High Detection Rates of Human Bocavirus in Infants and Small Children with Lower Respiratory Tract Infection from Croatia.

BACKGROUND: Human bocavirus (HBoV) is known to cause lower respiratory tract infections (LRTI) in children and may result in substantial morbidity and mortality. The aim of this study was to determine HBoV prevalence among hospitalized infants and small children with acute LRTI in Zagreb, Croatia, as well as to evaluate HBoV DNA quantity in samples in relation to the patients' age and co-infection with other respiratory viruses. METHODS: During winter season 2016/2017, a total of 295 children younger than three years of age who were admitted to hospitals with LRTI were tested for the presence of HBoV, respiratory syncytial virus (RSV), adenovirus (ADV), parainfluenza virus (PIV) types 1 to 3, and human metapneumovirus (HMPV). HBoV was detected with a real-time PCR method, and the other viruses were diagnosed using monoclonal antibodies in direct fluorescence assay. RESULTS: Viral etiology was proven in 225/295 (76.3%) of patients. The most commonly diagnosed virus was RSV (59.3%), followed by HBoV (23.1%), PIVs (4.4%), ADV (3.1%), and HMPV (1.4%). HBoV-infected children were older than RSV-infected children; likewise, detection rates of HBoV infection increased with age, while RSV infection rates decreased with age. In 51% of HBoV-positive samples an additional respiratory virus was also detected. There was no difference in HBoV DNA quantity between samples with single virus detection and those with multiple virus detection (p = 0.056), although samples positive only for HBoV showed higher cycle threshold values. There was no difference in HBoV DNA quantity in samples of different age groups (p > 0.05). CONCLUSIONS: Frequent detection of HBoV in small children with LRTI, even in combination with other viruses, highlights its role in the pathogenesis of respiratory disease.

Variability analysis and inter-genotype comparison of human respiratory syncytial virus small hydrophobic gene.

BACKGROUND: Small hydrophobic (SH) gene is one of the mostly diverse genomic regions of human respiratory syncytial virus (HRSV). Its coding region constitutes less than 50% of the complete gene length, enabling SH gene to be highly variable and the SH protein highly conserved. In standard HRSV molecular epidemiology studies, solely sequences of the second hypervariable region of the glycoprotein gene (HVR2) are analyzed. To what extent do the strains identical in HVR2 differ elsewhere in genomes is rarely investigated. Our goal was to investigate whether diversity and inter-genotype differences observed for HVR2 are also present in the SH gene. METHODS: We sequenced 198 clinical samples collected within a limited area and time frame. In this HRSV collection, rapid and significant changes in HVR2 occurred. RESULTS: Over 20% of strains from this pool (containing HRSV genotypes NA1, ON1, GA5, BA9 and BA10) would be incorrectly assumed to be identical to another strain if only the HVR2 region was analysed. The majority of differences found in SH gene were located in the 5' untranslated region (UTR). Seven indels were detected, one was genotype GA5 specific. An in-frame deletion of 9 nucleotides (coding for amino acids 49-51) was observed in one of group A strains. Fifteen different SH protein sequences were detected; 68% of strains possessed the consensus sequence and most of others differed from the consensus in only one amino acid (only 4 strains differed in 2 amino acids). The majority of differing amino acids in group A viruses had the same identity as the corresponding amino acids in group B strains. When analysis was restricted to strains with identical HVR2 nucleotide sequences and differing SH protein sequences, 75% of differences observed in the SH ectodomain were located within region coding for amino acids 49-51. CONCLUSIONS: Basing HRSV molecular epidemiology studies solely on HVR2 largely underestimates the complexity of circulating virus populations. In strain identification, broadening of the genomic target sequence to SH gene would provide a more comprehensive insight into viral pool versatility and its evolutionary processes.

Comparative genomics of human rubulavirus 2.

Although human rubulavirus 2 (HPIV2) is an important respiratory pathogen, little is known about its molecular epidemiology. We performed a comparative analysis of the full-length genomes of fourteen HPIV2 isolates belonging to different genotypes. Additionally, evolutionary analyses (phylogenetic reconstruction, sequence identity, detection of recombination and adaptive evolution) were conducted. Our study presents a systematic comparative genetic analysis that complements prior analyses and utilizes full-length HPIV2 genomes to provide a basis for future work on the clinical significance, molecular variation and conservation, and evolution of HPIV2.

