Pediatric Lyme Disease in Northern Italy: An 18-Year Single-Center Case Series.

Tracing the profile of pediatric Lyme borreliosis (LB) in Europe is difficult due to the interregional variation in its incidence and lack in notifications. Moreover, the identification of LB can be challenging. This study is an 18-year case series of 130 children and adolescents aged under 18 years referred to the Pediatric Infectious Diseases Unit at L. Sacco Hospital, Milan, with suspicion of LB, between January 2005 and July 2023. The routine serological workup consisted of a two-step process: an initial screening test followed by Western blot (WB). Forty-four (34%) patients were diagnosed with LB. The median age was six years, and 45% were females. Of the children with erythema migrans (EM), 33 (57%) were confirmed as having true EM, and, of these, 4 (12%) were atypical. Ten (23%) patients had early disseminated/late diseases, including facial nerve palsy (n = 3), early neuroborreliosis (n = 1), arthritis (n = 3), relapsing fever (n = 2), and borrelial lymphocytoma (n = 1). No asymptomatic infections were documented. Over seventy percent of confirmed LB cases (n = 31/44) recalled a history of tick bites; in this latter group, 19 (61%) were from the area of the Po River valley in Lombardy. Almost half of the children evaluated for LB complained of non-specific symptoms (fatigue, musculoskeletal pain, skin lesions/rash, and persistent headache), but these symptoms were observed in only two patients with confirmed LB. Most LB cases in our study were associated with EM; two-tier testing specificity was high, but we found frequent non-adherence to international recommendations with regard to the timing of serology, application of the two-step algorithm, and antibiotic over-prescription. Most children were initially assessed for a tick bite or a skin lesion suggestive of EM by a family pediatrician, highlighting the importance of improving awareness and knowledge around LB management at the primary healthcare level. Finally, the strengthening of LB surveillance at the national and European levels is necessary.

SNUPN deficiency causes a recessive muscular dystrophy due to RNA mis-splicing and ECM dysregulation.

SNURPORTIN-1, encoded by SNUPN, plays a central role in the nuclear import of spliceosomal small nuclear ribonucleoproteins. However, its physiological function remains unexplored. In this study, we investigate 18 children from 15 unrelated families who present with atypical muscular dystrophy and neurological defects. Nine hypomorphic SNUPN biallelic variants, predominantly clustered in the last coding exon, are ascertained to segregate with the disease. We demonstrate that mutant SPN1 failed to oligomerize leading to cytoplasmic aggregation in patients' primary fibroblasts and CRISPR/Cas9-mediated mutant cell lines. Additionally, mutant nuclei exhibit defective spliceosomal maturation and breakdown of Cajal bodies. Transcriptome analyses reveal splicing and mRNA expression dysregulation, particularly in sarcolemmal components, causing disruption of cytoskeletal organization in mutant cells and patient muscle tissues. Our findings establish SNUPN deficiency as the genetic etiology of a previously unrecognized subtype of muscular dystrophy and provide robust evidence of the role of SPN1 for muscle homeostasis.

Use of transcriptomics for diagnosis of infections and sepsis in children: A narrative review.

AIM: The aim of this review was to summarise the most recent evidence about the use of omics-based techniques as an instrument for a more rapid and accurate characterisation of respiratory tract infections, neurological infections and sepsis in paediatrics. METHODS: We performed a narrative review using PubMed and a set of inclusion criteria: English language articles, clinical trials, meta-analysis and reviews including only paediatric population inherited to this topic in the last 15 years. RESULTS: The examined studies suggest that host gene expression signatures are an effective method to characterise the different types of infections, to distinguish infection from colonisation and, in some cases, to assess the severity of the disease in children. CONCLUSIONS: 'Omics-based techniques' may help to define the aetiology of infections in paediatrics, representing a useful tool to choose the most appropriate therapies and limit antibiotic resistance.

An unusual way to improve lung function in congenital myopathies: the power of singing.

