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Analizirana je seroprevalencija, genotipovi i rizični faktori za HCV infekciju u populacijama rizičnog spolnog ponašanja: osoba koje često mijenjaju spolne partnere (PROM), muškaraca koji imaju seksualne odnose s muškarcima (MSM), prodavatelja/ica seksualnih usluga i njihovih korisnika (PSU) te osoba koje u anamnezi imaju spolno prenosive bolesti (SPB). Kontrolnu su skupinu sačinjavale trudnice. U istraživanje je bio uključen 821 ispitanik: od toga 716 (87,2%) muškaraca i 105 (12,8%) žena prosječne životne dobi od 35,8+11,5 godina. Medijan dobi iznosio je 32 godine (raspon 18-77 godina). Skupini s jednom vrstom rizičnog spolnog ponašanja pripadalo je 71,5% ispitanika, dok je 25,2% imalo dvije vrste, 3,2% tri vrste te 0,1% ispitanika četiri vrste rizičnog spolnog ponašanja. Medijan broja spolnih partnera u posljednjih 12 mjeseci iznosio je 2 (raspon 0-100). SPB u anamnezi imalo je 25,3% ispitanika slijedećom učestalošću: HBV 12,8%, gonoreju 4,6%, C. trachomatis 2,7%, HPV 2,7%, genitalni/analni herpes 1,3% te sifilis 1%. Ukupna seroprevalencija HCV infekcije u ispitanika rizičnog spolnog ponašanja iznosila je 4,6% (95% CI=3,2-6,1%): 6,3% (95% CI=3,9-8,8%) u skupini PROM ispitanika; 2,9% (95% CI=0,6-5,2%) u skupini MSM; 4,0% (95% CI=1,7-6,4%) u skupini PSU i korisnika PSU; 8,5% (95% CI=4,7-12,4%) u skupini ispitanika sa SPB te 0,5% (95% CI=0.0-1,5%) u kontrolnoj skupini. Dokazana je statistički značajna razlika HCV seroprevalencije u ispitanika rizičnog spolnog ponašanja u odnosu na kontrolnu skupinu u svim ispitivanim skupinama [PROM: OR=13,49, 95% CI (OR)=1,81-100,4; PSU: OR=8,39, 95% CI (OR)=1,07-65,5; SPB: OR=18,59, 95% CI (OR)=2,4-141,1] osim MSM [OR=6,00, 95% CI (OR)=0,72-50,2]. S obzirom na spol, seropozitivno je bilo 4,9% (95% CI=0,0-6,0) muškaraca te 2,9% (95% CI=3,3-6,0) žena (p=0,350). HCV seroprevalencija nije se značajno razlikovala između dobnih skupina ispitanika (p=0,230). Najviše seropozitivnih ispitanika bilo je u dobi od 20-29 godina (5,8%; 95% CI=3,2-8,4%) te 30-39 godina (5,7%; 95% CI=2,8-8,6%). Bračni/intimni status (p=0,681) te stručna sprema (p=0,978) ispitanika nisu bili povezani s HCV seropozitivitetom. Od preboljelih SPB, značajno viša HCV seroprevalencija dokazana je u ispitanika s preboljelom HBV infekcijom (p=0,002) te gonorejom (p=0,01). HCV-RNA dokazana je u 73,1% seropozitivnih te 2,1% seronegativnih ispitanika (novozaraženi ispitanici u periodu "imunološkog prozora"). Najviše ispitanika bilo je inficirano genotipom 1 (55,6%), genotipom 3 38,9% te genotipom 4 5,6% ispitanika. Najčešći dokazani podtipovi bili su 1a (38,9%) te 3a (38,9%). Ovisno o rizičnim skupinama, u skupini PROM ispitanika dokazana su sva tri genotipa (podtipovi 1a i 3a), u MSM genotipovi 3a i 4, u skupini PSU i korisnika PSU genotipovi 1a i 3a te u ispitanika sa SPB genotipovi 1 (podtipovi 1a i 1b) i 3a., We analyzed the seroprevalence, genotypes and risk factors for HCV infection in groups with high-risk sexual behavior: persons with multiple sexual partners, men who have sex with men (MSM), commercial sex workers and their clients (CSW) and persons with sexually transmitted diseases (STDs). Control group consisted of pregnant females. A total of 821 patients were enrolled in the study: 716 (87.2%) males and 105 (12.8%) females. The mean age was 35.8+11.5 years. Median age was 32 years (range 18-77 years). Seventy-one point five percent of participants reported one risk sexual behavior, 25.2% reported two, 3.2% reported three and 0.1% of participants reported four risk sexual behaviors. Median number of sexual partner in the preceding year was 2 (range 0-100). A prior history of STDs had 25.3% participants. Hepatitis B virus (HBV) infection was the most frequently reported (12.8%) followed by gonorrhea (4.6%), C. trachomatis (2.7%) and HPV infection (2.7%), genital/anal herpes (1.6%) and syphilis (1.0%). The overall HCV seroprevalence was 4.6%: 6.3% in persons with multiple sexual partners, 2.9% in MSM, 4.0% in CSW and their clients and 8.5% in persons with STDs compared to 0.5% in control group. HCV seroprevalence was significantly higher than in controls in all risk groups except MSM [persons with multiple sexual partners: OR=13.49; 95% CI (OR)=1.81-100.4; CSW and their clients: OR=8.39; 95% CI (OR)=1.07-65.5; persons with STDs: OR=18.59; 95% CI (OR)=2.4-141.1; MSM OR=6.00; 95% CI (OR)=0.72-50.2]. The difference of HCV seroprevalence between age groups was not statistically significant (p=0.230). The highest seroprevalence was reported in the 20-29 age group (5.8%) and in the 30-39 age group (5.7%). Neither marital status (p=0.681) nor level of education (p=0.978) were associated with HCV seropositivity. The prior HBV infection (p=0.002) and gonorrhea (p=0.011) were the only risk factors significantly associated with anti-HCV prevalence. No other factors reflecting risk sexual behavior such as sexual orientation (p=0.355), the number of sexual partners (p=0.318) and the number of risk factors (p=0.079) were associated with HCV seroprevalence. HCV-RNA was detected in 73.1% seropositive participants. Three of the seronegative cases (2.1%) were also found to be HCV-RNA positive (“window period”). The majority of participants were infected with genotype 1 (55.6%), 38.9% with genotype 3 and 5.6% with genotype 4. The most prevalent subtypes were 1a (38.9%) and 3a (38.9%). According to the risk groups, in persons with multiple sexual partners all three genotypes (subtypes 1a and 3a) were detected. MSM were infected with genotypes 3a and 4, CSW and their clients with genotypes 1a and 3a and persons with STDs were infected with genotypes 1 (subtypes 1a and 1b) and 3a.

Background: Parvovirus B19 (B19V) is associated with a wide range of clinical manifestations. The major presentation is erythema infectiosum. However, a persistent infection may cause pure red cell aplasia and chronic anemia in immunocompromized patients. The B19V seroprevalence varies with age and geographical location. Aim: To determine the B19V serological status and DNAemia in kidney, liver, and pancreas transplant candidates. Methods: Patients who underwent kidney, liver, or simultaneous kidney and pancreas/liver transplantation between January 2021 and May 2022 were included in the study. The serum samples were collected before transplantation. For detection of B19V DNA, a LightMix Kit B19V EC (TIB MOLBIOL, Berlin, Germany) was used. B19V IgM and IgG antibodies were detected using a commercial ELISA test (Euroimmun, Lübeck, Germany). Results: One hundred and thirty-one transplant candidates were included in the study, 71.0% male, with an average age of 53.27 years ± 12.71 years. There were 68.7% liver, 27.5% kidney, 3.0% simul taneous pancreas/kidney transplant (SPKT), and 0.8% simultaneous liver/kidney transplant recipients. No patients had detectable B19V DNA. B19V IgG seroprevalence was 77.1%. No acute or recent infections were detected (IgM antibodies). There was no difference in the mean age of seronegative and seropositive patients (51.8 years ± 12.9 years vs 53.7 years ± 12.7 years, t = -0.603; P = 0.548). Although seropositivity was lower in patients aged less than 30 years (66.6%) compared to the patients aged 30-59 years and > 60 years (80.4% and 78.1%, respectively), this difference was not significant. In addition, there was no difference in seropositivity between male and female transplant candidates, 76.3% and 78.9% (χ 2 = 0.104; P = 0.748). The seroprevalence did not differ among organ recipients, with 77.8%, 80.6%, and 50.0% for liver, kidney, and SPKT, respectively, (χ 2 = 5.297; P = 0.151). No significant difference was found in the seroprevalence in kidney transplant patients according to dialysis modality. Seroprevalence was 71.1% in hemodialysis patients, and 100% in peritoneal dialysis patients (χ 2 = 0.799; P = 0.372). Conclusion: The B19V seroprevalence is expectedly high among kidney, liver, and pancreas transplant candidates, but there are still 22.9% of seronegative individuals who remain at risk for primary disease and severe manifestations. Further research should elucidate the necessity of B19V screening in peri-transplant management.

