Search

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.

(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.

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.

The seasonality, clinical and laboratory features of West Nile virus (WNV) neuroinvasive infection overlap with those of enteroviruses. We present a case report on dual infection with WNV and echovirus 9 in a patient with aseptic meningitis. In August 2018, a 59-year-old woman with a history of arterial hypertension was hospitalized at the Clinic for Infectious Diseases on the third day of the illnesses. The disease was manifested by a daily fever up to 38.2°C, diffuse headaches and nausea without vomiting. On admission, her condition appeared severe. She was conscious, but she suffered from weakness and hypotonia. Neurologic examination revealed terminal neck stiffness. Routine laboratory parameters were normal. Cerebrospinal fluid (CSF) analysis showed pleocytosis (19x106 cells/L) with 53% mononuclear cells and elevated protein level (0.412 g/L). Electroencephalogram showed slow diffuse dysrhythmia. PCR of CSF for HSV 1/2, enteroviruses, CMV, EBV, B. burgdorferi and L. monocytogenes were negative. High titer of both WNV IgM (ratio 3.36 ; >1.1 positive) and IgG antibodies (142.81 RU/ml ; positive >22) of low avidity (avidity index 21% ; low

Norovirus is an important cause of non- bacterial gastroenteritis. Norovirus infects hosts of all ages and can cause large outbreaks in closed settings. Person-to-person transmission is essential in endemic cycle. Besides humans, norovirus can infect a range of different animals (e.g. pigs, cattle and mice). Therefore, zoonotic transmission of the infection is possible, indirectly, through contaminated food and water as vehicles, or directly, through animal contact. The aim of this study was to determine the prevalence and distribution of noroviruses in patients with gastroenteritis. The studied population included 107 patients aged from 4 months to 97 years (median age: 28 years) with clinical symptoms of gastroenteritis. Stool specimens from each patient were tested by using rapid immunochromatographic tests for rotavirus, adenovirus (Rota- AdenoGnost, BioGnost, Zagreb) and norovirus (Norovirus, Biognost, Zagreb), respectively according to the manufacturer’s recommendations. The overall prevalence rates for norovirus, rotavirus and adenovirus were 13.1%, 1.9% and 0.9%, respectively. Out of 14 documented norovirus infections, 2 (14.3%) were sporadic cases and 12 (85.7%) cases were identified as norovirus outbreaks in closed settings: 5 (35.7%) as nosocomial infections and 7 (50%) cases in a child care facility. The first outbreak occurred in March in a hospital setting, and the child care facility outbreak occurred in May. All 5 nosocomial norovirus infection and 1/7 cases in the child care facility occurred in adults. These findings highlight the importance of an early detection of norovirus cases for preventing the spread of the infection by implementing appropriate hand hygiene, environmental disinfection and isolation of the infected person in high-risk closed settings (e.g. healthcare). Our data also showed that norovirus is an important pathogen in children and, therefore, norovirus detection should be routinely preformed in children with gastrointestinal infections.

Introduction: West Nile virus (WNV) is one of the most widely distributed arboviruses. In Croatia, first clinical cases of WNV neuroinvasive disease were reported in 2012, thereafter cases were continuously detected. In addition, infections in horses and poultry were also documented. In 2018, the largest outbreak of WNV occurred with more than 60 human cases. For the first time in Croatia, WNV infections were detected in wild birds. Aims: The aims of this study were to analyze epidemiological characteristics and molecular epidemiology of WNV infections detected during 2018 transmission season. Methods: From January to December 2018, a total of 182 patients with neuroinvasive disease and 70 patients with symptoms compatible with WNV fever were tested for the presence of WNV RNA, IgM/IgG antibodies and IgG avidity. In addition, a total of 2759 horses were tested for the presence of WNV IgG antibodies. IgG positive serum samples were further tested for IgM antibodies for confirmation of recent infection. Thirty-five dead wild birds were tested for WNV RNA. Meteorological conditions (air temperature, precipitation) were also analyzed. Results: WNV infection (WNV neuroinvasive disease/WNV fever) was confirmed in 61 patients. Cases occurred in ten continental Croatian counties. Phylogenetic analysis of six detected strains showed circulation of WNV lineage 2. Acute asymptomatic infection was documented in 13/0.03% horses from three continental counties while IgG antibodies were detected in 232/8.4% horses. Seroprevalence rates were higher in continental counties (3.2- 26.0%) compared to counties on the Adriatic coast (0-4.8%). IgM positive horses were detected from March to June, while human infections occurred from July to October. Two WNV positive goshawks were detected in September. The early start of WNV season was associated with favorable climate conditions (very warm April and May). Conclusion: Our results confirm the need of continuous multidisciplinary ("One health") surveillance of this emerging viral zoonosis.