Congenital myopathies (CMs) are a clinically and genetically heterogeneous group of disorders characterized by early onset weakness, hypotonia and characteristic structural abnormalities in muscle fibres. Hypotonia and weakness can be present at birth or appear in infancy, and a static or slowly progressive clinical course may present with muscle weakness, loss of spontaneous movement, involuntary muscle activity, and muscle atrophy. Often patients develop a restrictive syndrome and respiratory failure and require respiratory support In our case, we described lung improvement and respiratory muscle training due to singing in a young patient, affected by CMs with a poor adherence to non-invasive mechanical ventilation.

The genetic puzzle of a SOD1-patient with ocular ptosis and a motor neuron disease: a case report.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease with a complex genetic architecture, showing monogenic, oligogenic, and polygenic inheritance. In this study, we describe the case of a 71 years-old man diagnosed with ALS with atypical clinical features consisting in progressive ocular ptosis and sensorineural deafness. Genetic analyses revealed two heterozygous variants, in the SOD1 (OMIM*147450) and the TBK1 (OMIM*604834) genes respectively, and furthermore mitochondrial DNA (mtDNA) sequencing identified the homoplasmic m.14484T>C variant usually associated with Leber's Hereditary Optic Neuropathy (LHON). We discuss how all these variants may synergically impinge on mitochondrial function, possibly contributing to the pathogenic mechanisms which might ultimately lead to the neurodegenerative process, shaping the clinical ALS phenotype enriched by adjunctive clinical features.

Stereotactic MR-guided adaptive radiotherapy (SMART) for primary rectal cancer: evaluation of early toxicity and pathological response.

Background: The purpose of this study is to measure the effects of stereotactic MR-guided adaptive radiotherapy (SMART) for rectal cancer patients in terms of early toxicity and pathological response. Materials and methods: For this prospective pilot study, patients diagnosed with locally advanced rectal cancer (LARC) with positive lymph node clinical staging underwent SMART on rectal lesion and mesorectum using hybrid MR-Linac (MRIdian ViewRay). Dose prescription at 80% isodose for the rectal lesion and mesorectum was 40 Gy (8 Gy/fr) and 25 Gy (5 Gy/fr), respectively, delivered on 5 days (3 fr/week). Response assessment by MRI was performed 3 weeks after SMART, then patients fit for surgery underwent total mesorectal excision. Primary endpoint was evaluation of adverse effect of radiotherapy. Secondary endpoint was pathological complete response rate. Early toxicity was graded according to the Common Terminology Criteria for Adverse Events (CTCAE v5.0). Results: From October 2020 to January 2022, twenty patients underwent rectal SMART. No grade 3-5 toxicity was recorded. Twelve patients were eligible for total mesorectal excision (TME). Mean interval between the completion of SMART and surgery was 4 weeks. Pathological downstaging occurred in all patients; rate of pathological complete response (pCR) was 17%. pCR occurred with a prolonged time to surgery (> 7 weeks). Conclusion: To our knowledge, this is the first study to use stereotactic radiotherapy for primary rectal cancer. SMART for rectal cancer is well tolerated and effective in terms of tumor regression, especially if followed by delayed surgery.

Repression of autocrine pheromone signaling leads to fusaric acid over-production.

Fusaric acid (FA), a picolinic acid derivative, is a natural substance produced by a wide variety of fungal plant pathogens belonging to the Fusarium genus. As a metabolite, fusaric acid exerts several biological activities including metal chelation, electrolyte leakage, repression of ATP synthesis, and direct toxicity on plants, animals and bacteria. Prior studies on the structure of fusaric acid revealed a co-crystal dimeric adduct between FA and 9,10-dehydrofusaric acid. During an ongoing search for signaling genes differentially regulating FA production in the fungal pathogen Fusarium oxysporum (Fo), we found that mutants lacking pheromone expression have an increased production of FA compared to the wild type strain. Noteworthy, crystallographic analysis of FA extracted from Fo culture supernatants showed that crystals are formed by a dimeric form of two FA molecules (1:1 molar stoichiometry). Overall, our results suggest that pheromone signaling in Fo is required to regulate the synthesis of fusaric acid.

A coordinated multiorgan metabolic response contributes to human mitochondrial myopathy.

