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Aim: To assess parents' attitudes toward childhood COVID-19 immunization in Croatia. Methods: In this multicenter cross-sectional study, we collected data from four tertiary care facilities in Zagreb, Split, and Osijek between December 2021 and February 2022. During the visit to the Pediatric Emergency Departments, parents were asked to fill out a highly-structured questionnaire about their attitudes toward COVID- 19 immunization in children. Results: The sample consisted of 872 respondents. A total of 46.3% of respondents were hesitant about vaccinating their child against COVID-19, 35.2% definitely did not intend to vaccinate their child, and 18.5% definitely intended to vaccinate their child. Parents who were themselves vaccinated against COVID-19 were more likely than unvaccinated parents (29.2% and 3.2%, P

An increase of acute hepatitis of unknown aetiology has been reported among children in multiple countries worldwide. With a rapid online survey among hospitals in and outside of Europe, we describe case numbers recorded from 1 January to 18 April 2022 vs the previous 5 years. Of 24 countries that responded, we identified 5/17 European and 1/7 non-European countries with an elevation in probable cases of unexplained acute hepatitis, and severe cases were elevated in five European countries.

Rotavirus A (RVA) belongs among the leading causes of gastroenteritis in young children and animals, varying in severity from mild to deadly. Rotavirus genome consists of 11-segmented, double-stranded RNA. We aimed to determine the complete genome constellations of the unusual RVA genotypes discovered in Croatian children hospitalized with acute gastroenteritis collected during seasons 2018 to 2020. Following multiplex RT-PCR and Sanger sequencing, samples with host-unusual genotypes were singled out for the Illumina-based whole-genome sequencing. Attained constellations for samples D230-ZG, D329-OB, D248-ZG, and D399-SD were G4-P[6]-I1-R1-C1-M1-A1-N1-T7-E1-H1, G4-P[6]- I1-R1-C1-M1-A1-N1-T1-E1-H1, G6-P[6]-I2-R2-C2-M2- A3-N2-T6-E2-H3, G8-P[14]-I2-R2-C2-M2-A3-N2-T6-E2- H3, respectively. Two G4P[6] strains presented with a Wa-like constellation that evolutionary shares its backbone with porcine RVAs, except for the human-originated NSP1 and NSP2 segments. Conversely, strains G4P[6] and G8P[14] displayed a bovine-like genotype constellation. These two strains revealed a very close evolutionary relationship in all segments, some to the clade level, with the RVAs from the Artiodactyla family, hence a plausible direct zoonotic transmission could be hypothesized. Despite the relatedness with the Artiodactyla, it is interesting to note that during concurrent One Health research, the P[14] genotype within the G8P[14] human RVA strain, was detected only in the Croatian wild canine population. RVA strains with zoonotic background, currently detected sporadically, could be more present in the overall human population, eluding the present study by causing clinically milder, rarely hospitalized cases. Therefore, future surveillance is essential to fully determine the influence the wildlife and domestic animal populations have on the epidemiology of the specified RVA strains circulating in humans.

Rotavirusi, a posebno vrsta rotavirus A (RVA), su glavni uzročnici virusnih gastroenteritisa kod djece i kod mlađih dobnih kategorija životinja. Segmentirani genom rotavirusa odgovoran je za preslagivanje između različitih sojeva uz povremenu pojavu novih, humano-animalnih i moguće emergentnih sojeva. Tijekom tri rotavirusne sezone (2018-2021), prikupili smo ukupno preko 3400 uzoraka fecesa od hospitalizirane djece (RVA ICA pozitivni ; N=602) te domaćih i divljih mesojeda (N=1042), biljojeda (N=882) i svejeda (N=886). Uzorci su pretraženi primjenom metode RT-qPCR za VP2 segment i konvencionalnih metoda pojedinačnih ili multipleks RT-PCR za VP7 i VP4 segmente RVA. Rezultati pokazuju prevalenciju RVA u rasponu od 9, 3-18, 3% kod divljih i 38, 8-49, 9% kod domaćih životinja, dok je većina humanih uzoraka potvrđena kao RVA pozitivna. Genotipizacija VP7 i VP4 segmenata kod humanih uzoraka otkriva prisutnost osam različitih G (G1-G4, G6, G8-G10) i pet različitih P genotipova (P[4], P[6], P[8], P[9], P[14]), uz dominantnu zastupljenost genotipske kombinacije G3P[8]. Svi ovi genotipovi, osim genotipa P[4] i P[9] su utvrđeni kod različitih vrsta životinja. Filogenetska analiza potvrđuje zoonotsku pozadinu u svega 1, 6% humanih RVA sojeva, uslijed preslagivanja genoma sa sojevima podrijetlom od goveda (G6, G8, G10), svinja (G4 i P[6]) i lisica/ čagljeva (G8 i P[14]). Zanimljivo, genotip P[14] se smatra tipičnim za parnoprstaše kod kojih u predmetnom istraživanju nije utvrđen. Međuvrsni prijenos RVA je dokazan i u obrnutom smjeru u vidu reverznog zoonotskog prijenosa s čovjeka na domaće svinje i pse (G1 i P[8]), a moguće i lisice (G1 i G2). Iako je značaj životinjskih sojeva u epidemiologiji rotavirusnih infekcija kod djece u ovom istraživanju marginaliziran, učestalost zoonotskog prijenosa je možebitno veća u populaciji djece koja nisu hospitalizirana uslijed RVA infekcije. Redoviti nadzor cirkulirajućih sojeva RVA u domaćih i divljih sisavaca, uz pristup „Jednog zdravlja“, omogućuje pravovremeno prepoznavanje pojave novih emergentnih sojeva RVA moguće važnih za humano zdravlje i zaštitni učinak dostupnih cjepiva.