West Nile virus (WNV) belongs to the family Flaviviridae, genus Flavivirus. Birds are natural reservoir for WNV, mosquitoes are vectors, while humans and other vertebrates are considered as accidental hosts. Infections caused by WNV in humans vary in the presentation, from asymptomatic infections to a severe neuroinvasive disease. In Croatia, WNV infections are detected in horses, birds and humans using serological and molecular methods. Currently, there are nine genetic lineages proposed for classification of WNV and the lineages 1 and 2 are responsible for extensive epidemics in animals and humans. The aim of this study was to present the molecular epidemiology of WNV infections in Croatia in the transmission seasons 2017 and 2018. Viral RNA was sequenced using Sanger method in four human samples in 2017 as well as eight human and one sample from a dead goshawk (Accipiter gentilis) in 2018. Analysis of the NS5 gene segment has shown that all the strains belong to the lineage 2 and show high similarity with strains detected in different hosts in other European countries. Grouping of nucleotide sequences from our country into different clusters regardless the transmission season, location and host was observed. This indicates to probably multiple introduction of the virus into Croatia, intensive circulation and local overwintering of the virus. For the viral RNA from the dead goshawk the whole genome was sequenced in 12 partially overlapping segments using Sanger method. The whole genome shows the highest similarity (99.8%) with strains detected the same year in birds from Germany. Despite the high similarity, a deletion of eight nucleotides was found in the 3’ non- coding region of the genome which is not present in any of the available WNV sequences. Further studies at the molecular level of WNV detected in Croatia are needed to obtain better insight into the WNV molecular epidemiology and its local specificity.

Molecular epidemiology of West Nile virus (WNV) and Usutu virus (USUV) in Croatia during 2017 and 2018 was described. A total of 11 human WNV strains, one avian WNW strain as well as one human USUV strain, one avian USUV strain and one mosquito USUV strain were analyzed at the molecular level and compared phylogenetically with contemporary WNV and USUV viruses from other countries.

Tick-borne encephalitis virus (TBEV) and West Nile virus (WNV) are the most common arboviruses detected in Croatia. Sporadic infections caused by Usutu virus (USUV) and Toscana virus (TOSV) were also reported. Antibodies to Tahyna virus (TAHV) and Bhanja virus (BHAV) were found in 1970s and 1980s, but clinical cases were not reported so far. In 2018, a total of 194 patients with neuroinvasive disease were tested for the presence of arboviruses. WNV was the most commonly detected in 54 (27.8%) patients, TBEV in 18 (9.3%), USUV in 3 (1.9%) and TOSV in one (0.5%) patient. TAHV and BHAV were not detected. Infections caused by WNV, TBEV and USUV occurred in continental Croatian regions, while TOSV infection was detected at the Croatian littoral. During the same period, acute asymptomatic WNV infection (IgM antibodies) was documented in 20/2574 (0.8%) tested sentinel horses and 307 (11.9%) were IgG seropositive. In addition, WNV antibodies were detected in 125/1069 (12.7%) poultry serum samples (chickens, turkeys). Acute WNV infections in horses were recorded in March with sporadic occurrence until November. Human WNV infections were notified from July to October. In 2018, a re-occurrence of human USUV infections was reported. Furthermore, WNV and USUV infections in wild birds were detected for the first time in Croatia. USUV was also detected in one pool of the Culex pipiens mosquitoes collected in Zagreb. Phylogenetic analysis of detected strains showed circulation of WNV lineage 2 and USUV Europe 2 lineage. Detection of arboviruses in humans, sentinel animals and vectors confirms the importance of multidisciplinary ("One Health") surveillance of emerging arboviral infections.

