Does multiple gastric aspirate collection increase sensitivity of M. tuberculosis detection in children with pulmonary tuberculosis?

This study aims to investigate the sensitivity of microscopy, culture and polymerase chain reaction on three gastric aspirates (GAs) in the microbiological confirmation of active pulmonary tuberculosis (TB) and to identify possible changes in sensitivity derived from the collection of a different number of aspirates. Children with clinical and radiological diagnoses of active pulmonary TB who underwent three GAs between March 2007 and June 2019 were retrospectively evaluated. Clinical, radiological, and microbiological data were collected. The sensitivity of microbiological tests on GAs was calculated. Moreover, differences in sensitivity according to age and radiological pattern were investigated. Overall, 156 children with active pulmonary TB were enrolled with a median age of 51.5 (IQR: 25.2-113.2) months. Microbiological investigations on the first GA showed a sensitivity of 34% (95%CI 26.7, 42), the cumulative sensitivity of first and second GAs was 40.4% (95%CI 32.7, 48.5) and of the three GAs was 47.4% (95%CI 39.8, 55.2). The collection of three GAs leads to an overall increase in sensitivity of the first GA by 13.4% (95%CI 2.8, 24.1%; p=0.014). Moreover, the increase in sensitivity was significantly higher in children ≤ 4 years of age and in those with uncomplicated TB (p=0.008).Conclusions: Performing a higher number of GAs increases the sensitivity of microbiological confirmation of active pulmonary TB, particularly in children ≤ 4 years and with an uncomplicated radiological pattern. What is known: • The diagnosis of paediatric tuberculosis is a challenge for paediatricians • Despite their low sensitivity gastric aspirates represent the standard sample for microbiological confirmation of active pulmonary tuberculosis in children • Most international guidelines recommend performing three sequential gastric aspirates on three consecutive days What is new: • A significant increase in global sensitivity by 13.4% was found by the collection of three gastric aspirates compared to the first one • Performing a higher number of gastric aspirates increases the sensitivity of microbiological confirmation, particularly in children ≤ 4 years and with an uncomplicated radiological pattern.

Artesunate monotherapy versus artesunate plus quinine combination therapy for treatment of imported severe malaria: a TropNet retrospective cohort study.

BACKGROUND: The addition of intravenous quinine (IVQ) to intravenous artesunate (IVA) has been recently suggested by World Health Organization  in areas where artemisinin resistance is highly prevalent. Since IVA is not yet widely available as "Good Manufacturing Practices" product, for several years combination treatment with IVA and IVQ was used in some Italian centers to mitigate the legal risks in using an unlicensed drug. METHODS: A retrospective cohort study was designed to compare IVA + IVQ and IVA treatment for imported severe malaria. We collected data from three Italian centers. Adult and pediatric cohorts were analyzed separately. RESULTS: Forty-nine patients treated with IVA and 44 with IVA + IVQ were enrolled, 45 were adults and 48 children. All acquired malaria in Sub-Saharan Africa. In the adult cohort, median of fever clearance time (FCT) was similar in both groups (48 h vs 48 h, p = 0.19) but number of patients who reached apyrexia within 48 h (FCT48) was higher in IVA group (20/24, 83.3% vs 8/17, 47%, p = 0.002). The parasite clearance time (PCT) measure did not differ (median 48 h vs 48 h, p = 0.669). In the pediatric cohort, FCT did not differ in the two groups (median 30 vs 48 h, p = 0.50) while PCT was longer in IVA + IVQ group (median 72 vs 48 h, p = 0.002). Adverse events (AEs) in adults were more common in the combination treatment group (6/19, 31.58% vs 2/26, 7.69%, p = 0.055). CONCLUSION: IVA + IVQ treatment did not show better outcome with respect to IVA monotherapy. AEs were more frequent in the IVA + IVQ group compared to the monotherapy. Further studies are necessary to investigate whether IVA + IVQ could be an efficient strategy to treat severe malaria cases in areas at high risk of artemisinin resistance.

Real-life isoniazid and rifampicin plasma concentrations in children: a tool for therapeutic drug monitoring of tuberculosis.