Rotavirus A (RVA), in countries like Croatia with low vaccination coverage, represents an important healthcare burden due to frequent hospitalizations. The aim of the study was to investigate rotavirus diversity in hospitalized children in Croatia and to evaluate the potential zoonotic background of circulating strains. During three consecutive RVA seasons (2018-2021), we have collected 602 RVA positive samples from children admitted to three regional hospitals. Multiplex semi-nested RT-PCR and Sanger sequencing were applied for VP7/VP4 genotyping. For G1 and G3 strains, NSP5 and VP6 segments were additionally genotyped and selected strains were further analyzed by NGS on Illumina platform. Overall, we detected eight and five different G and P genotypes, respectively. In descending order, G3, G9, G2, G1, G4, G6, G8, G10 and P[8], P[4], P[9], P[6], P[14]. The G3 genotype was predominant (54%) with rising season-to-season prevalence of G3 equine-like (G3e) lineage. Furthermore, P[8] genotype was detected in 79% of samples. Interestingly, a high share (15%) of mixed G genotypes was observed. On the contrary, strains with zoonotic background were infrequent (1.6%). The NSP5/VP6 genotyping and NGS confirmed the emergence of intergenogroup reassortant strains ; G1P[8] and G3eP[8], both in DS-1-like and Wa-like constellations. Notable genetic heterogeneity and sporadic detection of RVA strains with zoonotic background were reported in hospitalized children in Croatia. A significant finding is the circulation of intergenogroup reassortant strains. These results provide a valuable source of knowledge prerequisite in considering potential introduction of vaccination in the national immunization program.

Rotavirus is the leading cause of severe diarrhea among young children and animals with Rotavirus A (RVA), having the greatest impact considering its prevalence and pathogenicity. The segmented rotavirus genome is enclosed within three layered capsid, with an outer layer consisting of VP4 and VP7 proteins representing the foundation for binary classification. In years 2020 and 2021 we have collected and tested 191 fecal samples mostly from hospitalized children, 92 samples originating from the wider Zagreb area, and 99 from the wider Split area. All the obtained samples have tested positive with the immunochromatographic assay in their respective hospital centers. Using multiplex semi- nested PCR, either G or P genotype was determined in 189 samples and G/P genotype combination was found in 175 samples. For the unattained and the representative strains of each genotype, Sanger sequencing of amplified VP7 and VP4 gene segments was used for corroboration. Considering our data, G genotypes circulating among children in two Croatian regions were G3 (52.4%), G9 (14.7%), G2 (3.7%), G1 (1%), G8 (1%), mixed G genotypes (23.6%) and untypeable G genotype (3.7%). The P genotypes predominantly consisted of P[8] (85.3%), followed by P[4] (2.6%), P[9] (2.1%), P[6] (1%), P[14] (1%), mixed P genotypes (2.1%) and finally untypeable P genotypes (4.7%). We also noted eight different G and P combinations, listed in the order of predominance as follows: G3P[8], G9P[8], G2P[4], G2P[8], G9P[6], G9P[9], G1P[8], G8P[14], alongside various mixed genotypes. Among eight genotype combinations, three may be considered uncommon (G2P[8], G9P[6] and G9P[9]). An animal to human interspecies transmission was evidenced with the circulation of bovine-like G8, G10 and P[14] RVA strains. G10 strains were only detected in mixed genotype group. An additional fact to accentuate is that within the G3 genotype, emerging G3 equine-like RVA strains were predominant with a 97% share over classic G3 strains. This represents an increase from a 48.5% share detected in two previous RVA seasons. The observation of the high detection rate of mixed G genotype in the present research is usually more common in developing countries. To conclude, our results state the importance of monitoring dynamic changes in the prevalence of different RVA genotypes which may serve as a basis for the eventual introduction of RVA vaccines in the national immunization program in Croatia.