INTRODUCTION: West Nile virus (WNV) is one of the most widely distributed arthropod-borne virus. In the last decades, WNV infections are detected in many European countries. In Croatia, first clinical cases of WNV neuroinvasive disease were reported in 2012 and thereafter, cases were continuously notified. In addition to human cases, serologic evidence of WNV infection was also recorded in sentinel animals (horses, poultry). OBJECTIVES: To analyze the prevalence of WNV in humans, horses, poultry and mosquitoes in Croatia in 2017. MATERIAL: During 2017 transmission season, a total of 90 patients with neuroinvasive infection (meningitis/encephalitis), 172 asymptomatic persons, 560 horses, 1580 poultry and 1186 mosquitoes (Culex pipiens and Aedes albopictus) were tested for the presence of WNV RNA and/or WNV antibodies. In patients with neuroinvasive infection, cerebrospinal fluid (CSF), serum and urine samples were collected. In asymptomatic subjects and sentinel animals serum samples were collected. METHODS: WNV IgM/IgG antibodies in human, horse and poultry sera were detected using a commercial ELISA. WNV RNA was detected in human and mosquito samples using real- time and nested RT-PCR. WNV positive human serum samples were confirmed using a virus neutralization test. RESULTS AND CONCLUSION: Neuroinvasive WNV infection was confirmed in 8 (8.9%) patients by detection of WNV IgM and IgG antibodies of low avidity and/or WNV RNA in CSF and urine. Phylogenetic analysis of four detected strains showed circulation of WNV lineage 2. Four (2.3%) asymptomatic persons were found to be IgG seropositive to WNV. In one participant, recent WNV infection was documented by low IgG avidity. WNV IgG antibodies were detected in 69 (12.3%) sentinel horses and 165 (10.4%) poultry. No one of the tested mosquito pool was found to be WNV RNA positive. Our results confirm the importance of multidisciplinary "One health" approach in the surveillance of this emerging viral zoonosis.

In last two decades, number of emerging and re- emerging viral zoonoses has increased. In Croatia, autochthonous infections caused by tick-borne encephalitis (TBEV), West Nile (WNV), Usutu (USUV) and Toscana virus (TOSV) were reported. Serologic evidence of Tahyna virus (TAHV) was documented in 1980s. Sporadic imported dengue (DENV), chikungunya (CHIKV) and Zika virus (ZIKV) infections were continously notified in travelers. During 2017, a total of 90 patients with neuroinvasive infection (meningitis/encephalitis), 172 asymptomatic persons, 560 horses and 1580 poultry were tested for the presence of neuroinvasive arboviruses. In addition, 72 travelers were tested for the presence of DENV, CHIKV and ZIKV antibodies. Neuroinvasive arboviral disease was confirmed in 19/21.1% patients: TBEV in 11/12.2% and WNV infection in 8/8.8% patients. Phylogenetic analysis showed circulation of WNV lineage 2. USUV, TOSV and TAHV infection was not detected. In asymptomatic persons, TBEV IgG antibodies were detected in 3/1.7% participants, WNV in 4/2.3% and TOSV in 4/2.3% participants. In two participants recent TBEV and WNV infection was confirmed by detection of borderline/low IgG avidity. Seroepidemiological survey in sentinel animals showed WNV IgG seropositivity in 165/10.4% poultry and 69/12.3% horses and TBEV IgG seropositivity in 80/14.3% horses. Imported arboviral infection (DENV, CHIKV, ZIKV) was reported in four travelers returning from areas with documented arbovirus circulation. Detection of emerging and re-emerging arboviruses in humans and sentinel animals in Croatia highlight the need of continuous multidisciplinary surveillance in accordance with "One health" approach.

In last two decades, the number of emerging and re-emerging mosquito-borne arboviral infections has increased. In Croatia, dengue (DENV), West Nile (WNV) and Usutu virus (USUV) infections have been detected. Autochthonous dengue was reported in 2010 on the Pelješac penninsula (south Adriatic coast). Seroepidemiological study performed during 2011-2012 showed seropositivity to DENV of 0.59%. First clinical cases of human neuroinvasive WNV infection were reported in 2012 with 38 cases notified until 2018 in eastern and north-western counties. Phylogenetic analysis of four strains detected during the 2017 outbreak showed circulation of WNV lineage 2. USUV antibodies were first detected in two asymptomatic horses (2011) and one human (2012), while neuroinvasive USUV infection was confirmed in 2013 in three patients from Zagreb and its surroundings. Autochthonous chikungunya (CHIKV) and Zika (ZIKV) infections were not detected so far. A seroepidemiological study conducted during 2011-2012 showed CHIKV IgG antibodies in 0.7% of inhabitants of the Croatian littoral. Clinically manifest imported chikungunya and Zika infections were reported in six returning travelers during 2016 and 2017. The recent emergence of several arboviral diseases in Croatia highlighted the role of mosquitoes as vectors of emerging arboviruses. In Croatia, 52 mosquito species have been detected. Medically most important species are Culex pipiens complex, the primary vector of WNV and USUV, and Asian tiger mosquito, Aedes albopictus, a vector of DENV, CHIKV and ZIKV. Aedes albopictus is an invasive mosquito species detected for the first time in 2004 in Zagreb with rapid spread in the subsequent years. Aedes japonicus is another invasive mosquito species detected in 2013. Recent molecular investigation showed that one pool of Ae. albopictus (2016) and Cx. pipiens complex (2017) were positive for USUV RNA. Detection of arboviruses in humans, sentinel animals and mosquitoes emphasize the importance of continuous multidisciplinary ("One health") arbovirus surveillance in Croatia.