Mitochondrial diseases are a heterogeneous group of monogenic disorders that result from impaired oxidative phosphorylation (OXPHOS). As neuromuscular tissues are highly energy-dependent, mitochondrial diseases often affect skeletal muscle. Although genetic and bioenergetic causes of OXPHOS impairment in human mitochondrial myopathies are well established, there is a limited understanding of metabolic drivers of muscle degeneration. This knowledge gap contributes to the lack of effective treatments for these disorders. Here, we discovered fundamental muscle metabolic remodeling mechanisms shared by mitochondrial disease patients and a mouse model of mitochondrial myopathy. This metabolic remodeling is triggered by a starvation-like response that evokes accelerated oxidation of amino acids through a truncated Krebs cycle. While initially adaptive, this response evolves in an integrated multiorgan catabolic signaling, lipid store mobilization, and intramuscular lipid accumulation. We show that this multiorgan feed-forward metabolic response involves leptin and glucocorticoid signaling. This study elucidates systemic metabolic dyshomeostasis mechanisms that underlie human mitochondrial myopathies and identifies potential new targets for metabolic intervention.

Administrative Databases and Diagnostic Therapeutic and Assistance Paths -PDTA- in the Monitoring Treatment of Rheumatoid Arthritis: The Experience of ATS Pavia.

Background: The current flows of the SSN represent the set of interest whose interconnection alone justifies the current study. These flows can be interconnected with other sources, institutional or otherwise, in order to answer well-defined questions. Objective: The objective of the study is to verify, through the analysis of administrative databases, any differences in the consumption of health resources between biological off-patent originator drugs and biosimilars in real clinical practice, with particular reference to the rheumatology area. Methods: Through the use of assisted databases (BDA) of ATS Pavia we evaluated the differences in terms of consumption of health resources related to the different drugs under analysis. Annual and daily costs were calculated by total patient cost, stratified for different treatments, considering the sum of total costs for the prescriptions of drugs subject to the analysis. Another objective was to evaluate the adherence of the drugs of interest, by utilizing specific indicators (MPR). Results: A total of 145 patients were analyzed. Among enrolled patients, 26.9% of users were treated with a biosimilar drug, while 73.1% with a biologic originator. Adherence is higher if it is considered the population treated with biosimilar drugs (82.1%). Total cost (including drug prescriptions, hospitalizations, outpatient services, tests for any cause) during the observation period of 1 year is 14,274.08. 87.7% of the total is attributable to drugs. Non-hospitalized patients are the least expensive, whether they were treated with biologics or biosimilars. Conclusion: In our sample, biosimilar drugs tend to be underused: the treatment of a patient with a chronic autoimmune disease is a clinical process that involves many health professionals, and a criticality could also derive from the difficult communication between the various professional figures who get involved with the whole patient treatment.

Economic Evaluation for Palbociclib Plus Fulvestrant vs Ribociclib Plus Fulvestrant and Abemaciclib Plus Fulvestrant in Endocrine-Resistant Advanced or Metastatic Breast Cancer in Italy.

Background: To date, no study evaluated the cost-effectiveness of palbociclib (PAL) plus fulvestrant (FUL) vs ribociclib (RIB) plus FUL and abemaciclib (ABM) plus FUL in Italy. Cost-effectiveness analysis comparing the three cyclin-dependent 4/6 kinase inhibitors in combination with endocrine therapies for the management of postmenopausal women with HR+, HER2- advanced or metastatic breast cancer in Italy was developed. Material and Methods: To assess the cost-effectiveness of PAL plus FUL vs RIB plus FUL and ABM plus FUL, a cost-minimization has been carried out with a conservative scenario considering three CDK4/6 inhibitors with equal effectiveness in terms of overall survival (OS) (MAIC, Rugo et al 2021). Adverse events (AEs) associated with all therapies were obtained from clinical trials. Ad-hoc analysis was performed to estimate the cost-effectiveness considering the quality-of-life (QoL) data (Lloyd et al 2006). Results: Cost-minimization inputs were drugs, visits and exams, AE monitoring and best supportive care (BSC) before the progression state, active and BSC in the progression and terminal phase of the last two weeks of life. Given the comparability of PAL, RIB and ABM in terms of efficacy, this analysis demonstrated slight economic savings over a lifetime for PAL. Results showed saving per patient of €305 (lifetime) when PAL is compared with RIB; for PAL vs ABM a saving of €243 (lifetime) in a conservative scenario. Results of a budget impact analysis showed a potential savings of €319,563 for PAL vs RIB and €297,544 for PAL vs ABM. When QoL data were considered, results may favor PAL due to the lower impact of AE with savings and improvement in the QoL related to fewer AE. Conclusion: From the Italian perspective, a cost-saving profile associated with the use of PAL+FUL for the management of advanced/metastatic HR+/HER2- breast cancer compared to RIB+FUL and ABM+FUL emerged.