Izlaganje na konferenciji MICC 2018 - Medicinske spoznaje u javnom prostoru s tematikom utjecaja medija na antivakcinalni pokret

Izvanbolnička upala pluća jest potencijalno ozbiljna infekcija u djece. Dijagnostika izvanbolničkih upala pluća u djece temelji se na anamnestičkim podatcima i kliničkim simptomima i znacima, potpomognuto dodatnim dijagnostičkim pretragama: laboratorijskim, slikovnim i mikrobiološkim. Etiologija izvanbolničke upale pluća ovisi o brojnim čimbenicima, kao što su sezonstvo, geografski položaj, dob bolesnika i težina bolesti. Liječenje djeteta s izvanbolničkom upalom pluća uključuje primjenu simptomatskih mjera i u većine bolesnika antimikrobnu terapiju. U radu su prikazane kliničke preporuke Hrvatskog društva za pedijatrijsku pulmologiju radi ujednačenja postupaka i kriterija postavljanja dijagnoze, liječenja i prevencije izvanbolničkih upala pluća u djece.

Aim: To depict heterogeneous clinical features of atypical Guillain-Barré syndrome (GBS) variants overlapping between different GBS types and subtypes. Methods: Retrospective analysis of data comprising neurological features, cerebrospinal fl uid (CSF) analysis, ganglioside antibody testing results, electromyography (EMG) fi ndings, brain and spinal magnetic resonance imaging (MRI) in all pediatric patients with GBS treated during a 10-year period at a tertiary center. Results: Twenty-three children were treated for GBS during the study period. Atypical variants were found in fi ve patients and included bifacial and severe pharyngocervicobrachial weakness of descending type, sixth nerve lesion accompanied with lower extremity paresthesias, sensory atactic neuropathy and facial nerve lesion, acute ptosis with mydriasis and incomplete Miller Fisher syndrome, and bilateral facial nerve paresis (one case each). Initial CSF analysis revealed mostly normal proteinlevel in atypical variants. MRI evaluation was normal in all atypical variants except for enhancement of the cervical nerve roots in a patient with pharyngocervicobrachial subtype. EMG performed in the fi rst two weeks showed prolonged distal latency and proximal conduction block in 3/5 patients, in elicitable nerves and axonal loss on upper extremities in a patient with pharyngocervicobrachial subtype, and absent F-waves and neural potentials in 3/5 patients. Slight decrease of motor conduction velocity was present in 2/5 patients in distal nerve segments. Antiganglioside antibodies were positive in 4/5 patients. Conclusion: Clinical manifestations of GBS are very variable, whereas atypical variants/overlaps are not so uncommon. This study supports the proposed hypothesis of continuous spectrum of GBS requiring reconsideration of the existing diagnostic criteria for classic GBS in pediatric population supported by recently proposed (published) diagnostic guidelines., Cilj: Prikazati heterogene kliničke značajke inačica atipičnog Guillain-Barréova sindroma (GBS) te preklapanje između različitih GBS tipova i podtipova. Metode: Retrospektivna analiza podataka koji uključuju neurološke značajke, analizu cerebrospinalne tekućine, rezultate ispitivanja antigangliozidnih protutijela, nalaze elektromiografi je (EMG), magnetsku rezonanciju mozga i kralježnice (MRI), svih pedijatrijskih bolesnika liječenih zbog GBS-a u tercijarnom centru u 10-godišnjem razdoblju. Rezultati: Liječeno je ukupno 23 djece zbog GBS-a. Atipične varijante pronađene su u pet bolesnika i uključivale su bifacijalnu i tešku faringo-cerviko-brahijalnu silaznu slabost, leziju šestog živca popraćenu parestezijama donjih ekstremiteta, senzoričku ataktičnu neuropatiju i leziju facijalnog živca, akutnu ptozu s midrijazom i nepotpunim Miller Fisherovim sindromom te bilateralnu parezu facijalnog živca (sve po jedan slučaj). Inicijalna analiza cerebrospinalne tekućine pokazala je većinom normalnu razinu proteina. Radiološka obrada je bila uredna u svim atipičnim varijantama osim u bolesnika s faringo-cerviko-brahijalnom varijantom gdje je nađen pojačan signal u korijenu živaca cervikalne kralježnice. EMG je prva dva tjedna bolesti pokazao produljenu distalnu latenciju i proksimalni blok u 3/5 bolesnika, gubitak aksona na gornjim ekstremitetima u bolesnika s faringo-cerviko-brahijalnim podtipom te odsutne F-valove i neuronske potencijale u 3/5 bolesnika. Blago smanjenje brzine provođenja u distalnim segmentima živaca bilo je prisutno u 2/5 bolesnika. Antigangliozidna protutijela bila su pozitivna u 4/5 bolesnika. Zaključak: Kliničke manifestacije GBS-a vrlo su varijabilne, a atipične varijante/preklapanja nisu rijetke. Ovo istraživanje podupire predloženu hipotezu o kontinuiranom spektru GBS-a koja zahtijeva preispitivanje postojećih dijagnostičkih kriterija za klasični GBS u dječjoj populaciji te uključenje kriterija za varijante GBS-a koji su nedavno predloženi i objavljeni.