In the last two decades, the number of emerging and re-emerging zoonoses has increased. Autochthonous infections caused by tick borne encephalitis (TBE), dengue (DENV), West Nile (WNV), Usutu (USUV), hepatitis E (HEV) and hantavirus as well as imported infections caused by chikungunya (CHIKV) and Zika virus (ZIKV) have been detected in Croatia. TBE occurs continuously since 1553. Didease is endemic in North-Western Croatian counries. Autochthonous DENV infections were reported in 2010 on the Pelješac peninsula. Seroepidemiological investigation performed during 2011-2012 showed seropositivity to DENV of 0.5%. First clinical cases of neuroinvasive WNV infection were reported in 2012 with 31 cases notified until 2017. in eastern and nort- western counties. USUV antibodies were found in one asymtomatic person from eastern Croatia in 2012, while neuroinvasive USUV was confirmed in 2013 in three patients from Zagreb and its surroundings. CHIKV infections were not detected so far, however antibodies were sporadically detected in persons who travelled ti endemic regions. A seroepidemiological study conducted during 2011-2012 showed CHIKV IgG antibodies in 0.7% of inhabitants of the Croatian littoral. First clinically manifest imported CHIKV infection was reported in 2016. During the same year, three clinically manifest imported ZIKV were also reported in Croatia. Autochthonous HEV infection was first described in 2012. A pilot study conducted in different population groups during 2014-2015 showed seropositivity to HEV of 5.6%. Hantavirus infection in Croatia are caused by Puumala and Dobrava viruses. About 10-20 cases are reported annually with several outbreaks (last one in 2014-2015). Leptospirosis and borreliosis are continuously reported bacterial infections. The most commonly detected leptospira serovars are Sejroe, Australis and Icterohaemorrhagiae. Q- fever is endemic in many Croatian regions wth seroprevalence rates 21.2-41.2%. Mediterranian spotted fever (R. connori) is the most commonly detected rickettsiosis. Toxoplasmosis is the most prevalent parasite infection.

The TORCH acronym includes the most common etiological agents of infections in pregnant women and newborns: T. gondii, rubella virus (RUBV), cytomegalovirus (CMV), herpes simplex viruses (HSV) type 1 and 2, as well as several other viruses: varicella-zoster virus (VZV), parvovirus B19, hepatitis B and C viruses (HBV, HCV), HIV, enteroviruses and bacteria. The "others" category has rapidly expanded, including emerging and re-emerging viral zoonoses. Most of the TORCH infections cause mild maternal morbidity, but have serious consequences on fetal health. In the last two decades, some zoonotic viruses have emerged such as hepatitis E virus (HEV) and Zika virus (ZIKV) which can cause infections in pregnant women and newborns. HEV infection may be more severe during pregnancy with fatal outcome in 15-25% pregnant women. ZIKV infection during pregnancy is associated with severe congenital malformations, including microcephaly. Materno- fetal transmission of re-emerging viruses such as dengue virus (DENV), chikungunya virus (CHIKV) and West Nile virus (WNV) was also reported. DENV infection during pregnancy is associated with higher rate of spontaneous abortion, intrauterine fetal death and premature birth. Congenital dengue fever is caused by the transplacental transmission of the virus during delivery. Most materno-fetal CHIKV transmissions occur during the peripartum period. Congenital WNV infections are possible but very rare ; they are associated with malformations, particularly of the brain. In the period from 2005 to 2015, several studies were conducted at the Croatian National Institute of Public Health to define the seroprevalence of TORCH infections. In childbearing-aged women tested from 2005-2009, the overall IgG seroprevalence rate was found to be 75.3% to CMV, 96.4% to RUBV, 69.4-78.7% to HSV-1, 5.8-10.2% to HSV-2, 63.5% to parvovirus B19 and 83.4% to VZV. Among viral hepatitis markers, HBsAg was detected in 0.5%, anti-HBc in 3.8% and anti-HCV antibodies in 0.5% women. Two pilot studies on the seroprevalence of emerging and re-emerging arboviruses (2011-2015) found seropositivity of 0.38% to DENV and 0.76% to CHIKV. In addition, recently conducted study (2015-2016) on the HEV seroprevalence showed seropositivity of 2.3%. A total of ten asymptomatic pregnant women who returned from endemic areas were tested for ZIKV, DENV and CHIKV during 2016-2017, of whom all were negative. During the tested period, acute CMV and VZV infections (positive IgM antibodies/low IgG avidity) were confirmed in 0.09% childbearing-aged/pregnant women. HSV-1 and HSV-2 IgM antibodies were found in 1.2% and parvovirus B19 IgM antibodies in 6.9% participants. In 0.36% women, recent asymptomatic WNV infection (positive IgM antibodies/borderline IgG avidity) was confirmed. In addition, imported CHIKV and ZIKV infections were detected in two febrile childbearing-aged women who visited endemic areas. Acute infections caused by other TORCH agents were not detected during the tested period. Determination of serological status in childbearing-aged women will help to identify seronegative women who are at risk for TORCH infections during pregnancy and help the clinicians to appropriately counsel mothers on preventive measures to avoid these infections. Presented results highlight the importance of surveillance, as well as a need for expanding of TORCH diagnostics including emerging and re- emerging viral zoonoses.