The Impact of COVID-19 Pandemic Lockdown on the Relationship between Pediatric MAFLD and Renal Function.

BACKGROUND: Both direct and indirect effects of COVID-19 have been found in all age groups. In particular, adult data demonstrated significant changes in patients with chronic and metabolic disease (e.g., obesity, diabetes, chronic kidney disease (CKD), and metabolic associated fatty liver dysfunction (MAFLD)), while similar pediatric evidence is still limited. We aimed at investigating the impact of the COVID-19 pandemic lockdown on the relationship between MAFLD and renal function in children with CKD due to congenital abnormalities of the kidney and urinary tract (CAKUT). METHODS: A total of 21 children with CAKUT and CKD ≥ stage 1 underwent a comprehensive evaluation within 3 months before and 6 months after the first Italian lockdown. RESULTS: At follow-up, CKD patients with MAFLD presented higher BMI-SDS, serum uric acid, triglycerides, and microalbuminuria levels and lower eGFR levels than those without MAFLD (all p < 0.05). Higher ferritin and white blood cell concentrations were also found in patients with CKD diagnosed with MAFLD than peers without MAFLD (both p = 0.01). Compared to children without MAFLD, a higher delta of BMI-SDS, eGFR levels, and microalbuminuria levels was found in patients with MAFLD. CONCLUSIONS: Due to the negative influence of the COVID-19 lockdown on cardiometabolic health in childhood, a careful management of children with CKD is warranted.

Changes in the Use of Antibiotics for Methicillin-Resistant Staphylococcus aureus Bloodstream Infections in Children: A 5-Year Retrospective, Single Center Study.

Monitoring antibiotic use in the pediatric population is a challenge, especially when determining a relationship between specific pathogens, infections, and antibiotic use. We retrospectively analyzed the consumption of anti-methicillin-resistant Staphylococcus aureus (MRSA) drugs from 2017 to 2021 at Istituto Giannina Gaslini by means of defined daily dose (DDD) adopted for adults by World Health Organization. We observed a statistically significant increase in the use of daptomycin and ceftaroline, combined with a decrease in the use of vancomycin. In the same period, we observed an increase in the proportion of bloodstream infections due to MRSA with vancomycin minimally inhibitory concentration (MIC mg/L) = 1, that represented the 100% of cases in 2021. This aspect was combined with the observation that in the 59% of cases, where vancomycin plasma concentrations were evaluated, it was not possible to achieve a ratio of the 24-h area under the concentration-time curve and MIC (AUC0-24/MIC) of vancomycin ≥ 400 mg/L. This study confirms that DDD can be used in pediatrics to monitor antibiotic consumption in relationship with infections epidemiology. Moreover, it describes the presence of vancomycin MIC creep for MRSA also in pediatrics and the difficulties in obtaining effective vancomycin plasma concentrations in children.

Gut microbiota and COVID-19: An intriguing pediatric perspective.