Objectives: Multi-system inflammatory syndrome in children (MIS-C) is a rare complication of SARS-CoV-2 infection defined when there is no other obvious cause of multi-system inflmamation. Bacterial co-infection and secondary bacterial infection are considered critical risk factors for the severity and mortality rates of COVID-19. We present a 10-year-old boy with invasive meningococcal disease (IMD) and SARS-CoV- 2 co-infection. Case Report: Previously healthy 10-year-old boy presented with fever, cough, nausea and headache. On the 4th day of illness, diagnosis of COVID-19 was made by positive RT-PCR for SARS-CoV-2 from nasopharyngeal swab and the boy was discharged home. However, due to the ongoing fever and malaise he was admitted three days later as a possible MIS-C. At admission, he was hypothermic (34.9°C), hypotensive (RR 96/65 mmHg), with positive meningeal signs and petechial-purpuric rash on the extremities, abdomen and gluteus. Upon suspicion of IMD, the ceftriaxone treatment was started. His initial laboratory results included: procalcitonin of >100 mcg/L, CRP of 310 mg/L, WBC count of 19.9 x 109/L with relative neutrophilia of 91%, platelet count of 111 x 109/L, PV of 46%, D-dimers of 34.48 mg/L, NT-pro- BNP of 6099 ng/L and IL-6 of 1691 ng/L. Serology for SARS-CoV-2 was positive (IgG 77.9 IU/ml) and RT-PCR for Neisseria meningitidis from blood was positive for serogroup B. Chest X-ray, echocardiography and brain MSCT were unremarkable. Treatment included ceftriaxone for 10 days with no need for ICU treatment. During hospital stay he developed post-infectious erythema nodosum, but otherwise recovered completely. Conclusion: SARS-CoV-2 predisposition to invasive bacterial infections remains poorly explained due to lack of reported cases. As did in our patient, IMD may trail a preceding infection by around 2 weeks. Surveillance for invasive bacterial infection is important to characterise and to help protect against bacterial co-infection in the context of the ongoing COVID-19 pandemic.

Izvanbolnička upala pluća jest potencijalno ozbiljna infekcija u djece. Dijagnostika izvanbolničkih upala pluća u djece temelji se na anamnestičkim podatcima i kliničkim simptomima i znacima, potpomognuto dodatnim dijagnostičkim pretragama: laboratorijskim, slikovnim i mikrobiološkim. Etiologija izvanbolničke upale pluća ovisi o brojnim čimbenicima, kao što su sezonstvo, geografski položaj, dob bolesnika i težina bolesti. Liječenje djeteta s izvanbolničkom upalom pluća uključuje primjenu simptomatskih mjera i u većine bolesnika antimikrobnu terapiju. U radu su prikazane kliničke preporuke Hrvatskog društva za pedijatrijsku pulmologiju radi ujednačenja postupaka i kriterija postavljanja dijagnoze, liječenja i prevencije izvanbolničkih upala pluća u djece., Community-acquired pneumonia is potentially serious infection in children. The diagnosis can be based on the history and clinical signs and symptoms. Additional diagnostic tests (laboratory, imaging, and microbiology) may be helpful when diagnosis is unclear. The most likely etiology depends on a number of factors such as seasonality, geographic location, the age of child, and the severity of disease. Treatment involves symptomatic measures, and in most children antibiotic therapy. This paper presents the clinical guidelines of the Croatian Society of Pediatric Pulmonology to standardize the procedures for the diagnosis and treatment of children with community-acquired pneumonia.