In last decades, number of emerging and re-emerging zoonoses has increased. The appearance of tick-borne encephalitis, dengue, West Nile fever, Usutu, hepatitis E and hantaviruses is recorded in Croatia. Some infections are endemic (tick-borne encephalitis) while other have been recognized in the last decades.

West Nile virus (WNV) infects wide range of vertebrates. Birds are the main reservoir of the virus which is transmitted by mosquitoes to other alimals and humals. Humans and horses are particularly sensitive to the infection, ald can develop neuroinvasive disease even with lethal outcome. We used chickens and horses as sentinels for detection of seasonal incursions of WNV. During a three-year period (2O 13-20 1 5) , a total of I , 7 17 serum samples from sentinel outdoor chickens and 8, 131 serum samples from horses were tested for WNV antibodies using IgG ELISA. In addition, 278 serum samples from humans suspected of WNV infection and 1, 778 serum samples from asymptomatic subjects were tested for WNV. 198 (11.53%) chickens, 665 (8.187o)horses as well as22 (7.9o/o) humans suspected of WNV infection and 15 (O.84%l asymptomatic subjects were found to be positive. A significant correlation in geographical distribution of high WNV seroprevalence in tested animals with human WNV infections was found. Coordinated extensive monitoring of WNV infection in poultry and horses throughout Croatia, as an early warning system, enabled timely anti-epidemic measures, primarily thorough disinsection in affected areas. This resulted in prevention of mass occurrence of human neuroinvasive WNV infections as reported in certain neighbouring countries.

Objectives: Birds are the main reservoir for West Nile virus (WNV) while mammals, particularly humans and horses, represent dead end hosts. Infected birds usually do not show symptoms, although few avian species can suffer from severe disease even with lethal outcome. Geese and ducks are particularly sensitive to the disease and develop high viremia and severe symptoms. Mortality in young geese can be as high as 60%. On the other hand, infection in chickens and turkeys usually remain subclinical but with development of high titre antibodies and very low viremia. Hence, chickens can be used as sentinels for seasonal incursions of WNV which is usually introduced by migratory birds and transmitted by mosquitoes. WNV can cause neurological disease and death in humans as well as in horses. The objective of this study was to use chickens and horses as sentinels for early detection of WNV in order to timely conduct appropriate anti-epidemic measures in Croatia. Materials and methods: During a three-year period (2013-2015), a total of 1, 717 serum samples from sentinel outdoor chickens and 8, 131 serum samples from horses were tested for WNV antibodies using IgG ELISA. In addition, 278 serum samples from humans suspected of WNV infection and 1, 778 serum samples from asymptomatic subjects were tested for WNV IgM and IgG antibodies. WNV positive human samples were confirmed using virus neutralization test. Positive horse sera were retested using IgM ELISA in order to confirm current/recent WNV infection. Results & Conclusion: 198 (11.53%) chickens, 665 (8.18%) horses as well as 22 (7.9%) humans suspected of WNV infection and 15 (0.84%) asymptomatic subjects were found to be positive. Of 585 positive horse sera, WNV IgM antibodies were found in 34 (5.81%) samples. A significant correlation in geographical distribution of high WNV seroprevalence in tested animals with human WNV infections was found. In addition, this coincided in regions contiguous to countries where high incidence of human neuroinvasive WNV infection was reported during the investigated period. Coordinated extensive monitoring of WNV infection in poultry and horses throughout Croatia, as an early warning system, enabled timely anti-epidemic measures, primarily thorough disinsection in affected areas. This resulted in prevention of mass occurrence of human neuroinvasive WNV infections as reported in certain neighbouring countries.