BACKGROUND: Hepatitis E virus (HEV) is an emerging virus of global health concern. The seroprevalence rates differ greatly according to geographic region and population group. AIM: To analyze the seroprevalence of HEV in exposed (animal-related professions) and nonexposed populations, as well as solid organ and hematopoietic stem cell transplant patients. METHODS: Forestry workers (n = 93), hunters (n = 74), and veterinarians (n = 151) represented the exposed population. The general population (n = 126) and pregnant women (n = 118) constituted the control group. Transplant patients included liver transplant recipients (LTRs) (n = 83), kidney transplant recipients (KTRs) (n = 43), and hematopoietic stem cell transplant recipients (HSCRs) (n = 39). HEV immunoglobulin G antibodies were detected using the enzyme- linked immunosorbent assay and confirmed by the immunoblot test. RESULTS: The HEV seroprevalence significantly differed between groups: Veterinarians 15.2%, hunters 14.9%, forestry workers 6.5%, general population 7.1%, and pregnant women 1.7%. In transplant patients, the seropositivity was highest in LTRs (19.3%), while in KTRs and HSCRs, the seroprevalence was similar to the general population (6.9% and 5.1%, respectively). A significant increase in seropositivity with age was observed from 2.9% in individuals less than 30 years to 23.5% in those older than 60 years. Sociodemographic characteristics (sex, educational level, area of residence, and number of household members), eating habits (game meat, offal, and pork products consumption), and environmental and housing conditions (drinking water supply, type of water drainage/sewer, waste disposal, domestic animals) were not associated with HEV seropositivity. However, individuals who reported a pet ownership were more often seropositive compared to those who did not have pet animals (12.5% vs 7.0%). CONCLUSION: The results of this study showed that individuals in professional contact with animals and LTRs are at higher risk for HEV infection. In addition, age is a significant risk factor for HEV seropositivity.

Background: Tahyna orthobunyavirus (TAHV) is a neglected mosquito-borne arbovirus. The vertebrate hosts for TAHV are hares, rabbits, hedgehogs and rodents, while Aedes vexans mosquitoes are the main vectors. Although TAHV is widespread throughout continental Europe, due to the lack of commercially available assays, human clinical cases are rarely reported. Aim: The aim of this study was to analyze the prevalence of TAHV in humans, animals and mosquitoes ('One Health') in Croatia. Methods: The study included: patients with neuroinvasive disease (NID) of unsolved etiology from continental and coastal Croatian regions (2017-2021 ; N=218) ; asymptomatic individuals from Zagreb and its surroundings (2020-2022 ; N=218) ; animals from Zagreb and its surroundings (2020- 2022 ; horses N=98, dogs N=70, cats N=24) and Aedes vexans mosquitoes collected in the Zagreb area (2020-2022 ; N=4456). TAHV neutralizing (NT) antibodies were detected using a virus neutralization test. Mosquito pools were tested for the presence of TAHV RNA using RT-PCR. Results: TAHV NT antibodies were detected in 20 (10.1%) patients with NID and 8 (3.7%) asymptomatic individuals. In two patients presented with meningitis, NT antibodies were detected in the CSF suggesting recent infection. While in patients with NID seroprevalence increased significantly with age from 1.8% to 24.4%, in asymptomatic individuals there was no difference in the seropositivity between age groups (2.5-7.7%). The majority of seropositive humans were residents of floodplains along the rivers in continental Croatia. In addition, a total of 29 (29.6%) horses, and 11 (11.7%) pet animals were found to be TAHV seropositive. No one mosquito pool tested positive for TAHV RNA. Conclusion: The presented results showed the presence of TAHV in Croatia. Further studies on a large sample of humans and animals are needed to confirm the prevalence of this neglected virus in the Croatian population.

Aim: Neuroinvasive arboviruses such as tick-borne encephalitis virus (TBEV), West Nile virus (WNV) and Usutu virus (USUV) are widely distributed in continental Croatia. We analyzed the prevalence of neuroinvasive arboviral infections detected in humans and animals ('One Health') in the 2022 transmission season. Methods: Diagnosis in humans was confirmed by detection of viral RNA in the cerebrospinal fluid (CSF) or urine (WNV) and/or specific IgM antibodies in the CSF (TBEV, WNV). In horses, WNV IgM/IgG antibodies were detected in serum samples. In a bird, USUV RNA was detected in brain tissue by RT-PCR. Results: During the 2022 transmission season (up to 31 August), TBEV infection was confirmed in 15 patients (12 males and 3 females ; median age 58, IQR=23-69 years). Clinical presentations were 'febrile headache' (N=2), meningitis (N=12) and meningoencephalitis (N=1). WNV was detected in five patients presented with meningitis (N=2), encephalitis (N=2) and myelitis (N=1). The median patients' age was 54 (IQR=28- 65) years. TBEV infections occurred from April to August in Primorje-Gorski Kotar (cluster of 8 cases), Osijek-Baranja, Varaždin, Virovitica- Podravina and Brod-Posavina County. WNV was detected in August in Osijek-Baranja and Zagreb County. Two TBEV infections were imported from Bavaria (Germany) and Poland. In addition, acute asymptomatic WNV infections (IgM positive) was recorded in four horses from Zagreb, Zagreb County and Brod- Posavina County. For the first time in Croatia, clinically manifest WNV neuroinvasive infection was confirmed in one horse from Osijek- Baranja County. USUV was confirmed in a dead blackbird (Turdus merula) from Zagreb presented with neurologic symptoms. Conclusion: TBEV infections were continuously detected in continental Croatian regions. Human WNV infections re-emerged in 2022 after a three- year absence. For the first time in Croatia, clinically manifest neuroinvasive infection was confirmed in a horse (WNV) and a blackbird (USUV).

Posljednjih se godina na području Europe sve češće bilježi pojava arbovirusnih infekcija. Značajno mjesto među njima zauzimaju West Nile (WNV) i Usutu virusna (USUV) infekcija, čiji vektori su komarci. Ovaj rad prikazuje rezultate testiranja komaraca na prisutnost flavivirusa WNV i USUV, koji su prikupljeni na području Grada Zagreba u razdoblju od 2015. do 2019. godine. Komarci su uzorkovani CDC klopkama, BG Sentinel klopkama i aspiratorom. Za dokazivanje virusa ukupno su pripremljene 15.863 jedinke komaraca prikupljene tijekom 466 uzorkovanja. Determinirano je 10 vrsta komaraca. Jedinke su razvrstane u 700 skupnih uzoraka koji su testirani na prisutnost WNV i USUV RNA metodom RT- qPCR u stvarnom vremenu. RNA USUV je dokazana u tri skupna uzorka: jednom skupnom uzorku jedinki Ae. albopictus iz 2016. godine i dva skupna uzorka jedinki Cx. pipiens kompleks (2018. i 2019. godina). RNA WNV nije dokazana niti u jednom testiranom skupnom uzorku komaraca. Filogenetskom analizom pozitivnih skupnih uzoraka iz 2018. i 2019. godine dokazana je USUV europska linija 2. USUV pozitivni skupni uzorak komaraca Cx. pipiens kompleks iz 2018. godine dodatno je analiziran NGS metodom pri čemu su dokazane virusne sekvence iz različitih porodica virusa od kojih većina pripada virusima koji inficiraju komarce, a od virusa koji inficiraju sisavce i ptice dokazan je USUV. Dokaz RNA USUV u skupnim uzorcima komaraca u Zagrebu tijekom godina istraživanja pokazuje da je virus kontinuirano prisutan na istraživanom području te da je područje sjeverozapadne Hrvatske postalo endemsko područje za USUV. Dokazana RNA USUV u skupnom uzorku vrste Ae. albopictus 2016. godine prvi je dokaz flavivirusa u komarcima u Hrvatskoj.

Aim To assess the humoral immunity to COVID-19 in nursing home residents six months after vaccination. Methods This seroepidemiological research enrolled 118 residents of one nursing home in Zagreb. All participants received two doses of BioNTech/Pfizer COVID-19 and had no previously detected SARS-CoV-2 infection. The samples were tested for the presence of neutralizing antibodies using a virus neutralization test. A SARS-CoV-2 strain isolated in Vero E6 cells from a Croatian COVID-19 patient was used as a stock virus. Neutralizing antibody titer was defined as the reciprocal of the highest serum dilution that showed at least 50% neutralization. Neutralizing antibody titer ≥8 was considered positive. Results Sixty-four (54%) participants had a positive neutralizing antibody titer, 27 (23%) had a low positive titer (titer 8), and 27 (23%) had a negative titer. Women had a significantly higher median titer than men (16 [interquartile range, IQR 24] vs 8 [IQR 12], Mann-Whitney U = 1033, P = 0.003). Age was negatively but not significantly correlated with neutralizing antibody titer (Spearman’s rho -0.132, P = 0.155). Conclusion Almost half of the participants (46%) had a negative or low positive titer six months after having been fully vaccinated. This study suggests that humoral immunity among nursing home residents considerably wanes six months after BioNTech/Pfizer COVID-19 vaccination. Our results could contribute to the discussion about the need for a booster dose.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a novel, highly transmissible virus that has led to the ongoing coronavirus disease 2019 (COVID-19) global pandemic. It emerged in late 2019 in Wuhan, China and since then virus has been rapidly spreading worldwide. The World Health Organization (WHO) officially declared it a pandemic in March 2020. Clinical features of COVID-19 vary from asymptomatic or mild common cold-like symptoms to more severe multi-organ manifestations. Emerging evidence suggests that subacute thyroiditis (SAT) is one of the rare and potentially neglected manifestations of COVID-19. SAT is a self-limiting inflammatory thyroid disease thought to be linked to a viral infection or post-viral inflammatory reaction. According to recent reports, SARS-CoV-2, amongst several other familiar viruses, may also precipitate development of SAT. It is often characterized by anterior cervical pain, transient abnormalities in thyroid function and systemic symptoms such as fever, fatigue, myalgia and anorexia. Diagnosis is usually based on clinical presentation, physical examination, thyroid hormones analysis and imaging. Until 28th November 2020, there are 13 published studies of SAT in 28 patients with COVID-19. Median age at the time of diagnosis was 41 years and 75% of patients were females. The median between COVID-19 diagnosis and the onset of SAT symptoms is reported to be 29 days. Common clinical symptoms were palpitations (80%), fever (65%), fatigue (50%) and anterior cervical pain radiating to jaw (50%). The median follow-up was 40 days (range 10-70 days) and 71% of patients regained normal thyroid function within that period. However, 11% of patients presented with subclinical hypothyroidism, 11% with hypothyroidism and 7% of them with hyperthyroidism. Although limited number of SAT cases after SARS-CoV-2 infection are reported, physicians should consider SAT as a differential diagnosis in COVID-19 patients. The routine assessment of thyroid function in patients with clinical suspicion of SAT or with the suspicion of any other thyroid dysfunction is advised.