Cilj panel diskusije je prikazati epidemiološku situaciju prije i nakon uvođenja cjepiva protiv pneumokokne bolesti u nacionalni imunizacijski program.

Introduction Acute hepatitis associated aplastic anemia (AHAAA) is a rare condition in which acute hepatitis is complicated by development of aplastic anemia. It is more common in young males, and presents with pancytopenia 2-3 months after an episode of acute hepatitis. The etiology of hepatitis mostly remains unknown and it is thought that aplastic anemia is caused by immune dysregulation provoked by perhaps infection triggered cytokine production that injures hematopoetic stem cells. Options for treating severe AHAAA are bone marrow transplantation or immunosuppressive therapy (corticosteroids, cyclosporin A, antithymocyte globulin, antilymphocite globulin). Case Report We present a case of a previously healthy 10-year old boy, admitted due to sudden onset of jaundice and nausea. Laboratory results showed highly elevated aminotransferase (AST 2089 U/L, ALT 3228 U/L), conjugated hyperbilirubinemia (total bilirubin 391 umol/L, conjugated 302 umol/L) and initially preserved synthetic liver function. Extensive workup excluded infectious, toxic and metabolic causes. Hypergammaglobulinemia was not present, but anti smooth muscle antibodies were positive in two consecutive testing (1:20). Liver biopsy showed unspecific mixed type acute inflammation. In the next few days he developed liver failure (INR 2.6) so immunosuppressive therapy for autoimmune hepatitis (AIH) was initiated (corticosteroids and azathioprine), together with symptomatic therapy (fresh frozen plasma).Due to potential adverse toxic effect (decreased activity of the enzyme thiopurinmethyltransferase) azathioprin was changed to mycophenolate mofetil. In the following weeks patient’s synthetic liver function completely recovered and aminotransferase and bilirubin levels improved. Six weeks after initial onset of his symptoms patient developed severe thrombocytopenia and leukopenia followed by mild anemia. Bone biopsy showed hypoplastic to aplastic bone marrow. Screening for opportunistic pathogens revealed positive Pneumocystis carinii, and also CMV and VZV reactivation so treatment with trimethoprim-sulfametoxazole and valganciclovir was started together with antifungal therapy for oral candidiasis. He also required repeated platelet transfusions. After two months his bone marrow started showing signs of recovery and he was discharged from hospital. At last follow up, his leukocyte counts was 4.3 x 109/L, hemoglobin 142 g/L and platelet count 57 x 109/L. Discussion Our patient followed the typical presentation of AHAAA described in the scarce literature. The notable aspect of our case is potentially spontaneous recovery of bone marrow aplasia without the need for bone marrow transplantation. Mycophenolate mofetil might be viable therapeutic step in treatment for this type of disease.

To gain insight into the peculiarities of clinical presentation and time of diagnosis in the population of patients with combined immunodeficiency treated in our center. We retrospectively analyzed 22 patients with combined immunodeficiency treated in our center in the past 25 years. Clinical findings were collected, including age of presentation and age of diagnosis, genetic defects, consanguinity, family history, number and site of infections, presence of diarrhea, failure to thrive, skin rash, vaccine complications and type of treatment. Immunological findings included WBC with absolute counts, lymphocyte subsets and immunoglobulin levels. Patients (n=22) with severe combined immunodeficiency (n=17) and other combined immunodeficiencies (n=5) were included in the study. Sixteen patients (77%) were male. The most common clinical manifestations were opportunistic infections. Most of the patients were diagnosed after 3 months of age. Prenatal diagnosis was performed in one patient (4.5%), two patients (9%) did not undergo genetic analysis. The spectrum of genetic defect in our cohort show wide heterogeneity. Seventeen patients (77%) underwent HSCT, one patient (4.5%) is on enzyme replacement therapy. None of our patients have undergone gene therapy. Twelve patients (54.5%) died, of which four (18%) died before HSCT. Combined immunodeficiencies are fatal without definitive therapy (HSCT or gene therapy). The outcomes are better if infants are diagnosed and treated in pre-symptomatic stage, preferably before 3.5 months of age. More and more countries are implementing newborn screening for primary immunodeficiency diseases in their health care systems to diagnose patients as early as possible and to prevent infections. For the same reason, we advocate for expanding our national neonatal screening program