Objectives: Several arboviruses have emerged in Croatia in the last few years such as dengue virus (DENV), West Nile virus (WNV) and Usutu virus (USUV). In addition, antibodies to DENV and chikungunya virus (CHIKV) were sporadically reported in travelers to endemic areas. We analyzed seroprevalence of the emergent arboviruses in humans and animals in Istria County during a three-year period (2012-2014). Methods: Human serum samples were tested for DENV, WNV, USUV and CHIKV IgG antibodies. Horse and chicken serum samples were tested for WNV IgG antibodies. All tested chickens were less than one year old. Serologic tests were performed using commercial enzyme-linked immunosorbent assays. Except in chickens, all IgG reactive samples were further tested for the presence of IgM antibodies for confirmation of acute/recent infection. Results: In 2012, 2/136 (1.4%) human samples from inhabitants of Pula and Labin tested positive for CHIKV IgG antibodies while IgM antibodies were not detected. Antibodies to WNV, DENV and USUV were not found in any tested sample. Among horses, WNV IgG seropositivity was 1.8% (5/276) in 2012, 1.7% (5/289) in 2013 and 2.3% (2/87) in 2014. Acute infections were not detected during the tested period except one documented seroconversion during 2014. Chicken seropositivity to WNV was 33.3% (3/9) in 2013 and 90% (18/20) in 2014. Conclusions: Our results indicate circulation of WNV among horses and chickens in Istria County during three consecutive transmission seasons. Although USUV infections were not detected in Istria County, emergence of human USUV neuroinvasive disease in Croatia highlighted the importance of flaviviruses surveillance. In addition, current data from Europe and Croatia indicate that indigenous transmission of DENV and CHIKV is possible. Permanent vector control measures should be regularly performed, particularly in regions with established Ae. albopictus population.

Patogeni koji se mogu prenositi hranom uključuju crijevne bakterije, viruse, parazite, bakterije koje proizvode toksine i prione. Od navedenih patogena najčešći uzročnici bolesti koje se prenose hranom jesu virusi. U ovom su radu pregledno prikazani virusi koji se prenose hranom s posebnim naglaskom na načine širenja i mogućnosti profilakse kao najvažnijim područjima u kojima sudjeluje veterinarska struka. U navedenom području, kao i u javnom zdravstvu u cjelini, samo bliska suradnja veterinarske i liječničke struke može rezultirati uspostavom cjelovitog sustava nadzora i suzbijanja zaraznih bolesti s ciljem očuvanja zdravlja ljudi. Prvi korak pritom jest dostupnost najnovijih spoznaja i informacija o rizicima i mjerama zaštite širem krugu stručnjaka u svrhu sprečavanja nastanka epidemija uzrokovanih virusima koji se prenose hranom, što je i bio cilj ovog preglednog prikaza., Foodborne pathogens include bacteria, viruses, parasites, toxin producing bacteria and prions. Of these, by far the most common causes of foodborne illness are viruses. In this article a review is given of the most important food borne viruses with the emphasis on the modes of transmission and prophylaxis as the most important areas where veterinary practitioners are included. In these areas and in protecting public health in general only close collaboration between human and veterinary medical specialists will result in establishing an optimal system for surveillance and prevention of foodborne infections in humans. The aim of this article is to present new information regarding risk factors and protection measures to a wide number of professionals, as a step in the prevention of epidemics caused by foodborne viruses.

West Nile virus (WNV) is a small, enveloped, spherical virus that belongs to the family Flaviviridae, genus Flavivirus, Japanese encephalitis serocomplex. Natural reservoirs of WNV are birds, and the main vectors are mosquitoes of the genus Culex. There are seven genetic lineages of WNV. Lineages 1 and 2 are the most widely distributed (Africa, North and South America, Europe, Asia and Australia). About 80% of infections are asymptomatic. In 20% of patients nonspecific febrile disease occurs (West Nile fever). Less than 1% of infected persons will develop neuroinvasive WNV disease (meningitis, encephalitis, and poliomyelitis). In Croatia, antibodies to WNV were demonstrated in humans,bears and horses. In August-September 2012 clinical cases of human WNV neuroinvasive disease and asymptomatic acute infection in horses were reported for the first time in three eastern Croatian counties. The diagnosis was confirmed by serologic tests (enzyme immunoassay, IgG avidity, plaque-reduction neutralization test).