BACKGROUND: Low plasma levels of first-line antitubercular drugs can be counted among the main causes of poor response to antitubercular therapy, and therapeutic drug monitoring has been proposed as a method to promote tailored treatments for both child and adult patients. The main aim of the study was to evaluate serum concentrations of isoniazid (INH) and rifampicin (RIF) and to investigate reasons for sub-therapeutic plasma concentrations in order to fix dosages. METHODS: Children with TB were prospectively enrolled from January to August 2019. Two venous blood samples were collected (the first at least 15 days after the beginning of antitubercular treatment, and the second between 1 and 8 weeks later). Plasma concentrations were determined by a validated high-performance liquid chromatography method. RESULTS: In all, 45 children were included. Seventy blood samples for INH plasma concentration were collected between 120 and 240 min after drug intake. Adjusting for dose (mg/kg/day) and time of INH administration, when considering three different age groups (≤ 2 years, 2-12 years, > 12 years), a statistically significant lower INH plasma concentration was observed in younger children compared to the older age groups in the multivariate analysis (p < 0.001 and p < 0.001). A total of 68 blood samples were evaluated for RIF concentrations. Both for INH and RIF a statistically significant lower plasma concentration was also observed in adolescents (p < 0.001). Fifteen children (15/45, 33%) presented drug concentrations under the referral therapeutic range. CONCLUSIONS: Based on our findings, monitoring patients' drug plasma concentrations in children under 2 years of age and in adolescents can make treatment more patient-tailored.

Clinical features and outcomes of lymphadenopathy in a tertiary children's hospital.

AIM: The aim of the present study is to describe the clinical features and outcomes of childhood lymphadenopathy and to define factors able to predict neoplastic aetiology or may improve its prognosis. METHODS: All children evaluated for lymphadenopathy in our tertiary children's hospital and who underwent their first examination between 1 January, 2015 and 31 December, 2017 were enrolled in this retrospective observational study. Data were analysed using SPSS.Statistics, 24.0. RESULTS: A total of 322 children, aged between 0 and 18 years (median 4.5; interquartile range 2.5-9), were enrolled. A specific diagnosis was achieved in almost half of the cases (n = 159, 49.4%) by using one or more methods, including serological, microbiological, biomolecular or histological investigations on surgical samples. Epstein Barr virus and non-tuberculous mycobacteria were the most common etiological agents among acute/sub-acute and chronic lymphadenopathy, respectively. At the end of the study period, two-thirds (210, 65.2%) of enrolled patients were successfully treated. Malignancies and non-tuberculous mycobacteria infections had the longest time to resolution. CONCLUSIONS: Our data suggest that lymphadenopathy is a benign condition in most cases. Of note in our study, 2.5% of lymphadenopathy cases were found to be due to oncologic conditions. The most frequent infective causes were Epstein Barr virus, bacteria and non-tuberculous mycobacteria infections. No haematic or ultrasonographic features were independently able to provide sufficient evidence for a conclusive diagnosis. However, utilising these findings alongside evaluation for clinical criteria can guide decision-making for physicians. Lymphadenectomy is the most appropriate process to follow in the event of chronic lymphadenopathy with undefined diagnosis.

Role of Mycobacterium avium lysate INF-γ, IL-17, and IL-2 ELISPOT assays in diagnosing nontuberculous mycobacteria lymphadenitis in children.

Nontuberculous mycobacteria are the most frequent cause of chronic cervical lymphadenitis in childhood. The aim of the study was to evaluate the performance of IL-2, IL-17, and INF-γ in-house enzyme-linked immunospot assays using a Mycobacterium avium lysate, in order to identify a noninvasive diagnostic method of nontuberculous mycobacteria infection. Children with subacute and chronic lymphadenopathies or with a previous diagnosis of nontuberculous mycobacteria lymphadenitis were prospectively enrolled in the study. Sixty children with lymphadenitis were included in our study: 16 with confirmed infection (group 1), 30 probable infected (group 2) and 14 uninfected (group 3). Significantly higher median cytokine values were found in group 1 vs group 2, in group 1 vs group 3, and in group 2 vs group 3 considering IL-2-based enzyme-linked immunospot assay (p = 0.015, p < 0.001, p = 0.004, respectively). INF-γ-based enzyme-linked immunospot assay results were significantly higher in group 2 vs group 3 (p = 0.010). Differences between infected and uninfected children were not significant considering IL-17 assays (p = 0.431). Mycobacterium avium lysate IL-2 and INF-γ-based enzyme-linked immunospot assays seem to be promising noninvasive diagnostic techniques for discriminating children with nontuberculous mycobacteria lymphadenitis and noninfected subjects.