Objectives: Literature data on the immunopathogenesis of neuroinvasive arboviral infections, particularly in the context of complex cytokine immune responses, are limited. The aim of this study was to compare cytokine expression patterns in the cerebrospinal fluid (CSF) and serum in human neuroinvasive arboviral infections caused by tick-borne encephalitis virus (TBEV), West Nile virus (WNV) and Toscana virus (TOSV). Methods: Antiviral cytokine response in the serum and CSF was analysed by using a multiplex bead- based assay for simultaneous quantification of 13 human cytokines (LEGENDplex Human Th cytokine panel, BioLegend, USA) on FACS Canto II flow cytometer (Beckton Dickinson, USA). The panel included cytokines associated with innate and early pro-inflammatory immune responses (TNF-α, IL-6), Th1 type immune response (IL-2, IFN-γ), Th2 immune response (IL-4, IL-5, IL-9 and IL-13), Th17 immune response (IL-17A, IL-17F, IL-21 and IL-22) and the key anti-inflammatory cytokine IL-10. Results: Analysis of cytokine expression pattern in the CSF and serum of patients with neuroinvasive infection with TBEV (32 patients), WNV (68 patients) and TOSV (3 patients) revealed partially similar patterns. Significantly higher concentrations of IL-6 in the CSF vs. serum were found in both TBEV (median 780.88 pg/mL, IQR 130.81-3, 077.66 vs. median 23.25 pg/mL, IQR 7.79- 27.09, respectively ; p

U lipnju 2019. godine, Služba za epidemiologiju zaraznih bolesti Hrvatskog zavoda za javno zdravstvo je obaviještena o grupiranju oboljelih od krpeljnog meningoencefalitisa (KME) na području Gorskog kotara. Od šest oboljelih pacijenata, pet ih je konzumiralo nepasterizirano (sirovo) kozje mlijeko s obiteljske farme u razdoblju od dva tjedna prije pojave simptoma, dok je jedna oboljela osoba u anamnezi navela nedavni ugriz krpelja. Kako bi procijenili rizične faktore za infekciju proveli smo slučaj-kontrola studiju u kojoj je izabrano šest kontrolnih ispitanika među zdravim osobama, članovima obitelji ili stanovnicima sela u kojem je zabilježeno grupiranje oboljelih. Provedeno je epidemiološko anketiranje i virološko testiranje oboljelih osoba i kontrolnih ispitanika. Izračunali smo omjer šansi za pojedine rizične izloženosti. Uzorci seruma koza, konja i psa s obiteljske farme testirani su na prisutnost neutralizacijskih protutijela na virus KME (KMEV). Uzorci kozjeg mlijeka i 65 krpelja vrste Ixodes ricinus prikupljenih sa životinja testirani su na prisutnost KMEV RNA metodom RT-PCR. Oboljele su 4 osobe ženskog i 2 osobe muškog spola, a medijan dobi oboljelih osoba bio je 41 godinu (10-85 godina). Oboljele osobe imale su 25 (95% interval pouzdanosti 0.8-1410.2, p = 0.021) puta više šanse da konzumiraju sirovo kozje mlijeko u posljednja dva tjedna u odnosu na zdrave kontrole. Nitko od oboljelih niti kontrola nije bio cijepljen protiv KME. Virus KME nije dokazan u uzorcima mlijeka niti krpeljima, dok je serološko testiranje životinja s farme ukazalo na izloženost virusu: šest koza iz stada imale su neutralizacijska protutijela na KMEV. Rezultati istraživanja pokazali su da je izvor epidemije bila konzumacija nepasteriziranog kozjeg mlijeka s obiteljske farme. Nakon preporuke za prestanak konzumacije proizvoda od nepasteriziranog mlijeka, nije bilo novooboljelih osoba. Ova epidemija ukazuje na potrebu provođenja edukacije među proizvođačima i potrošačima o rizicima koje donosi konzumacija nepasteriziranih mliječnih proizvoda. Preporuča se dodatno testiranje ljudi, domaćih životinja i krpelja radi boljeg razumijevanja epidemiologije KME u Gorskom kotaru, te za ciljano donošenje preporuka o cijepljenju protiv KME.

Više od godinu dana od početka COVID-19 (Coronavirus disease 2019) pandemije sve je više dokaza da je učestalost infekcija SARS-CoV-2 (Severe acute respiratory syndrome coronavirus 2) u kućnih ljubimaca daleko viša nego se prvotno smatralo. Ovo istraživanje provedeno je od 1. srpnja do 31. prosinca 2020. godine te je obuhvaćalo 78 pasa koji žive u kućanstvima s potvrđenim slučajem infekcije SARS-CoV-2 u ljudi. Pretraženo je i 1069 ostatnih uzoraka seruma pasa zaprimljenih u istom razdoblju na klinike Veterinarskog fakulteta Sveučilišta u Zagrebu, a koji su s područja grada Zagreba. Prikupljenih uzorci seruma pretraženi su imunoenzimnim testom (ELISA) koji je za potrebe ovog istraživanja razvijen i validiran. Svi ELISA pozitivni uzorci pretraženi su i mikrotitracijskim neutralizacijskim testom (MNT). Od ukupnog broja pretraženih seruma pasa iz COVID-19 pozitivnih kućanstava, ELISA testom su IgG protutijela dokazana u 43, 59% uzoraka, dok su neutralizirajuća protutijela dokazana u 25, 64% seruma. Vjerojatnost infekcije pasa koji borave u domaćinstvu s inficiranom osobom se nije razlikovala od one ranije utvrđene u ljudi. U pasa zaprimljenih na klinike Veterinarskog fakulteta, 14, 69% uzoraka je bilo ELISA pozitivno. Ovo je gotovo dvostruko viša seroprevalencija nego što je bila zabilježena krajem prvog vala pandemije, kada je iznosila 7, 45%. Značajno je napomenuti da je na kraju istraživanja, u prosincu 2020. godine, gotovo svaki peti uzorak seruma pasa koji su zaprimljeni na klinike Veterinarskog fakulteta bio ELISA pozitivan. Razmjer proširenosti infekcije u populaciji pasa najbolje pokazuje usporedba sa seroprevalencijom SARS-CoV-2 infekcije u ljudi. Tijekom prosinca 2020. godine prikupljeni su ostatni uzorci seruma ljudi koji žive na području grada Zagreba. Serološkom pretragom provedenom u Hrvatskom zavodu za javno zdravstvo, protutijela za SARS-CoV-2 su dokazana u 20, 52% uzoraka. U istom mjesecu seroprevalencija u pasa je bila 18, 56% te nije postojala statistički značajna razlika. Ovo istraživanje je pokazalo da je infekcija SARS-CoV-2 učestala u populaciji pasa. Pokusne infekcije su dokazale značajno veću prijemljivost mačaka na infekciju. U prirodnim uvjetima, vjerojatno zbog razlike u načinu držanja, psi budu podjednako često ili čak češće inficirani. Sa stanovišta javnog zdravstva, ovajje podatak izrazito značajan. Visoki postotak infekcija u pasa, uz činjenicu da je COVID-19 u određenog broja ljudi asimptomatska infekcija, otvara mogućnost korištenja pasa kao sentinel životinja što je potrebno dodatno istražiti. Po drugoj strani, za sada nema indicija da psi imaju značajnu ulogu u epidemiologiji COVID-19 u ljudskoj populaciji, međutim zbog nastanka novih varijanti virusa te smanjenom prijenosu SARS-CoV-2 s čovjeka na čovjeka, ulogu pasa, i domaćih životinja uopće, u epidemiologiji SARS-CoV-2 infekcije je potrebno nastaviti pratiti.

