the incidence of CMV reactivation in a pediatric HSCT cohort and analyze the potential impact of recipient/donor-related or transplant-related factors on this complication. Furthermore, we analyzed the management of CMV reactivation in order to purpose criteria for pre-emptive therapy.

Gut-microbiota in children and adolescents with obesity: inferred functional analysis and machine-learning algorithms to classify microorganisms.

The fecal microbiome of 55 obese children and adolescents (BMI-SDS 3.2 ± 0.7) and of 25 normal-weight subjects, matched both for age and sex (BMI-SDS - 0.

Pediatric SARS-CoV-2-Related Diplopia and Mesencephalic Abnormalities.

Objective: This case report describes a patient with mesencephalic MRI signal abnormality and diplopia, possibly associated with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection.

Methods: We describe a boy with binocular diplopia and nystagmus. The pattern of serology positivity and negative direct research of SARS-CoV-2 RNA in our patient allowed us to consider novel coronavirus as the trigger of possible immune-mediated phenomena against the central nervous system.

Contact tracing, use of surgical masks, hand hygiene and social distancing represent a bundle of effective measures to control SARS-CoV-2 spreading among healthcare workers in a paediatric hospital.

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) is the causative agent of pandemic coronavirus disease 2019 (COVID 19) Protection from virus exposure in children's hospital is a pivotal aspect of SARS-COV-2 pandemic control. Healthcare workers (HCW) could play an important role in viral infection in-hospital spread. Infection control measures were thus implemented to protect fragile patients and healthcare workers.

Pathways and microbiome modifications related to surgery and enterocolitis in Hirschsprung disease.

Background: Hirschsprung disease (HSCR) is a congenital anomaly of the enteric nervous system. Abnormal microbiome composition was reported in HSCR patients. In this study, we addressed and analyzed microbiome modifications with relation tosurgery and HSCR associated enterocolitis (HAEC).

Gut Microbiota in T1DM-Onset Pediatric Patients: Machine-Learning Algorithms to Classify Microorganisms as Disease Linked.

Aims: The purpose of this work is to find the gut microbial fingerprinting of pediatric patients with type 1 diabetes.

Methods: The microbiome of 31 children with type 1 diabetes at onset and of 25 healthy children was determined using multiple polymorphic regions of the 16S ribosomal RNA. We performed machine-learning analyses and metagenome functional analysis to identify significant taxa and their metabolic pathways content.

A fast and reliable method for detecting SNP rs67384697 (Hsa-miR-148a binding site) by a single run of allele-specific real-time PCR.

Surface expression of human leukocyte antigen (HLA)-class I molecules is critical for modulating T/natural killer lymphocytes' effector functions. Among HLA molecules, HLA-C, the most recently evolved form of class I antigens, is subjected to both transcriptional and multiple post-transcriptional regulation mechanisms affecting its cell surface expression. Among the latter a region placed in the 3' untranslated region of HLA-C transcript contains the single nucleotide polymorphism (SNP) rs67384697 "G-ins/del" that has been found to be strictly associated with surface levels of HLA-C allomorphs because of the effect on the binding site of a microRNA (Hsa-miR-148a).

A Novel System to Discriminate HLA-C mir148a Binding Site by Allele-Specific Quantitative PC R.

The levels of expression of the HLA-class I molecules are critical for modulating T/NK lymphocytes effector functions. Among HLA molecules, HLA-C, the most recent developed form of class I antigens, is subjected to multiple post transcriptional level of regulation that affect its cell surface expression.We describe a new method of allele-specific real-time PCR that monitor the integrity/disruption of the binding site of the microRNA Hsa-miR-148a, a key factor associated to the levels of HLA-C expression in the Caucasian populations.

Adverse events linked with the use of chimeric and humanized anti-CD20 antibodies in children with idiopathic nephrotic syndrome.

Aims: Anti-CD20 antibodies are increasingly being used to treat idiopathic nephrotic syndrome (INS) in children. While they may allow steroid and calcineurin inhibitor withdrawal, repeated infusions of anti-CD20 antibodies are often required to maintain remission. Data on their potential toxicity in INS are needed, to consider repeated infusions.