Genetic diversity of human metapneumovirus in hospitalized children with acute respiratory infections in Croatia.

Human metapneumovirus (HMPV) is recognized as a global and frequent cause of acute respiratory tract infections among people of all ages. The objectives of this study were molecular epidemiology and evolutionary analysis of HMPV strains which produced moderate and severe acute respiratory tract infections in children in Croatia during four consecutive seasons (2011-2014). A total of 117 HMPV-positive samples collected from hospitalized pediatric patients presenting with acute respiratory tract infections and tested by direct immunofluorescence assay were first analyzed by amplifying a part of the F gene. Sixteen samples were further analyzed based on complete F, G, and SH gene sequences. HMPV genome was identified in 92 of 117 samples (78%) and the circulation of multiple lineages of HMPV was confirmed. In 2011, 2012, and 2014, subgroups A2 and B2 co-circulated, while B1 gained prevalence in 2013 and 2014. The study established the presence of a novel subcluster A2c in Croatia. This subcluster has only recently been detected in East and Southeast Asia. This study provides new insights into epidemiology and genetic diversity of HMPV in this part of Europe.

Genetic Variability and Sequence Relatedness of Matrix Protein in Viruses of the Families Paramyxoviridae and Pneumoviridae.

BACKGROUND: The families Paramyxoviridae and Pneumoviridae comprise a broad spectrum of viral pathogens that affect human health. The matrix (M) protein of these viruses has a central role in their life cycle. In line with this, molecular characteristics of the M proteins from variable viruses that circulated in Croatia were investigated. METHODS: Sequences of the M proteins of human parainfluenza virus (HPIV) 1-3 within the family Paramyxoviridae, human metapneumovirus (HMPV), and human respiratory syncytial virus from the family Pneumoviridae were obtained and analyzed. RESULTS: M proteins were very diverse among HPIVs, but highly conserved within each virus. More variability was seen in nucleotide sequences of M proteins from the Pneumoviridae family. An insertion of 8 nucleotides in the 3' untranslated region in 1 HMPV M gene sequence was discovered (HR347-12). As there are no samples with such an insertion in the database, this insertion is of interest and requires further research. CONCLUSION: While we have confirmed that M proteins were conserved among individual viruses, any changes that are observed should be given attention and further researched. Of special interest is inclusion of HPIV2 M proteins in this analysis, as these proteins have not been studied to the same extent as other paramyxoviruses.

Genetic diversity among human parainfluenza virus type 2 isolated in Croatia between 2011 and 2014.

The dynamics and evolution of the human parainfluenza virus type 2 (HPIV2) in Croatia, and also globally, are largely unknown. Most HPIV2 infections are treated symptomatically outside the hospital setting. Thus, the diagnosis is missing making it difficult to follow the genetic variation and evolution of the HPIV2. This study explores hospitalized HPIV2 cases in Croatia during 4-year period (2011-2014). Most cases in this period were reported in October or November (68.75%) and most of patients were under 2 years of age (81.25%). For molecular analyses, we used the F and HN gene sequences and showed that although both regions are equally suitable for phylogenetic analyses it would be advantageous to use regions longer than 2 kb for HPIV2 analyses of isolates which are spatially and temporally closely related. We show here that the dominant cluster in this area was cluster G3 while only one strain isolated in this period was positioned in the distant cluster G1a. Further monitoring of the HPIV2 will determine whether cluster G3 will remain dominant or it will be overruled by cluster G1a. This will be important for the surveillance of virus circulation in population and significance of the viral infection. J. Med. Virol. 88:1733-1741, 2016. © 2016 Wiley Periodicals, Inc.

IgG Avidity: an Important Serologic Marker for the Diagnosis of Tick-Borne Encephalitis Virus Infection.

A total of 52 serum samples from patients with symptoms suggestive of tick-borne encephalitis virus (TBEV) infection and positive IgM and/or IgG antibodies were tested for IgG avidity. Acute/recent TBEV infection was confirmed by low/borderline avidity index (AI) in 94.8% IgM positive/IgG positive samples, while in 5.2% high AI was found indicating persisting IgM antibodies. Majority of IgM negative/IgG positive samples (78.6%) showed high AI consistent with past TBEV infection. However, in 21.3% patients without measurable IgM antibodies current/recent infection was confirmed by AI. IgG avidity represents an additional serologic marker that improves diagnosis of TBEV, especially in cases of atypical antibody response.