Pojava flavivirusnih infekcija tijekom posljednja dva desetljeća sve se češće bilježi na području Europe. Među infekcijama značajno mjesto zauzimaju West Nile (WNV) i Usutu virusna (USUV) infekcija, čiji vektori su komarci. Na području sjeverozapadne Hrvatske (Grad Zagreb i Međimurska županija) u razdoblju od 2015. do 2020. godine odrasle jedinke komaraca su uzorkovane radi testiranja na prisutnost WNV i USUV virusa. Komarci su uzorkovani CDC klopkama, BG Sentinel klopkama i aspiratorom. Ukupno je obavljeno 618 uzorkovanja, 613 u Zagrebu (2015.-2020. godine) i 5 u Međimurskoj županiji (2017. godina). Za dokazivanje virusa ukupno su pripremljene 20.363 jedinke komaraca, od čega s područja Zagreba 20.291, a s područja Međimurske županije 72 jedinke. Određeno je 11 vrsta komaraca. Jedinke su razvrstane u 899 skupnih uzoraka komaraca koji su testirani na prisutnost WNV i USUV RNA metodom RT-qPCR u stvarnom vremenu. RNA USUV je dokazana u četiri skupna uzorka komaraca: jednom skupnom uzorku jedinki Ae. albopictus iz 2016. godine (Zagreb), dva skupna uzorka jedinki Cx. pipiens kompleks iz Zagreba (2018. i 2019. godina) i jednom skupnom uzorku Cx. pipiens kompleks iz Preloga (Međimurje). RNA WNV nije dokazana niti u jednom testiranom skupnom uzorku komaraca. Filogenetskom analizom pozitivnih skupnih uzoraka iz 2018. i 2019. godine dokazana je USUV europska linija 2. USUV pozitivni skupni uzorak komaraca Cx. pipiens kompleks iz 2018. godine (uzorkovan na području ŠRC Jarun) dodatno je analiziran NGS metodom pri čemu su dokazane virusne sekvence iz različitih porodica virusa od kojih većina pripada virusima koji inficiraju komarce, a od virusa koji inficiraju sisavce i ptice dokazan je USUV. Dokaz RNA USUV u skupnim uzorcima komaraca uzastopno tijekom četiri godine na području sjeverozapadne Hrvatske pokazuje da je virus kontinuirano prisutan na istraživanom području te da je navedeno područje postalo endemsko područje za USUV.

SARS-CoV-2 kao RNA virus s velikim genomom podložan je učestalim mutacijama tijekom replikacije. Između beznačajnih i pogubnih mutacija nalaze se i one koje virusu donose selektivnu prednost. Krajem 2020. i početkom 2021. godine pojavile su se varijante koje se čine zaraznije nego postojeći sojevi, izbjegavaju imunološki odgovor i izazivaju težu kliničku sliku. Tri varijante koje su na određenim zemljopisnim područjima brzo postale dominantne te nazvane varijantama od značaja (VOC ; eng. Variant of Concern) su: B.1.1.7, B.1.351 i P.1 Do današnjeg dana ECDC prati još dvije VOC: B.1.1.7+E484K i B.1.617.2 U svrhu praćenja epidemiološke situacije, sukladno preporukama ECDC provedeno je sekvenciranje cijelog genoma (WGS, eng. Whole Genome Sequencing) na 5-10% SARS- CoV-2 pozitivnih uzoraka u Hrvatskoj. WGS se provodi na tjednoj dinamici od 09.02.2021. godine. Dobivene sekvence se učitavaju u GISAID bazu podataka koja izolate kategorizira u filogenetske grane ovisno o jednostrukim polimorfizmima prisutnima u genomu. Do kraja svibnja 2021. godine poslana su 5734 uzorka, od čega je uspješno sekvencirano 4745. Između prva dva vala epidemije dominirali su virusni sojevi G, GR i GV grana, a pojačana prisutnost sojeva iz GRY grane označila je početak trećeg vala epidemije u Hrvatskoj. Dobiveni rezultati sekvenciranja pokazuju značajan porast u udjelu prisutnosti VOC B.1.1.7 od 21.23% u prvom tjednu WGS ispitivanja, do 98, 4% u zadnjem tjednu svibnja 2021. godine. S druge strane, sljedeća VOC B.1.351 GH grane dokazana je sporadično, u 19 uzoraka koji su većinom povezani s putovanjima izvan Hrvatske te njihovim bliskim kontaktima. Pri kraju trećeg vala pojavila se varijanta B.1.617.2, koja je dokazana u dva uzorka.

Koronavirusi (CoV) su velika obitelj RNA virusa koja uzrokuje bolesti dišnog sustava, a one se klinički mogu manifestirati u rasponu od obične prehlade do znatno težih oblika bolesti poput pneumonije praćene akutnim respiratornim sindromom (SARS ; eng. Severe Acute Respiratory Syndrom). COVID-19 je bolest koju uzrokuje novi SARS-CoV-2 koji se lako širi između ljudi, neovisno o prisutnosti simptoma u zaražene osobe. Prvi slučajevi ove bolesti zabilježeni su u Kini u provinciji Hubei sredinom prosinca 2019. godine. Prvi slučaj u Hrvatskoj zabilježen je 25. veljače 2020. godine. Restriktivne epidemiološke mjere uvedene su u ožujku 2020. godine kako bi se spriječilo i kontroliralo širenje bolesti. Postupno popuštanje mjera započelo je u svibnju 2020. godine što je dovelo do postupnog rasta broja zaraženih tijekom ljeta te značajno ubrzanog rasta novozaraženih koje je započelo u rujnu 2020. godine. Do sada je u Hrvatskoj laboratorijski potvrđen 356 181 slučaj zaraze uzrokovane SARS- CoV-2, a 8026 osoba je preminulo. Kako bismo procijenili stvarnu proširenost COVID-19 analizirali smo seroprevalenciju i prisutnost neutralizacijskih protutijela (NT) u uzorcima seruma opće populacije u Hrvatskoj nakon prvog (veljača - srpanj 2020. godine) odnosno drugog (kolovoz 2020. - veljača 2021. godine) pandemijskog vala. Početno serološko testiranje provedeno je komercijalnim imunoenzimskim (ELISA) testom (Vircell, Microbiologists, Granada, Španjolska), a za reaktivne uzorke kao potvrdni test korišten je test neutralizacije virusa (VNT ; eng. virus neutralization test). Zabilježena je statistički značajna razlika u rezultatima ukupne seroprevalencije između prvog (ELISA 2, 2% ; VNT 0, 2%) i drugog pandemijskog vala (ELISA 25, 1% ; VNT 18, 7%). Seropozitivne osobe su zabilježene u svim dobnim skupinama uz statistički značajnu razliku po dobi. Najniža prevalencija NT protutijela nađena je djece (

Coronavirus disease 2019 (COVID-19) is an emerging infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The usual presentation of the disease is a common cold-like illness but it can present with more severe and sometimes fatal manifestations. Immunocompromised patients such as those with common variable immunodeficiency (CVID) also are among the infected population. A limited number of reports have been published concerning CVID patients with COVID-19. The main reported symptoms were fever, cough, dyspnea and fatigue while the median duration of illness was 19 (interquartile range 14-26.5) days. Total recovery rate was 88.4%. It is still unknown whether primary immunodeficiency interacts as a predisposing or protective factor against the severe forms of COVID-19. Substitute immunoglobulin (IG) therapy is the only treatment option for CVID. Some reports suggest that early administration of intravenous IGs or convalescent plasma infusion may positively influence the outcome of COVID-19 in these patients.

Posljednjih nekoliko desetljeća, kontinuirano se bilježi porast broja autohtonih i importiranih emergentnih arbovirusnih infekcija. Arbovirusi predstavljaju veliku javnozdravstvenu prijetnju u endemskim područjima uz tendenciju zemljopisnog širenja. Putnici nemaju samo značajan rizik od zaraze ovim virusima, već doprinose i njihovom širenju u ne-endemska područja. Od ožujka 2017. do rujna 2020. godine u Nacionalnom referentnom laboratoriju za arboviruse, Hrvatskog zavoda za javno zdravstvo testirano je ukupno 149 povratnika iz endemskih područja na prisutnost Zika (ZIKV), chikungunya (CHIKV) i dengue (DENV) RNA te IgM i IgG protutijela. Serološka dijagnostika učinjena je pomoću komercijalnih imunoenzimnih (ELISA) i indirektnih imunofluorescentnih testova (IFA). Početno reaktivni rezultati dodatno su testirani immunoblot testom (LIA) za istodobnu detekciju navedenih arbovirusa. Molekularna dijagnostika učinjena je metodom RT- PCR u stvarnom vremenu. Pozitivni uzorci dodatno su testirani klasičnom RT-PCR metodom te je učinjeno sekvenciranje. Udio muškaraca u testiranoj skupini iznosio je 84 (56, 3%), a udio žena 65 (43, 6%). U skupini žena, 46 (70, 8%) je bilo u generativnoj dobi, od kojih su 9 (19, 5%) bile trudnice. Najčešća mjesta potencijalne izloženosti bile su Srednja i Južna Amerika (27, 5%), Azija (20, 8%), Europa (10%) i Afrika (6, 7%). Većina testiranih putnika razvila je kliničke simptome (60, 4%), dok je njih 39, 6% bilo asimptomatsko. Od kliničkih simptoma najčešće su zabilježeni vrućica (78, 8%), osip (15, 5%), artralgija (18, 8%), mijalgija (14, 4%) i glavobolja (6, 6%). Detekcijom IgM/IgG protutijela dokazana je akutna/nedavna importirana arbovirusna infekcija kod 11 bolesnika. Zika virusna infekcija zabilježena je kod dvije bolesnice (povratnice iz Vijetnama i Tajlanda), chikungunya groznica kod jedne bolesnice (povratnica iz Brazila) te dengue groznica kod 8 bolesnika (povratnici s Tajlanda, Kube, Tanzanije, Somalije, Indonezije i Filipina). U dva su ispitanika dokazana IgG protutijela na DENV te u dva ispitanika IgG protutijela na CHIKV. U samo jednom testiranom uzorku dokazana je prisutnost DENV RNA metodom RT-PCR. Filogenetska analiza potvrdila je da se radi o DENV tip 1.