Gastrointestinal (GI) involvement has been reported in approximately 50% of patients with coronavirus disease 2019 (COVID-19), which is due to the pathogenic role of inflammation and the intestinal function of the angiotensin-converting enzyme 2 and its receptor. Accumulating adult data has pointed out that gut dysbiosis might occur in these patients with a potential impact on the severity of the disease, however the role of gut microbiota in susceptibility and severity of COVID-19 disease in children is still poorly known. During the last decades, the crosstalk between gut and lung has been largely recognized resulting in the concept of "gut-lung axis" as a central player in modulating the development of several diseases. Both organs are involved in the common mucosal immune system (including bronchus-associated and gut-associated lymphoid tissues) and their homeostasis is crucial for human health. In this framework, it has been found that the role of GI dysbiosis is affecting the homeostasis of the gut-liver axis. Of note, a gut microbiome imbalance has been linked to COVID-19 severity in adult subjects, but it remains to be clarified. Based on the increased risk of inflammatory diseases in children with COVID-19, the potential correlation between gut microbiota dysfunction and COVID-19 needs to be studied in this population. We aimed to summarize the most recent evidence on this striking aspect of COVID-19 in childhood.

Economic Burden of Non-Alcoholic Steatohepatitis (NASH) Among Diabetic Population in Italy: Analysis and Perspectives.

Background: Aim of our study is to evaluate the economic impact of NASH among diabetic population in Italy and potential benefits of treatments that can slow the disease progression. Methods: A Markov model was conducted from the Italian National Healthcare System perspective reporting results at 3, 5, 10 and 15 years. The model included NASH and T2DM patients with all stages of fibrosis (F0-F3), compensated cirrhosis (CC), decompensated cirrhosis (DCC), hepatocellular carcinoma (HCC), liver transplant (LT), post-LT and death. A 1-year model cycle length was considered, with each patient passing through the stages and exiting the model when reached one of mortality states. Transition probabilities and annual cost related to health states were derived from published literature. Moreover, the model made it possible to develop a scenario analysis to simulate the impact of treatments capable of slowing the disease progression in phases F0-F4 (CC). Results: The results highlighted an economic burden of NASH in T2DM patients of approximately € 1.4 billion, € 3.1 billion, and € 9.4 billion, respectively, after 3, 5 and 10 years, reaching about € 17.3 billion after 15 years. The slowing down of the progression in the early stages of the disease (fibrosis F0-CC) has led to significant savings corresponding to € 2.3 billion at 15 years. These savings were generated by the reduction of the patients in the advanced stages of the disease, which is linked to a reduction in deaths, equal to 92,208 deaths avoided over a 15-year time horizon. Conclusion: Patients with NASH and T2DM reported an important burden in Italy. It is important to investigate the potential clinical and economic benefits of antidiabetic drugs that have been shown to be effective in preventing the transition to advanced disease, simultaneously acting on the therapeutic goals of diabetic disease.

Rapamycin rescues mitochondrial dysfunction in cells carrying the m.8344A > G mutation in the mitochondrial tRNALys.

BACKGROUND: Myoclonus, Epilepsy and Ragged-Red-Fibers (MERRF) is a mitochondrial encephalomyopathy due to heteroplasmic mutations in mitochondrial DNA (mtDNA) most frequently affecting the tRNALys gene at position m.8344A > G. Defective tRNALys severely impairs mitochondrial protein synthesis and respiratory chain when a high percentage of mutant heteroplasmy crosses the threshold for full-blown clinical phenotype. Therapy is currently limited to symptomatic management of myoclonic epilepsy, and supportive measures to counteract muscle weakness with co-factors/supplements. METHODS: We tested two therapeutic strategies to rescue mitochondrial function in cybrids and fibroblasts carrying different loads of the m.8344A > G mutation. The first strategy was aimed at inducing mitochondrial biogenesis directly, over-expressing the master regulator PGC-1α, or indirectly, through the treatment with nicotinic acid, a NAD+ precursor. The second was aimed at stimulating the removal of damaged mitochondria through prolonged rapamycin treatment. RESULTS: The first approach slightly increased mitochondrial protein expression and respiration in the wild type and intermediate-mutation load cells, but was ineffective in high-mutation load cell lines. This suggests that induction of mitochondrial biogenesis may not be sufficient to rescue mitochondrial dysfunction in MERRF cells with high-mutation load. The second approach, when administered chronically (4 weeks), induced a slight increase of mitochondrial respiration in fibroblasts with high-mutation load, and a significant improvement in fibroblasts with intermediate-mutation load, rescuing completely the bioenergetics defect. This effect was mediated by increased mitochondrial biogenesis, possibly related to the rapamycin-induced inhibition of the Mechanistic Target of Rapamycin Complex 1 (mTORC1) and the consequent activation of the Transcription Factor EB (TFEB). CONCLUSIONS: Overall, our results point to rapamycin-based therapy as a promising therapeutic option for MERRF.