Should Pediatricians Be Aware of Cystic Echinococcosis? A Literature Review.

Among parasitic hepatic cystic lesions, the most common disease is represented by cystic echinococcosis (CE), especially in high endemic countries. European epidemiology of CE in children is difficult to assess because of under-reporting but is increasing, because of high immigration flows from endemic countries and an increased awareness. Hydatidosis can be localized in every part of the body. The liver and lungs are the most common localizations in both children and adults. Multiorgan involvement is rarely reported in children. Different tests are available. Usually the sensitivity of serological screening tests is variable, ranging between 60% and 90%. The immunoblot assay is used as a confirmatory test because of its higher sensitivity and specificity. Radiological tests are the criterion standard for diagnosis of CE, with an ultrasound accuracy of approximately 90%. In case of inactive and uncomplicated cysts the watch-and-wait approach is recommended. Albendazole, currently used for 3 to 6 months consecutively represents the most commonly used drug in children even if there is limited experience in treating children younger than 6 years of age. Percutaneous treatment with the puncture, aspiration, injection, and reaspiration technique is a minimally invasive procedure. Surgery is indicated based on cyst characteristics in case of big cysts with multiple daughter cysts, single superficial cysts at risk of spontaneous or traumatic rupture, cysts related with the biliary tract in which the percutaneous treatment is contraindicated, and cysts compressing related structures.

Underestimation of Invasive Meningococcal Disease in Italy.

Knowing the incidence of invasive meningococcal disease (IMD) is essential for planning appropriate vaccination policies. However, IMD may be underestimated because of misdiagnosis or insufficiently sensitive laboratory methods. Using a national molecular surveillance register, we assessed the number of cases misdiagnosed and diagnoses obtained postmortem with real-time PCR (rPCR), and we compared sensitivity of rPCR versus culture-based testing. A total of 222 IMD cases were identified: 11 (42%) of 26 fatal cases had been misdiagnosed or undiagnosed and were reclassified as IMD after rPCR showed meningococcal DNA in all available specimens taken postmortem. Of the samples tested with both rPCR and culture, 58% were diagnosed by using rPCR alone. The underestimation factor associated with the use of culture alone was 3.28. In countries such as Italy, where rPCR is in limited use, IMD incidence may be largely underestimated; thus, assessments of benefits of meningococcal vaccination may be prone to error.

Tandem mass spectrometry, but not T-cell receptor excision circle analysis, identifies newborns with late-onset adenosine deaminase deficiency.

BACKGROUND: Adenosine deaminase (ADA)-severe combined immunodeficiency (SCID) is caused by genetic variants that disrupt the function of ADA. In its early-onset form, it is rapidly fatal to infants. Delayed or late-onset ADA-SCID is characterized by insidious progressive immunodeficiency that leads to permanent organ damage or death. Quantification of T-cell receptor excision circles (TRECs) or tandem mass spectrometry (tandem-MS) analysis of dried blood spots (DBSs) collected at birth can identify newborns with early-onset ADA-SCID and are used in screening programs. However, it is not clear whether these analyses can identify newborns who will have delayed or late-onset ADA-SCID before symptoms appear. OBJECTIVE: We performed a retrospective study to evaluate whether tandem-MS and quantitative TREC analyses of DBSs could identify newborns who had delayed-onset ADA-SCID later in life. METHODS: We tested stored DBSs collected at birth from 3 patients with delayed-onset ADA-SCID using tandem-MS (PCT EP2010/070517) to evaluate levels of adenosine and 2'-deoxyadenosine and real-time PCR to quantify TREC levels. We also analyzed DBSs from 3 newborns with early-onset ADA-SCID and 2 healthy newborn carriers of ADA deficiency. RESULTS: The DBSs taken at birth from the 3 patients with delayed-onset ADA-SCID had adenosine levels of 10, 25, and 19 µmol/L (normal value, <1.5 µmol/L) and 2'-deoxyadenosine levels of 0.7, 2.7, and 2.4 µmol/L (normal value, <0.07 µmol/L); the mean levels of adenosine and 2'-deoxyadenosine were respectively 12.0- and 27.6-fold higher than normal values. DBSs taken at birth from all 3 patients with delayed-onset ADA deficiency had normal TREC levels, but TRECs were undetectable in blood samples taken from the same patients at the time of diagnosis. CONCLUSION: Tandem-MS but not TREC quantification identifies newborns with delayed- or late-onset ADA deficiency.