Zoonoze, bolesti koje se prirodno prenose sa životinja na ljude, oduvijek su bile zajedničko područje rada veterinara i liječnika. Kroz blisku suradnju mnoge su odavno poznate zoonoze uspješno stavljene pod nadzor ili čak i potpuno iskorijenjene na pojedinim područjima. S druge strane, posljednjih desetljeća suočeni smo s novim izazovima kroz jasno naglašen trend pojave sve većeg broja emergentnih bolesti čiji nadzor je nemoguć bez interdisciplinarne suradnje u skladu s pristupom Jedno zdravlje. Analizirajući pojavu tih sve učestalijih emergentnih bolesti, jasno se ističe da su one u najvećem postotku zoonoze, te da među njima poseban značaj imaju one prenosive vektorima. Ovo je lako objašnjivo globalizacijom prometa ljudi, životinja, roba i usluga, uz istovremene klimatske promjene koje omogućavaju širenje staništa različitih vektora zaraznih bolesti kao i unošenje novih vektorskih vrsta na nova područja. Možda i najbolji primjer ovakvih trendova su flavivirusne infekcije čijem širenju svjedočimo posljednjih godina na globalnoj razini, ali i na području Republike Hrvatske. Na području naše domovine samo u posljednjem desetljeću svjedočili smo autohtonoj dengue groznici, prvim slučajevima bolesti Zapadnog Nila i Usutu virusne infekcije u ljudi koje su danas udomaćene, kao i povremenim re-emergencijama krpljenog encefalitisa, dijelom uvjetovano alimentarnim infekcijama mlijekom i mliječnim proizvodima. Uz opisano, importirani slučaj Zika virusne infekcije, dovoljno naglašava nužnost sustavnog nadzora zaraznih bolesti uzrokovanih flavivirusima radi očuvanja javnog zdravstva. Nadzor i preventivno djelovanje na suzbijanju ovakvih bolesti može se provoditi bilježenjem oboljenja ljudi i posljedičnim provođenjem epidemioloških mjera. Problem ovakvoga sustava je kasno djelovanje, u trenutku kada je već narušeno zdravlje i ugroženi ljudski životi. Za razliku od toga, provođenje nadzora flavivirusnih infekcija u životinja omogućava nam preventivno usmjereno djelovanje otkrivanjem područja virusne aktivnosti prije oboljenja ljudi. Na ovaj način usmjeravajući protuepidemijske mjere na područja s dokazanim izvorima infekcije, umanjujemo vjerojatnost infekcije i oboljenja ljudi. Primjeri koji podupiru ovakvo djelovanje su zasigurno dokaz prisustva i aktivnosti virusa Zapadnog Nila kroz program nadzora infekcija konja dvije godine prije prvog oboljenja u ljudi zabilježenoga 2012. godine. Upravo provedba ovog programa bila je osnova za predviđanje područja i razdoblja pojave prvih oboljenja ljudi što je omogućilo pravovremeno prepoznavanje i suzbijanje ove bolesti. Kasnijim unaprjeđenjem ovog sustava nadzora kroz praćenje infekcija divljih ptica i peradi te povremeno praćenjem prisustva uzročnika u vektorima, zasigurno je spriječen znatan broj infekcija i oboljenja ljudi.Nadzorom flavivirusnih infekcija u konja također je 2011. godine, dvije godine prije prvih oboljenja ljudi, potvrđeno prisustvo Usutu virusa, još jednog emergentnog flavivirusa, u središnjoj Hrvatskoj. Njegovo prisustvo dalje se pratilo i kroz infekcije ptica što je omogućilo i molekularnu tipizaciju te zahvaljujući tome dodatne epidemiološke spoznaje. Uz navedeno praćenjem infekcija koza, konja i pasa određivala su se rizična područja za infekciju virusom krpeljnog encefalitisa. Uspostava i provedba dijagnostike ovih infekcija u životinja omogućila je i dokaz alimentarne infekcije mlijekom i mliječnim proizvodima koza kao dodatni doprinos razumijevanju epidemiologije ove bolesti s mogućim teškim posljedicama. Svi navedeni uspjesi i doprinosi javnom zdravstvu ostvareni kroz praćenje flavivirusnih infekcija u životinja ostvareni su kroz blisku interdisciplinarnu suradnju koja je potvrđeno najbolji put za suočavanje s izazovima emergentnih bolesti. S druge strane zasigurno zabrinjava što usprkos jasnim trendovima u pojavljivanju emergentnih bolesti te objektivnim uspjesima ovakvog praćenja infekcija u životinja, nije dovoljno razvijena svijest o potrebi stalnih i dugoročnih programa nadzora nego se opseg i izvedba programa učestalo mijenja temeljem političkih, a ne stručnih odluka. Neovisno o tome, uspostavljenom bliskom stručnom i znanstvenom suradnjom, u okviru objektivnih mogućnosti, ovakvi programi se nastavljaju što i dalje rezultira novim spoznajama koje su temelj za nadzor i suzbijanje emergentnih flavivirusnih infekcija. Ovakva suradnja u praćenju flavivirusnih infekcija najbolja je potvrda uspješnih rezultata zajedničkog rada, u skladu s pristupom Jedno zdravlje, u očuvanju zdravlja ljudi i životinja od ovih, ali i mnogih drugih sadašnjih i budućih emergentnih zoonoza.

Arbovirusi čine veliku skupinu s >500 poznatih virusa, od kojih su većina uzročnici zoonoza. Na području Hrvatske, dokazane su kliničke infekcije uzrokovane virusom krpeljnog encefalitisa (TBEV), West Nile (WNV), Usutu (USUV), virusom papataci groznice i Toscana (TOSV) uz serološki dokazanu prisutnost Tahyna (TAHV) i Bhanja virusa (BHAV). Infekcije uzrokovane virusom dengue (DENV), Zika (ZIKV) i chikungunya (CHIKV) se bilježe sporadično u putnika povratnika iz endemskih područja. Za velik broj arbovirusnih infekcija ljudi predstavljaju slučajne krajnje domaćine s kratkotrajnom i niskom viremijom (iznimka su npr. žuta groznica, dengue i chikungunya), zbog čega je izravna dijagnostika često neuspješna. Stoga se u dijagnostici ovih infekcija najčešće primjenjuju serološki postupci. U rutinskoj se dijagnostici koriste imunoenzimni test (EIA ; enzyme immunoassay) i indirektni imunofluorescentni test (IFA ; indirect immunofluorescence assay) kojima je moguće razlikovati pojedine klase protutijela. Imunoblot testovi (western blot, LIA ; line immunoassay) imaju veću specifičnost u odnosu na EIA i IFA test jer dokazuju protutijela na različite virusne antigene. Međutim, pojedine skupine virusa kao npr. alfavirusi ili flavivirusi često pokazuju križne reakcije, posebice između virusa koji pripadaju istoj seroskupini te je u tim slučajevima reaktivne rezultate potrebno potvrditi dokazom neutralizacijskih protutijela (test neutralizacije virusa ; VNT, neutralizacijski test redukcije plakova ; PRNT). Rijetko su moguće križne reakcije i u VNT testu, no u tim je slučajevima homologni titar obično znatno viši od heterolognog. Kao dodatni test za potvrdu akutne/nedavne primarne infekcije koristi se određivanje aviditeta IgG protutijela. Ovaj se test pokazao korisnim u dijagnostici flavivirusnih infekcija za razlikovanje akutne/nedavne infekcije u bolesnika s dugom perzistencijom IgM protutijela, te križno-reaktivnim IgM protutijelima (dengue, West Nile infekcije, krpeljni encefalitis). U Referentnom centru Ministarstva zdravstva za dijagnostiku i praćenje virusnih zoonoza pri Hrvatskom zavodu za javno zdravstvo rutinski se provodi molekularna i serološka dijagnostika arbovirusnih infekcija (tablica 1). U suradnji s Veterinarskim fakultetom Sveučilišta u Zagrebu i Hrvatskim veterinarskim institutom, VNT je uveden u dijagnostiku flavivirusa (TBEV, WNV, USUV). Po potrebi, uzorci se šalju na potvrdno testiranje u OIE Referentni centar za West Nile virus, Istituto Zooprofilattico Sperimentale "G. Caporale", Teramo, Italija.