Activating Killer Immunoglobulin Receptors and HLA-C: a successful combination providing HIV-1 control.

Several studies demonstrated a relevant role of polymorphisms located within the HLA-B and -C loci and the Killer Immunoglobulin Receptors (KIRs) 3DL1 and 3DS1 in controlling HIV-1 replication. KIRs are regulatory receptors expressed at the surface of NK and CD8+ T-cells that specifically bind HLA-A and -B alleles belonging to the Bw4 supratype and all the -C alleles expressing the C1 or C2 supratype. We here disclose a novel signature associated with the Elite Controller but not with the long-term nonprogressor status concerning 2DS activating KIRs and HLA-C2 alleles insensitive to miRNA148a regulation.

Moyamoya vasculopathy shows a genetic mutational gradient decreasing from East to West.

Background: Moyamoya disease (MMD) is a chronic, occlusive cerebrovascular disease characterized by bilateral steno-occlusive changes at the terminal portion of the internal carotid arteries and an abnormal vascular network at the base of the brain determining stroke in children. Patients with a similar vasculopathy and associated conditions are affected by the moyamoya syndrome (MMS). Most of the studies focused on MMD were carried out on East-Asian population.

Real-time PCR detection of Mycoplasma pneumoniae in the diagnosis of community-acquired pneumonia.

Polymerase chain reaction is a useful technique in microbial diagnostics to detect and quantify DNA or RNA of low abundance. Bacterial and viral nucleic acid can be amplified by PCR upon clinical sample extraction using specific primers for classical qualitative PCR and primers and probes for real-time PCR. Here we describe the Scorpion-probe real-time PCR-based assay that offers thermodynamic advantages due to its kinetic reaction and provides faster performances compared to a classical double-labeled probe-based assays.

The administration of a polyvalent mechanical bacterial lysate in elderly patients with COPD results in serological signs of an efficient immune response associated with a reduced number of acute episodes.

The administration of a polyvalent mechanical bacterial lysate (PMBL) in elderly patients with COPD has been shown to reduce the number of exacerbation. This is largely related to the involvement of cells belonging to the innate and the adaptive immune system (including dendritic cells, granulocytes, T and B lymphocytes and NK cells) that actively cooperate inducing the production of specific opsonizing antibodies directed to the antigens of PMBL. We have evaluated the production of antibodies directed to respiratory and systemic pathogens in a group of elderly COPD patients, recruited in a clinical trial, ancillary to a larger multicenter double blind, placebo-controlled, parallel-designed clinical trial in which patients were randomized to daily receive either PMBL or placebo.

Development and validation of a multiplex quantitative polymerase chain reaction assay for the detection of Mollicutes impurities in human cells, cultured under good manufacturing practice conditions, and following European Pharmacopoeia requirements and the International Conference on Harmonization guidelines.

Background Aims: The clinical applications of in vitro manipulated cultured cells and their precursors are often made use of in therapeutic trials. However, tissue cultures can be easily contaminated by the ubiquitous Mollicutes micro-organisms, which can cause various and severe alterations in cellular function. Thus methods able to detect and trace Mollicutes impurities contaminating cell cultures are required before starting any attempt to grow cells under good manufacturing practice (GMP) conditions.

Detection of ganciclovir resistance mutations by pyrosequencing in HCMV-infected pediatric patients.

Background: Human cytomegalovirus (HCMV) is an opportunistic pathogen especially for immuno-suppressed subjects that might develop pharmacological resistance in patients undergoing prolonged antiviral treatment. Ganciclovir (GCV) is the drug used as first choice therapy in affected children and a GCV-resistant phenotype is mainly linked to mutations of the viral protein kinase UL97.

Objectives: Here a new quantitative pyrosequence (PSQ) method is presented that allows detection and quantification of the viral species carrying the more frequent UL97 mutations responsible for GCV resistance in clinical samples (>80% of known cases).