Severe infections caused by Panton-Valentine leukocidin-positive Staphylococcus aureus in infants: report of three cases and review of literature.

UNLABELLED: We report three cases of severe infections in infants caused by Panton-Valentine leukocidin positive Staphylococcus aureus and evolved with a positive outcome. The literature of Panton-Valentine leukocidin positive Staphylococcus aureus infections in infants is reviewed. CONCLUSION: Our findings suggest that a prompt identification of Panton-Valentine leukocidin positive Staphylococcus aureus and an appropriate therapy can reduce mortality and long-term sequelae. Further research is needed to specify features of Panton-Valentine leukocidin positive Staphylococcus aureus infections in infants.

Unbalanced serum immunoglobulins in clinical subtypes of pediatric tuberculosis disease.

Immune response to tuberculosis (TB) has been extensively studied in the past decades and classically involves cellular immunity. However, evidence suggests that humoral immunity may play a relevant role. Past studies regarding serum immunoglobulin (Ig) levels in TB are dated and only involve adult subjects. In this study, we retrospectively studied a cohort of 256 children with TB disease and analyzed 111 patients screened for total serum Ig at diagnosis. According to the severity and extent of organ involvement, subjects were divided into four groups, namely, uncomplicated pulmonary TB (UCPTB, 56.3% of patients), complicated pulmonary TB (CPTB, 22.5%), lymph node extrapulmonary TB (LN-EPTB, 7.2%), and extra-nodal extrapulmonary TB (EN-EPTB, 13.5%). Serum IgG and IgA levels were significantly higher in more severe and extended TB disease. Median IgG levels progressively increased from uncomplicated to complicated pulmonary and nodal forms, reaching their highest values in diffuse extra-pulmonary TB. In parallel, UCPTB showed significantly lower frequencies of patients presenting a substantial increase in IgG levels when compared with the other three groups. No relevant differences in IgM levels were detected. Ig screening at follow-up showed a significant reduction in IgG and IgA levels. Finally, we unveiled three cases of selective IgA and one case of selective IgM deficiencies (SIgMD), the latter with a severe clinical course. Serum IgG and IgA may be a useful clinical tool to assess the severity and monitor the treatment response in pediatric TB disease. Moreover, immunological workup in children with TB disease may unmask primary defects of humoral immunity.

Early limb paralysis in infants: do not forget about congenital syphilis.

We describe the case of a 2-month-old baby with congenital syphilis, presenting with limb paralysis. The radiological investigations showed periosteal thickenings of the limb. Despite negative maternal serology during the first trimester of pregnancy, clinical and radiological features led to the suspicion of pseudoparalysis of Parrot, which was confirmed by blood tests. Delayed diagnoses or misdiagnoses are possible when uncommon presentations of forgotten diseases occur. The needing for a second screening for syphilis in high-risk pregnant women should be evaluated.

Resistance Patterns from Urine Cultures in Children Aged 0 to 6 Years: Implications for Empirical Antibiotic Choice.

PURPOSE: Urinary tract infection (UTI) is a frequent disorder of childhood, caused mainly by Gram negative Enterobacterales. The aim of this study is to evaluate etiology and antimicrobial susceptibility patterns of bacterial isolates in urine cultures of children under the age of 6 and to analyze the relationship between previous hospitalization or antibiotic prescriptions and antimicrobial resistance rates. PATIENTS AND METHODS: A retrospective study on positive urine cultures from 13 public laboratories in Tuscany, Italy was conducted. Data were obtained by reviewing records of the "Microbiological and Antibiotic-Resistance Surveillance System" (SMART) in Tuscany, Italy. A total of 2944 positive urine cultures were collected from 2445 children. RESULTS: Escherichia coli represented the majority of isolates (54,2%), followed by Enterococcus faecalis (12,3%), Proteus mirabilis (10,3%) and Klebsiella pneumoniae (6,6%). Isolated uropathogens showed high resistance rates to amoxicillin-clavulanate (>25%), particularly in children under one year of age or hospitalized within the 12 months before the sample collection. High susceptibility rates were reported of aminoglycosides, cephalosporins and quinolones (>90%). Previous antibiotic prescriptions by general pediatricians did not increase resistance rates. CONCLUSION: Our results show a rate of amoxicillin-clavulanate resistance of 25%. Higher resistance rates were reported in children under one year of age and with previous hospitalization. Hence, amoxicillin-clavulanate should be used carefully in young children and those with severe symptoms.