Svjetska zdravstvena kriza nastala brzim širenjem korona virusne bolesti (COVID-19) uzrokovala je velike probleme u svim područjima zdravstva. Transplantacijska medicina jedan je od najpogođenijih segmenata budući da se oslanja na brojne djelatnosti koje su sada preusmjerene na liječenje COVID-19 pacijenata. Pandemija je tako imala mnogostruke učinke na transplantacijsku medicinu, ne samo zbog smanjenog broja provedenih transplantacija, već i zbog odgode kontrolnih pregleda i testova probira što bitno narušava kvalitetu skrbi i onemogućuje pravovremeno liječenje. U oporavku od pandemije, može se očekivati prekoračenje zdravstvenih kapaciteta, povećan broj pacijenata dijagnosticiranih u težim stadijima bolesti i posljedično porast broja potrebnih transplantacija. U Hrvatskoj, prvi COVID-19 slučaj zabilježen je 25. veljače 2020. godine, što je dovelo do privremene stanke u transplantaciji solidnih organa osim za urgentne kandidate za transplantaciju jetre. Hrvatska transplantacijska mreža uspostavila je svoje uobičajene aktivnosti početkom svibnja 2020. godine. Od tada se transplantacije solidnih organa odvijaju nesmetano, uz obavezno COVID-19 testiranje prije zahvata svih darivatelja i primatelja. U jeku COVID-19 pandemije američka i europska transplantacijska društva objavila su svoje prve preporuke koje se kontinuirano dorađuju. U slučaju pozitivnog COVID-19 darivatelja, organi se neće koristiti za transplantaciju, dok će se u slučaju pozitivnog COVID-19 primatelja, transplantacija odgoditi do negativizacije nalaza. Prethodno preboljela (14-21 dan) COVID-19 infekcija nije prepreka za transplantaciju organa. Potrebno je uzeti u obzir i mogućnost lažno negativnih testova te sve pacijente i darivatelje s karakterističnim simptomima COVID-19 infekcije tretirati kao potencijalno zaražene uz provođenje dodatnih dijagnostičkih testova. Transplantirani pacijenti ugroženi su stoga zbog poteškoća u organizaciji zdravstvene skrbi, no i zbog mogućnosti same SARS-CoV-2 infekcije te za sada još nedovoljno istraženih posljedica te infekcije. Nakon transplantacije imunosupresivna terapija sprečava aloimuni odgovor, međutim, zbog imunokompromitiranosti, transplantirane osobe su pod povećanim rizikom od svih infekcija. COVID-19 infekcija zahvaća sve dobne skupine transplantiranih pacijenata te se prvenstveno povezuje s respiratornim simptomima, no može uzrokovati i sustavnu bolest te zahvatiti probavni sustav, srce, bubrege i imunološki sustav. Za sada još uvijek nema dovoljno dokaza da je COVID-19 infekcija teža u transplantiranih nego u ne-transplantiranih osoba te nije poznato predisponira li imunosupresivna terapija pacijente za teži oblik bolesti. Prekid imunosupresivnih lijekova kod transplantiranih pacijenata može dovesti do odbacivanja presatka ili ponovne pojave osnovne bolesti, što bi u konačnici imalo nepovoljniji ishod za pacijenta od same COVID-19 infekcije.Oporavak od SARS-CoV-2 infekcije u transplantiranih pacijenata može potrajati dulje nego u ne- transplantiranih osoba. Pri tome je važno da redukcija imunosupresivne terapije, ako je potrebna, bude strogo nadzirana od strane transplantacijskog tima. Današnja prva saznanja o COVID-19 i transplantaciji jetre u Europi, temelje se na podacima Europskog registra za transplantaciju jetre (ELTR) koji sadrži podatke za 103 COVID-19 pozitivne osobe, uglavnom s područja Italije, Španjolske i Francuske. Nakon transplantacije jetre COVID-19 infekcija u većine se očitovala prepoznatljivim simptomima vrućice, kašlja i nedostatka daha. Oko 20% pacijenata bez značajnijih dišnih tegoba praćeno je kod kuće, a ostali su zaprimljeni u bolnicu. Većina hospitaliziranih imala je upalu pluća te zahtijevala terapiju kisikom, a 15% liječeno je mehaničkom ventilacijom. Od ukupno 103 COVID-19 pozitivna pacijenta nakon transplantacije jetre, 16% je umrlo, a svi pacijenti kod kojih je došlo do smrtnog ishoda bili su stariji od 60 godina i više od dvije godine nakon transplantacije. U Hrvatskoj su podaci o COVID-19 infekciji nakon transplantacije jetre oskudni. Pilot studija seroprevalencije COVID-19 provedena tijekom rujna 2020. godine koja je uključila 137 osoba nakon transplantacije jetre pokazuje IgG seropozitivitet dokazan ELISA testom od 5, 1%. Neutralizacijska protutijela dokazana su u 2, 2% pacijenata. Današnja znanja o COVID-19 infekciji u transplantiranih osoba još su uvijek limitirana i ne možemo donositi čvrste zaključke, stoga prevencija ostaje najvažnija mjera u borbi protiv ove bolesti.

Krajem 2019. godine pojavio se novi koronavirus SARS-CoV-2 koji je do danas uzrokovao globalnu pandemiju. U terenskim uvjetima dokazi infekcije u pasa i mačaka su sporadični. Početna istraživanja dokazala su prijemljivost pasa i mačaka na infekciju ovim uzročnikom u laboratorijskim uvjetima, ali je značaj kućnih ljubimaca u epidemiologiji SARS-CoV-2 i dalje nejasan. Ovo istraživanje prikazuje rezultate prvog opsežnog serološkog pretraživanja uzoraka seruma pasa i mačaka u Europi. Od 26. veljače 2020. godine, samo jedan dan nakon prvog potvrđenog slučaja infekcije SARS-CoV-2 u ljudi u Hrvatskoj, do 15. lipnja 2020. godine prikupljani su ostatni uzorci seruma pasa i mačaka zaprimljenih u tri veterinarske organizacije s područja Zagreba i Splita. Ukupno je prikupljeno 656 uzoraka seruma pasa i 131 uzorak seruma mačaka koji su pretraženi virus neutralizacijskim testom razvijenim za potrebe ovog istraživanja. Od navedenih uzoraka nasumično je odabrano 172 uzorka seruma pasa koji su dodatno pretraženi imunoenzimnim testom. Korišteni imunoenzimni test nastao je preinakom komercijalno dostupnog kompleta koji je prilagođen pretraživanju uzoraka seruma pasa na prisutnost protutijela za SARS-CoV-2 virus. Neutralizirajuća protutijela su dokazana u 0, 76% seruma mačaka. Seroprevalencija SARS-CoV-2 infekcije u mačaka u Hrvatskoj je niska, osobito kada se usporedi sa seroprevalencijom u mačaka u Wuhanu. S druge strane, prevalencija neutralizirajućih protutijela u mačaka je slična udjelu mačaka koje su pokazale pozitivan rezultat imunoenzimnog testa u Francuskoj. Kako se čini u terenskim uvjetima infekcija u mačaka u Europi je rijetka. Kod pasa u 0, 31% seruma dokazana je neutralizacijska aktivnost te su imunoenzimnim testom protutijela dokazana i u 7, 56% pretraženih životinja. U dostupnoj literaturi ne postoje podaci o sličnim istraživanjima za usporedbu. Što je zanimljivo prvi, imunoenzimnim testom, pozitivni uzorak seruma psa uzorkovan je u Zagrebu 17. travnja, oko sedam tjedana nakon prvog dokaza infekcije u ljudi u istom području, a najveći broj pozitivnih pasa je uzorkovan šest tjedana nakon najvećeg zabilježenog broja infekcija u ljudi što govori u prilog da dinamika infekcije SARS-CoV-2 u pasa prati dinamiku infekcije u ljudi te nije bilo naznaka o pojavi neovisne epizootije među pretraživanim psima. Zaključno, niska seroprevalencija i nizak titar neutralizirajućih protutijela u pozitivnih životinja govori u prilog da su infekcije u kućnih ljubimaca rijetke. Usporedbom rezultata seroprevalencije u kućnih ljubimaca i širenja infekcije SARS-CoV-2 u populaciji ljudi u Hrvatskoj nema dokaza da se ovaj virus širio neovisno među kućnim ljubimcima već da su primaran izvor infekcije za pse i mačke zaraženi ljudi.