A Second Case With the V374A KCND3 Pathogenic Variant in an Italian Patient With Early-Onset Spinocerebellar Ataxia.

Background and Objectives: To date, approximately 20 heterozygous mainly loss-of-function variants in KCND3 have been associated with spinocerebellar ataxia (SCA) type 19 and 22, a clinically heterogeneous group of neurodegenerative disorders. We aimed at reporting the second patients with the V374A KCND3 mutation from an independent family, confirming its pathogenic role. Methods: We describe the clinical history of a patient with SCA and conducted genetic investigations including mitochondrial DNA analysis and exome sequencing. Results: This male patient was reported to have unstable gait with tremors at the lower limbs and dysarthric speech since childhood. A neurologic examination also showed dysarthria, nystagmus, action tremor, dysmetria, and weak deep tendon reflexes. He had marked cerebellar atrophy at brain MRI, more evident at vermis. Molecular analysis, including exome sequencing and an in silico panel analysis of genes associated with SCA, revealed the c.1121T>C [p.V374A] mutation in KCND3. Discussion: This report consolidates the pathogenicity of the V374A KCND3 mutation and suggests that the ataxic paroxysmal exacerbations are not a key phenotypic feature of this mutation.

Mammalian RNase H1 directs RNA primer formation for mtDNA replication initiation and is also necessary for mtDNA replication completion.

The in vivo role for RNase H1 in mammalian mitochondria has been much debated. Loss of RNase H1 is embryonic lethal and to further study its role in mtDNA expression we characterized a conditional knockout of Rnaseh1 in mouse heart. We report that RNase H1 is essential for processing of RNA primers to allow site-specific initiation of mtDNA replication. Without RNase H1, the RNA:DNA hybrids at the replication origins are not processed and mtDNA replication is initiated at non-canonical sites and becomes impaired. Importantly, RNase H1 is also needed for replication completion and in its absence linear deleted mtDNA molecules extending between the two origins of mtDNA replication are formed accompanied by mtDNA depletion. The steady-state levels of mitochondrial transcripts follow the levels of mtDNA, and RNA processing is not altered in the absence of RNase H1. Finally, we report the first patient with a homozygous pathogenic mutation in the hybrid-binding domain of RNase H1 causing impaired mtDNA replication. In contrast to catalytically inactive variants of RNase H1, this mutant version has enhanced enzyme activity but shows impaired primer formation. This finding shows that the RNase H1 activity must be strictly controlled to allow proper regulation of mtDNA replication.

TWNK in Parkinson's Disease: A Movement Disorder and Mitochondrial Disease Center Perspective Study.

BACKGROUND: Parkinsonian features have been described in patients harboring variants in nuclear genes encoding for proteins involved in mitochondrial DNA maintenance, such as TWNK. OBJECTIVES: The aim was to screen for TWNK variants in an Italian cohort of Parkinson's disease (PD) patients and to assess the occurrence of parkinsonism in patients presenting with TWNK-related autosomal dominant progressive external ophthalmoplegia (TWNK-adPEO). METHODS: Genomic DNA of 263 consecutively collected PD patients who underwent diagnostic genetic testing was analyzed with a targeted custom gene panel including TWNK, as well as genes causative of monogenic PD. Genetic and clinical data of 18 TWNK-adPEO patients with parkinsonism were retrospectively analyzed. RESULTS: Six of 263 PD patients (2%), presenting either with isolated PD (n = 4) or in combination with bilateral ptosis (n = 2), carried TWNK likely pathogenic variants. Among 18 TWNK-adPEO patients, 5 (28%) had parkinsonism. CONCLUSIONS: We show candidate TWNK variants occurring in PD without PEO. This finding will require further confirmatory studies. © 2022 Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson Movement Disorder Society.