In 2012 the active WNV surveillance system in horses was introduced in Croatia. Between June and October 2012, in six Croatian counties, 1804 horse sera were sampled and tested for IgM WNV antibodies as the confirmation of an acute infection. Additional 1472 samples were tested for the presence of IgG WNV antibodies in the seroprevalence study. The study showed the increased seroprevalence of WNV infection in horses (was) in four and acute infections in horses in three Eastern Croatian counties. In the same transmission season, the first human WNV clinical cases were reported in the counties with a confirmed increase in WNV seroprevalence in horses. Even more, the first detection of horses acutely infected with WNV had been two weeks before the first confirmed human clinical case. The results confirmed the active serosurveillance system in horses as a valuable tool for WNV surveillance in Croatia and as the source of important veterinary medicine and public health data.

During the last two decades, several West Nile virus (WNV) outbreaks have occurred throughout the world. In Croatia, clinical cases of WNV infection have not been documented until now, although cases have occurred in the neighboring countries(Italy, Hungary, Romania and Greece). Moreover, in July 2012, the first outbreak of WNV disease occurred in Serbia with 44 laboratory diagnosed cases and six deaths. We are reporting the first outbreak of WNV neuroinvasive infection in humans and evidence of acute WNV infection in horses in Croatia (August-September 2012). WNV IgM/IgG testing and IgG avidity were performed using commercial ELISA and confirmed by plaque-reduction neutralization test (PRNT) and micro-neutralization test (micro-NT). Samples were also tested for other flaviviruses (tick-borne encephalitis virus-TBEV, dengue virus-DENV and Usutu virus-USUV) to rule out cross-reactions. Seven human cases of WNV neuroinvasive disease were laboratory confirmed between 06 and 21 September 2012 in Croatia. The median patient age was 62 years (range 48-77 years). Five patients presented with meningoencephalitis and two patients with meningoencephalitis followed by acute flaccid paralysis. Most of them had underlying diseases, the most common being hypertension. In six patients, WNV-specific IgM antibodies with low IgG avidity antibodies were detected while one patient showed only IgM antibodies. Crossreactivity was seen mainly for DENV IgG. WNV infection was confirmed by PRNT and micro-NT in all samples. Patients with WNV neuroinvasive disease are residents of three counties located in eastern part of Croatia, near the Serbian state border. The first case was imported from Serbia while the others represent the first autochthonous WNV cases. Five patients fully recovered. Before human cases were reported, acute WNV infection had been demonstrated by detection of IgM antibodies in twelve asymptomatic sentinel horses in counties where human cases occurred. In September 2012, WNV neuroinvasive disease in humans was documented for the first time in Croatia. Asymptomatic acute infection in sentinel horses had been reported in the same geographic region before human cases occurred. Adulticidal and larvicidal treatments were administered immediately in the respective place of residence. Epidemic control measures prevented further spreading of disease.

Dengue is acute viral disease transmitted to humans by Aedes mosquitoes (Ae. aegypti, Ae. albopictus). Dengue virus belongs to the family Flaviviridae, genus Flavivirus. There are four dengue virus serotypes (1-4) which are maintained endemically. The disease is endemic in tropical and subtropical areas between latitudes 35 degrees N and 35 degrees S. Infections may be asymptomatic or may produce a wide spectrum of diseases: non-specific febrile illness, dengue fever, dengue haemorrhagic fever or dengue shock syndrome. For the first time in Croatia, Ae. albopictus was registered in Zagreb in October 2004. In autumn 2005, additional records of Ae. albopictus presence were made in many places along the Adriatic coast. During 2007, two cases of imported dengue fever were reported in Croatia, after which similar imported cases appeared continually. In August 2010, the first autochthonous case of dengue fever was recorded on the peninsula Peljesac. Though Croatia is not endemic for dengue, the existence of a corresponding vector and a latent threat by imported dengue cases demand state-of-the-art and timely diagnostics. The most commonly used methods in laboratory diagnosis of dengue infections involve detection of viral RNA, antigen detection and serologic methods (detection of antibodies).