Sredinom prosinca 2020. godine zabilježeno je grupiranje bolesnika s atipičnom virusnom pneumonijom nepoznate etiologije za koje je utvrđeno da su uzrokovane SARS-CoV-2 virusom zoonotskog porijekla, a bolest je nazvana COVID-19. Uskoro je postalo jasno da infekcija neće ostati lokalizirana u Wuhanu, u Kini već da će se proširiti te su poduzeti koraci za uspostavljanje dijagnostike u laboratorijima u kojima se inače provodi detekcija virusa influence. Nužno je bilo da laboratorij posjeduje biozaštitni kabinet 2. kategorije. Uskoro je razvijeno nekoliko protokola koje je podijelila Svjetska zdravstvena organizacija kako bi olakšala i ubrzala uvođenje dijagnostike COVID-19. Tako je i Nacionalni centar za gripu, HZJZ-a također započeo s pripremama za uspostavljanje dijagnostike. Neposredno nakon izbijanja epidemije u Italiji, u Hrvatskoj je zabilježen prvi slučaj 25. veljače 2020. godine. Dijagnostički pristup HZJZ-a je slijedio preporuke SZO te je postavljena metoda Corman i sur. Od početka epidemije u Hrvatskoj, testirali smo više od 74 000 uzoraka na SARS-CoV-2, od čega je 4647 bilo pozitivnih. Dokaz prisutnosti virusa u uzorku molekularnim metodama (lančana reakcija polimeraze s reverznom transkripcijom, RT-PCR) još uvijek je metoda izbora za dokaz akutne infekcije. Međutim, za procjenu proširenosti infekcije u populaciji jako je važno provesti serološka testiranja. Serologija je važna i kao dodatna metoda za potvrdu klinički jasnih slučajeva s negativnim RT-PCR nalazom na COVID-19 u osoba koje su bliski kontakti oboljelih. Napravljena je usporedba deset seroloških testova za dijagnostiku SARS-CoV-2. Korištena su pet 'point-of care' (POC) imunokromatografskih testova (ACRO, AMP, ENCODE, NG BIOTECH i WIZ BIOTECH), četiri imunoenzimska testa (EIA ; DiaPro, Vircell ; Euroimmun i NovaTec) te imunoenzimski test s fluorescentnom detekcijom (ELFA ; Mini Vidas). Testirani su uzorci seruma bolesnika s COVID-19 infekcijom te asimptomatskih osoba. EIA testovi su pokazali višu osjetljivost detekcije IgM/IgA protutijela u usporedbi s POC testovima, dok je osjetljivost detekcije IgG protutijela bila podjednaka u POC i EIA testovima. Budući da je za validaciju novih seroloških testova za detekciju protutijela na SARS-CoV-2 bilo preporučeno inicijalno reaktivne rezultate potvrditi testom neutralizacije virusa (VNT) koji je još uvijek "zlatni standard" za potvrdu seroloških testova bilo je potrebno izolirati SARS-CoV-2. Virus je uzgojen u staničnoj kulturi na Vero E6 stanicama iz nazofaringealnog obriska COVID-19 bolesnika u BSL-3uvjetima (biozaštitni kabinet 3. kategorije). Maksimalni citopatski učinak bio je vidljiv 4. dan nakon inokulacije i prisutnost virusa potvrđena je RT-PCR-om. Usporedili smo i rezultate VNT s rezultatima surogat VNT testa (GENSCRIPT SARS-CoV-2 Surrogate Virus Neutralization ELISA) koji je pokazao znatno nižu osjetljivost u uzorcima s niskim titrom neutralizacijskih protutijela detektiranih klasičnim VNT (8 i 16). Dodatno smo analizirali prisutnost SARS-CoV-2 u okolini COVID-19 bolesnika kako bismo procijenili obim prisutnosti virusa u okolišu oboljelih. Od testiranih 41 uzoraka, nađeno je 3 (7, 3%) pozitivnih okolišnih uzoraka (mobitel, upravljač igraće konzole, prozorska kvaka).

SARS-CoV-2 is a novel coronavirus that has spread among the human population at the end of 2019. It has been reported over 130 million outbreaks and around 3 million deaths worldwide so far. An animal origin of this coronavirus is generally accepted in the scientific community, and it brings the question about the epidemiological role of animals in COVID-19. SARS-CoV-2 infections in dogs have been described, and their susceptibility to infection has also been proven in experimental infections. Dogs as pet animals have a potentially significant role in the epidemiology of COVID-19 due to their close contact with owners. From July 1st to December 31st 2020, 1069 remaining dog sera samples from the city of Zagreb and Zagreb County were collected. Additionally, collected data included signalment, medical history and clinical signs. Serum samples were tested by enzyme- linked immunosorbent assay (ELISA), which was developed and validated for this study. For the statistical analysis of results, a chi-squared test was used. Of the total number of dogs tested, SARS-CoV-2 antibodies were confirmed by an ELISA test in 14.69%. The highest number of positive samples was in September and December 2020. There was gender predisposition with significantly increased seroprevalence in male dogs (p=0, 007). Unlike in the human population, age predisposition has not been proven (p=0.2). Breed predisposition was also pronounced (p=0.01). Belgian Shepherds were significantly more likely to be infected than any other dog breeds. The serological status of dogs could not be related to the occurrence of respiratory or gastrointestinal clinical signs (p = 0.37). There was no relation between the SARS- CoV-2 status of dogs and the reason for the veterinary visit. Dogs from shelters were not at higher risk of SARS-CoV-2 infection than private- owned ones (p = 0.96). This study shows that SARS- CoV-2 has become a significant pathogen among the dog population. At the end of the pandemic's first wave, the seroprevalence in Croatia was 7.45% and now is two times higher. Although it has not been proven that dogs have a significant role in the epidemiology of infection in humans, there is a question of the possible impact of SARS-CoV-2 on animal health.

In the season 2020/2021 two outbreaks of highly pathogenic avian influenza (HPAI) were detected in Croatia, one in northern and the other in eastern Croatia at a distance of approximately 200 kilometers apart. In both cases, the H5N8 subtype virus was detected. The first outbreak was recorded in mid-November 2020 at a fattening turkey farm, and the second outbreak in early March 2021 in three dead mute swans found within a four-kilometer diameter. The whole genome sequencing revealed that both outbreaks were caused by viruses of the Guandong/96 lineage, clade 2.3.4.4b. Viruses from both outbreaks in the 2020/2021 season are similar to HPAI viruses of H5 subtype detected in wild birds and poultry in Russia and Kazakhstan since July 2020 and with these viruses detected in northern, western and central Europe since October 2020. A more detailed analysis indicates a very close association of the virus detected in Croatia in a turkey farm in November 2020 with viruses detected in Western Europe in the autumn of the same year, especially with viruses detected in Germany, Sweden, Italy, the Czech Republic and Poland. In contrast, the virus detected in dead swans in March 2021 is closely related to viruses detected in early 2021 in Russia, particularly to viruses detected in poultry in Krasnodar and Kurgan. Although the introduction of the virus into the turkey farm in November 2020 remains unclear, it can undoubtedly be concluded that it is not directly related to viruses detected in wild birds in Croatia in March 2021 and there were at least two independent introductions of this virus. In order to prevent the introduction of HPAI viruses into poultry farms and avoid major economic losses, active wild bird monitoring is necessary, especially during autumn in areas where these viruses were detected.

Coronavirus disease 2019 (COVID-19) is an infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). It usually presents with mild common cold-like symptoms. However, it can lead to series of complications with some of them being fatal. One of the rare and potentially neglected manifestations of COVID-19 is subacute thyroiditis (SAT). Viral causes of SAT documented so far include several viruses, but according to the newest reports, SARS-CoV-2 should also be added to the list. The median time between COVID-19 diagnosis and the onset of SAT symptoms is reported to be 29 days. Patients mostly present with triphasic course of symptoms - thyrotoxicosis, followed by hypothyroidism and euthyroidism. Although limited number of SAT cases after SARS-CoV-2 infection are reported, physicians should consider SAT as a differential diagnosis in COVID-19 patients. The routine assessment of thyroid function in patients with clinical suspicion of SAT or with the suspicion of any other thyroid dysfunction is advised., Koronavirusna bolest 2019 (COVID-19) infektivna je bolest koju uzrokuje SARS-CoV-2 (engl. severe acute respiratory syndrome coronavirus 2). Uglavnom se očituje blagim simptomima nalik na običnu prehladu, ali može izazvati niz teških komplikacija, od kojih pojedine mogu biti i smrtonosne. Jedna od rijetkih i potencijalno zanemarenih manifestacija bolesti COVID-19 je subakutni tireoiditis (SAT). Do sada je dokazano nekoliko virusnih uzročnika SAT-a, no kako najnovija istraživanja upućuju, SARS-CoV-2 bi također trebao biti uvršten na taj popis. Nadalje, izvještaji ukazuju da medijan između dijagnoze COVID-19 i početka simptoma SAT-a iznosi 29 dana. Klinički se mogu razlikovati tri stadija bolesti - stadij tireotoksikoze nakon čega slijede hipotireoza te eutireoza. Unatoč ograničenom broju objavljenih slučajeva SAT-a nakon SARS-CoV-2 infekcije, liječnici bi trebali razmotriti SAT kao diferencijalnu dijagnozu kod pacijenata oboljelih od COVID-19. Tijekom praćenja i kontrole oboljelih savjetuje se i rutinska procjena funkcije štitnjače u onih pacijenata kod kojih postoji klinička sumnja na SAT ili neki drugi poremećaj u radu štitnjače.