A multiplex calibrated real-time PCR assay for quantitation of DNA of EBV-1 and 2.

Accurate and highly sensitive tests for the diagnosis of active Epstein-Barr virus (EBV) infection are essential for the clinical management of individuals infected with EBV. A calibrated quantitative real-time PCR assay for the measurement of EBV DNA of both EBV-1 and 2 subtypes was developed, combining the detection of the EBV DNA and a synthetic DNA calibrator in a multiplex PCR format. The assay displays a wide dynamic range and a high degree of accuracy even in the presence of 1μg of human genomic DNA.

Molecular characterization of hospital-acquired methicillin-resistant Staphylococcus aureus strains in pediatric outbreaks using variable tandem repeat analysis with spa and ClfB typing.

To analyze 67 clinical methicillin-resistant Staphylococcus aureus (MRSA) isolated from pediatric hospital infections, we used multilocus variable-number tandem-repeat DNA sequence-based techniques, targeting the protein A polymorphic X region and the clumping factor B complete R domain. We define a "clfB similarity score" and then compare the double loci analysis of closely related MRSA isolates with pulsed-field gel electrophoresis (PFGE). We found an endemic clone (MLST-ST8, spa-t008, SCCmecIV, ClfB lineage 1) able to originate 3 possible outbreaks and a second clone (MLST-ST152, spa-t355, SCCmecV, ClfB lineage 4) responsible for limited cases of MRSA infections, indicating that the combination of spa and clfB-lineage typing is useful to trace MRSA pediatric outbreaks.

HHV-8-related visceral Kaposi's sarcoma following allogeneic HSCT: report of a pediatric case and literature review.

An HHV-8-related visceral KS was diagnosed in a 10-yr-old boy after partially matched allogeneic HSCT. This complication occurred 463 days after HSCT and involved tonsils, lymph nodes, hard palate, lung, skin, and paranasal sinuses. Treatment with pegylated liposomal doxorubicin induced long-term remission (33 months) of this disease.

NKp44 expression, phylogenesis and function in non-human primate NK cells.

Molecular and functional characterization of the natural cytotoxicity receptor (NCR) NKp44 in species other than Homo sapiens has been elusive, so far. Here, we provide complete phenotypic, molecular and functional characterization for NKp44 triggering receptor on Pan troglodytes NK cells, the closest human relative, and the analysis of NKp44-genomic locus and transcription in Macaca fascicularis. Similar to H.

Low dosage cidofovir without probenecid as treatment for BK virus hamorrhagic cystitis after hemopoietic stem cell transplant.

We describe a single-center pediatric experience with 1 mg/kg/wk cidofovir without probenecid in 7 children with BK virus-associated hemorrhagic cystitis. Clinical improvement was observed in all cases, without adverse events, although significant reduction of urinary viral load was observed 2 weeks after the end of cidofovir in 5 out of 6 patients who completed the treatment.

Patients with oral erosive and cutaneous lichen planus may have antibodies directed against the chronic ulcerative stomatitis protein antigen of 70-kDa.

Objective: The aim of this study is to verify whether stratified epithelium-specific antinuclear antibodies are present in the sera of patients with erosive oral lichen planus and cutaneous lichen planus.

Methods: We studied the pre-immune and immune serum of a rabbit immunized with a peptide corresponding to the N-terminus of the 70-kDa antigen chronic ulcerative stomatitis protein; sera from two patients, one with oral erosive lichen planus and one with cutaneous lichen planus who presented stratified epithelium-specific antinuclear antibodies at high titer; and a third serum from a patient with cutaneous lichen planus without stratified epithelium-specific antinuclear antibodies.

Results: We demonstrated that the protein bands recognized by the serum of the rabbit immunized with an epitope of chronic ulcerative stomatitis protein co-migrated by SDS-PAGE with the protein bands recognized by the serum of a patient affected by oral erosive lichen planus and by the serum of a patient with cutaneous lichen planus, both containing antibodies directed against a 70-kDa antigen.