Aim: This investigation was performed at Department of Virology, Croatian National Institute of Public Health, the same diagnostic laboratory using the same serologic method as in earlier studies (in 1982 and 1992) to determine the incidence of the most common agents of atypical pneumonia in Croatia between January I and December 31, 2002. Patients and Methods: The study included 630 patients from nearly all regions of Croatia with a clinical diagnosis of atypical pneumonia based on epidemiologic data, clinical symptoms, laboratory findings and chest X-rays. Most of them were from Zagreb (n=370), followed by those from Slavonia, Istria, northwest Croatia, and Dalmatia. In all of them paired sera were collected at an interval of two weeks or more and tested for complement-fixing (CF) antibodies against the most common causative agents of the atypical pneumonia syndrome using CF test (micromethod). Results: An etiologic diagnosis was established in 25% (158/630) patients (81 male and 77 female). Respiratory viruses were the most frequently demonstrated pathogens in 2002, accounting for 72% of cases (adenoviruses 47%, parainfluenza viruses 14%, influenza viruses 9% and respiratory syncytial virus (RSV) 2%). These were followed by Chlamydophila (C.) psittaci ; 19%), Mycoplasma (M.) pneumoniae ; 6%) and Coxiella (C.) burnetii (3%). There were 3 cases of double infection: a combination of adenovirus and M. pneumoniae, of RSV and parainfluenza virus, and of RSV and M. pneumoniae in one patient each. Adenoviral, mycoplasmal and psittacosal pneumonia occurred throughout the year ; influenza and most of RSV pneumonias occurred in winter months. Parainfluenza viruses caused pneumonias throughout the year but were more common in winter months. CF test does not distinguish type-specific antibodies to parainfluenza viruses. In March, a small epidemic of psittacosis (11 patients) was registered in the Split area while was responsible for the high incidence of psittacosal pneumonia in 2002. While M. pneumoniae-caused pneumonia occurred mainly in children and adolescents, viral pneumonias were distributed across all age groups. Psittacosal pneumonia occurred in only one child but was more common in adolescents and especially adults. Q-fever pneumonia occurred only in adults.

The "atypical" pathogens are important causes of pneumonia, causing illness ranging from mild to severe and life threatening. The most important atypical pathogens are Mycoplasma pneumoniae (M. pneumoniae) and Chlamydia pneumoniae (C. pneumoniae). Others include Legionella spp., Chlamydia psittaci (C. psittaci), Coxiella burnetti (C. burnetti) and viruses (influenza, parainfluenza, adenoviruses, respiratory syncycial virus-RSV). Variations in etiology have been detected in different times of life and different geographical regions. The season of the year may also influence etiology and some infections follow a cyclical pattern. Direct diagnostics is sometimes difficult because most of these agents are intracellular and cannot be easily cultured. Thus, diagnosis relies on serology. The complement fixation (CF) test is the most widely used serologic test in diagnosis of atypical pneumonia. We performed two epidemiological studies (in 1992 and 2002) to determine the frequency of the most common agents of atypical pneumonia in Croatia in the same diagnostic laboratory using the same serologic method. The studies included 581 and 630 patients, respectively from nearly all regions of Croatia with clinical diagnosis of atypical pneumonia based on clinical symptoms, laboratory findings and chest X-rays. In all of them, paired sera were collected at an interval of two weeks or more and tested for CF-specific antibodies against the most common causative agents of the atypical pneumonia syndrome. An etiological diagnosis of pneumonia was established in 27% (156/581), respectively 25% (158/630) of the patients. M. pneumoniae was the most frequently demonstrated pathogen in Croatia in 1992 (22%). It was followed by RSV (21%), adenoviruses (14%), parainfluenza viruses (12%), C. burnetti (12%), C. psittaci (10%) and influenza viruses (9%. Most cases of atypical pneumonia in 2002 were due to adenoviruses (47%), C. psittaci (19%), parainfluenza viruses (14%), influenza viruses (9%), M. pneumoniae (6%), C. burnetti (3%) and RSV (2%). Whereas adenoviral, mycoplasmal and psittacosal pneumonias occured throughout the year, influenza and most parainfluenza and RSV pneumonias occured in winter months. In March 2002, small epidemics of psittacosis (11 patients) were registered in the Split area. Q-fever pneumonias occurred in winter and spring. It was noted that RSV pneumonias appeared only in children (except for one man older than 60 years). While M. pneumoniae-caused pneumonia occurred mainly in children and adolescents, the parainfluenza and adenoviral pneumonias were distributed in all age groups. Influenza pneumonia was distributed in all age groups in 1992, but only in adults in 2002. Psittacosis occured in only two children but was more common in adolescents and especially adults. Q-fever pneumonia occurred only in adults.