Background: Tick-borne encephalitis (TBE) is the most important re-(emerging) arboviral zoonosis in Europe. Although up to 80% of TBE cases are asymptomatic or presented as mild febrile illness, 20-40% patients develop neuroinvasive disease (meningitis, meningoencephalitis, meningoencephalomyelitis). While clinical characteristics of TBE are well described, data on the antiviral cytokine response to TBEV are limited. Methods: A total of 32 serum and 16 cerebrospinal fluid (CSF) samples from 33 patients with TBE were included in the study. A multiplex bead-based assay for the simultaneous quantification of 13 human antiviral cytokines was used. The panel included cytokines associated with innate and early pro-inflammatory immune responses: TNF-α, IL-6, IL-2, IFN-γ ; Th2 immune response: IL-4, IL-5, IL-9, IL-13 ; Th17 immune response: IL-17A, IL-17F, IL-21, IL-22 and the key anti-inflammatory cytokine IL-10. Results: Neuroinvasive infection was documented in 29/87.8% patients (meningitis 18/54.5%, meningoencephalitis 11/33.3%), while 4/12.2% of patients presented with nonspecific febrile disease. Significantly higher concentrations of IL-6 in the CSF vs. serum were found in paired CSF/serum samples (median 780.88 pg/mL, IQR 130.81-3, 077.66 vs. median 23.25 pg/mL, IQR 7.79-27.09 ; p

(Re-)emerging flavivirus infections represent a public health problem in many parts of the world. Among neuroinvasive flaviviruses, tick-borne encephalitis virus (TBEV), West Nile virus (WNV) and Usutu virus (USUV) are most widely distributed. WNV strains are classified into several genetic lineages of which lineage 1 and 2 are the most widespread. Recent data from the European countries showed that WNV strains detected in humans mainly belong to lineage 2. USUV strains are grouped into 8 lineages: five European (1-5) and three African (1-3). Most human USUV strains belong to European USUV lineages (mainly Europe 2), however African lineages were also documented. The TBEV European subtype is most prevalent in western, northern and Eastern Europe and European parts of Russia, with Far East and Siberian subtypes co-circulating in Eastern Europe. We analyzed the molecular epidemiology of neuroinvasive flaviviruses detected in Croatia during the four consecutive transmission seasons (2017-2020). TBEV was confirmed in 43 patients (2017-2020), WNV in 61 (2017-2018) and USUV in three patients (2018). Viral RNA was detected in cerebrospinal fluid and/or urine samples using a real-time RT-PCR. Samples identified as positive using the real-time RT-PCR assays were subjected to conventional RT-PCR and Sanger sequenced. One TBEV strain detected in the urine sample of a patient with severe neuroinvasive infection (2017) belonged to TBEV European subtype. All 11 sequenced WNV strains (3 from 2017 and 8 from 2018) showed circulation of WNV lineage 2. One USUV strain from a fatal encephalitis case (2018) clustered within Europe 2 lineage. Molecular epidemiology of flaviviruses detected in Croatian patients showed a similar pattern as in other European countries.

At the end of 2019, a new zoonotic coronavirus SARS-CoV-2 appeared, which caused a global pandemic and represents the world most important public health challenge. Initially, sporadic cases of SARS-CoV-2 infection have been reported in dogs, but with the increased number of disease outbreaks in humans, the number of recorded infections of dogs worldwide is increasing. The primary source of infection in dogs is humans, but dogs' role in the epidemiology of SARS-CoV-2 is still insufficiently investigated. At the Faculty of Veterinary Medicine University of Zagreb, a study is being conducted about the impact of living in COVID-19 positive households on pet animals' health. Owners who had recovered from COVID-19 were invited to sign their dogs for physical examination and serological testing for SARS-CoV-2. Based on epidemiological and epizootiological parameters, 78 dogs were selected for the serosurvey. Serum samples were tested by enzyme-linked immunosorbent assay (ELISA) and microneutralisation test (MNT) as a confirmatory method. Both used methods were developed and validated in the Virology Unit of the Faculty of Veterinary Medicine, University of Zagreb. Of the total number of dogs tested, IgG antibodies to SARS-CoV-2 were confirmed by ELISA in 43.59%, while neutralisation antibodies were confirmed in 25.64% of dog serum samples. Seroprevalence in this study was significantly higher than studies in the United States, Italy and France. In later studies, neutralisation antibodies were detected in between 11.9% and 15.4% of tested dogs. In the general population of dogs in Zagreb, whose owners have an unknown status for COVID-19, the seroprevalence determined by ELISA during the first wave of the pandemic was 7.45%. In comparison, during the second wave, the percentage of infected increased to 14.69%. Compared to the above, the seroprevalence determined by this study is significantly higher, which confirms the increased risk of SARS-CoV- 2 virus infections in dogs from COVID-19 positive households and confirms that COVID-19 is a reverse zoonosis. The high percentage of infections in dogs and the fact that COVID-19 is an asymptomatic infection in a certain number of people also raises the question of the possible use of dogs as sentinel animals, but this needs further investigation.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a novel coronavirus that emerged in Wuhan, China in December 2019. Coronavirus disease (COVID-19) spread rapidly and in March 2020, WHO declared a pandemic. With global attention on SARS-CoV-2, many other viral infections remain neglected in the shadow of COVID- 19. However, (re-)emerging arboviruses still represent an emerging public health threat. Arboviruses detected in humans in Croatia (acute infections or serologic evidence) are tick-borne encephalitis virus (TBEV), West Nile virus (WNV), Usutu virus (USUV), Toscana virus (TOSV), Tahyna virus (TAHV) and Bhanja virus (BHAV). During the 2020 transmission season, a total of 82 patients with neuroinvasive disease, 15 patients with non- specific febrile disease and 34 travelers returning from endemic areas were tested for arboviruses. TBEV was confirmed u 10 patients, while other neuroinvasive arboviruses were not detected. Clinical presentations were febrile headache, meningitis and meningoencephalitis. All patients recovered. Majority of them reported risk behaviors such as frequent visiting of forest areas and frequent tick bites. TBEV infections occurred from May to July in continental Croatian counties. Previous TBEV, WNV, USUV and TOSV infection (IgG antibodies) was detected in 1.2%, 4.9%, 2.4% and 1.2% of patients, respectively. Additionally, 2.4% of patients showed IgG antibodies to sandfly fever Naples virus. Imported Zika virus infection was documented in a traveler returning from Thailand. During the same period, a total of 188 asymptomatic persons were tested to determine the seroprevalence of neuroinvasive arboviruses. IgG prevalence was 1.6% to TBEV, 3.2% to WNV and 1.1% to USUV. Like in other European countries, arbovirus infections in Croatia in 2020 were, at least partly overshadowed by COVID-19 pandemic.

SARS-CoV-2 as an RNA virus with a large genome is subject to rapid mutation as it replicates. Among silent or deleterious mutations are the one that creates variants with a selective advantage. In late 2020 and early 2021 emerged variants that appear to be more transmissible than existing strains, evade immunologic response and induce more severe disease. Three variants that rapidly became dominant in specific area and were designated variant of concern (VOC) are: VOC- 202012/01 (B.1.1.7), 501Y.V2 (B.1.351) and P.1 (B.1.1.28.1). To this date GISAID acknowledges two more VOCs: GH/452R.V1 (B.1.429+B.1.427) and G/484K.V3 (B.1.525). To monitor the epidemiologic situation we followed ECDC recommendation and conduct whole-genome sequencing (WGS) on 5-10% SARS-CoV-2 positive samples in Croatia. WGS is performed weekly from 09.02.2021. Obtained sequences are uploaded to GISAID database and isolates are categorized in phylogenetic clades depending on single nucleotide polymorphism (SNP) present in the genome. By the end of March 2021 we processed 2632 samples. 1901 samples were successfully sequenced. Between the second and third waves of the epidemic, virus lineages from clade G, GR and GV were dominant, but marked weekly increase of GRY clade indicated the beginning of the third epidemic wave in Croatia. Delivered results showed marked continuous increase in B.1.1.7 presence from 21.23% in the first week of testing, to 94.27% in the last week of March. Variant B.1.351 from GH clade was detected in 15 samples, mainly related to travelling abroad and their close contacts. B.1.526 detected in 4 samples carry mutation essential for the loss of neutralizing activity of some monoclonal antibodies and is considered variant of interest.

The rapid spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has led to the coronavirus disease 2019 (COVID-19) worldwide pandemic which caused a worldwide crisis with profound effects on public health. According to the World Health Organization (WHO), there have been over 130 million confirmed cases and almost 3 million fatalities since December 2019. In Croatia, the first case was detected on February 25, 2020. Reference Centre for laboratory diagnosis of viral respiratory diseases Croatian Ministry of Health at the Croatian Institute of Public Health (CIPH) implemented SARS-CoV-2 molecular diagnostics according to the WHO protocols. Molecular testing is still the mainstay of diagnosis, supplemented by point-of-care antigen testing. CIPH reference laboratory analysed more than 200000 samples and 17.8% were positive. Rapid antigen testing was started in the November 2020 and till now we analysed more than 4000 samples, mostly symptomatic persons (42% were positive). While antibody testing is generally not useful for acute management, it can be used for retrospective diagnosis and seroprevalence studies to understand disease prevalence in the population. Two seroprevalence studies were conducted in the general population, after the 1st and 2nd wave, respectively. A total of 1088 and 1436 serum samples were tested for the presence of IgG SARS-CoV-2 antibodies, and IgG antibodies were detected in 2.2% and 25.2% participants, respectively. SARS-CoV-2 is believed to spread predominantly via short-range airborne aerosol, respiratory droplets, and direct or indirect contact with infectious respiratory droplets. Therefore, we have analysed environmental samples from COVID-19 patients’ surroundings and found the virus in 3/41 (7.3%) samples.