Epidemiology and clinical features of intestinal protozoan infections detected by Real-time PCR in non-native children within an Italian tertiary care children's hospital: A cross-sectional study.

BACKGROUND: Enteric parasite infections are underestimated due to the limited sensitivity and specificity of microscopy, which remains the diagnostic gold standard in routine clinical practice. This could be a major problem in high-income countries, where the burden of parasitic diseases is low. In recent years, Multiplex Real-Time polymerase chain reaction (RT-PCR) based methods have been implemented. Therefore, the aim of this study was to evaluate the prevalence of four enteric protozoan species detected by RT-PCR in non-native children in Italy, and to describe their clinical characteristics. METHODS: Adopted and immigrant children, evaluated for migration health assessment between 2017 and 2020 in a tertiary care children's hospital in Italy, were enrolled. Molecular analysis for Giardia lamblia, Dientamoeba fragilis, Blastocystis hominis, and Entamoeba histolytica, was conducted by in-house RT-PCR. RESULTS: Overall, 209 children were enrolled and 70% of them resulted positive by RT-PCR for at least one enteric parasite. B. hominis (47.8%) was the most commonly identified protozoa, followed by D. fragilis (44.5%). Co-infections with multiple pathogens were detected in 35.4% of the samples. Almost 80% of parasite-positive children were asymptomatic and the most common symptom was flatulence (60.7% of symptomatic children). Eosinophils were significantly increased in RT-PCR positive children compared to the negative ones and children with D. fragilis presented the highest eosinophils count. CONCLUSIONS: The In-house Multiplex RT-PCR assay provides a valid molecular detection system for selected enteric parasites. This novel and accurate diagnostic method can help in increasing the detection rate of parasite infection, especially in high-risk population.

Strongyloidiasis in Children Outside the Tropics: Do We Need to Increase Awareness?

Strongyloidiasis belongs to the group of neglected tropical diseases, due to diagnostic difficulties and the lack of systematic screening. Studies on strongyloidiasis prevalence are often heterogenous and mainly performed in adults in endemic countries. We retrospectively enrolled 2633 children referred to a tertiary care hospital in Italy between 2009 and 2020 and tested for S. stercoralis infection. Sixty-one (2.3%) had a positive serology and for 55 of them, clinical and epidemiological information were available. Thirteen cases (24%) were diagnosed in Italian children without history residency or travel to foreign countries, while the remaining were internationally adopted or migrant children. Seropositive patients were mostly asymptomatic, and often eosinophilia was the only sign of strongyloidiasis. Sero-reactivity to Toxocara canis was found in 1/3 of patients. Ivermectin was used in 37 (75.5%) treated patients. A significant reduction of eosinophil levels and IgG titer was seen after treatment. Our study confirms that strongyloidiasis is usually asymptomatic in children. However, due to the ability of the parasite to cause a life-long infection together with the risk of a severe form in case of immunosuppression, it is important to identify and treat infected children. Special consideration should be reserved to high-risk groups, such as immigrants and international adoptees, where screening for S. stercoralis is indicated. However, the study highlights that sporadic cases of autochthonous strongyloidiasis in Italy may occur. Therefore, pediatricians should be aware of this condition, which is often under-recognized.

Criteria for referral of pediatric SARS-CoV-2 infection: a real-life experience in the pandemic era.

A practical guidance on the management of children with COVID-19 to insure homogeneous criteria for referral to a higher-level facility, according to the disease severity, is pivotal in the pandemic era. A panel of experts in pediatric infectious diseases and intensive care at the tertiary-care Meyer Children's University Hospital, Florence, Italy, issued a practical document shared with Tuscany hospitals. The rationale was to target the referral for those children at risk of requiring an intensive support, since the above mentioned hospital has the pediatric intensive care unit. Overall, 378 patients between 0 and 19 years of age were diagnosed with COVID-19 infection in the Tuscany region with 24 (6.3%) hospitalizations. Only three children were centralized to Meyer Children's University Hospital according to reported criteria. Considering that appropriate referral criteria have been associated with reduced mortality in other conditions, our document might be useful to improve outcomes of children with COVID-19.