Early Evolution of Human Respiratory Syncytial Virus ON1 Strains: Analysis of the Diversity in the C-Terminal Hypervariable Region of Glycoprotein Gene within the First 3.5 Years since Their Detection.

OBJECTIVE: Characterization of the phylogeny and diversity of human respiratory syncytial virus (HRSV) genotype ON1 that occurred during its early evolution (within the first 3.5 years since the detection of the first ON1 strains). ON1 strains have a 72-nucleotide-long in-frame duplication within the second hypervariable domain of the glycoprotein gene (HVR2). METHODS: All available HVR2 sequences of strains belonging to the ON1 genotype published prior to June 20, 2014 were collected. Multiple sequence alignments, phylogeny, phylogeography, sequence clustering and putative protein analyses were performed. RESULTS: The worldwide spread and diversification of ON1 strains are presented. Only in a minority of ON1 strains do the two replicas remain identical, and various ON1 strains possess common differences between the first and the second copy (segments A and B). Mutations of the progenitor sequence were more frequent in segment B, a higher overall diversity on the protein level and more putative glycosylation sites exist in segment B, and, unlike in segment A, positive selection acts on that protein region. CONCLUSIONS: The fast spread of the novel HRSV genotype ON1 has been accompanied by its rapid concurrent diversification. Differences in variability of the two replicas within HVR2 were detected, with C-terminal replica being more variable.

[Usutu virus: a novel flavivirus in Croatia]. / Usutski virus: novi flavivirus u hrvatskoj.

Usutu virus (USUV) belongs to the family Flaviviridae, genus Flavivirus, Japanese encephalitis serocomplex. The virus was discovered in 1959 in South Africa and has emerged since 1996 causing epizootics with high avian mortality in Europe. The importance of USUV in humans is not fully understood. However, several human clinical cases of USUV infection described so far indicate the role of this virus as an antropozoonotic agent. In Croatia, serologic evidence of USUV was first documented in 2011 in two horses from Zagreb and Sisak-Moslavina County. In 2012, USUV neutralizing antibodies were found in one human sample from a resident of a Vukovar-Srijem County. Human clinical cases of USUV infection were detected for the first time during the West Nile virus outbreak from July to September 2013. Three patients with USUV neuroinvasive disease were detected in the City of Zagreb and Zagreb County. Our results indicate USUV circulation in Croatia. Further human cases could be expected in the next transmission seasons.

[POLIOMYELITIS ERADICATION--ONE STEP TO ACHIEVE THE GOAL]. / ERADIKACIJA POLIOMIJELITISA--KORAK DO CILJA.

Poliomyelitis is a very old disease of humans, caused by poliovirus. With appearance of the epidemics in the 20th century, poliomyelitis became a global public health issue. In 1988, the World Health Organization started a campaign for global eradication of poliomyelitis and till now poliomyelitis cases have been reduced by more than 99%. In Croatia, the introduction of vaccination in 1961 resulted in dramatic reduction of paralytic disease. The European region, including Croatia was certified polio free in 2002. However, the final goal of the "polio-free world" has not yet been reached. To reinforce the campaign, the global polio eradication initiative has come up with the Polio Eradication & Endgame Strategic Plan 2013-2018 with detailed program how to resolve the main challenges: (a) continued transmission of wild polioviruses in endemic reservoirs; (b) reinfection of polio-free areas; and (c) outbreaks due to the circulating vaccine-derived polioviruses (cVDPV). Global oral polio vaccine cessation will follow, with the introduction of universal use of inactivated polio vaccine.

[TICK-BORNE ENCEPHALITIS VIRUS: EPIDEMIOLOGICAL AND CLINICAL PICTURE, DIAGNOSIS AND PREVENTION]. / VIRUS KRPELJNOG ENCEFALITISA: EPIDEMIOLOSKA I KLINICKA SLIKA, DIJAGNOSTIKA I PREVENCIJA.

Tick-borne encephalitis virus (TBEV) is a small, enveloped virus that belongs to the family Flaviviridae, genus Flavivirus, tick-borne encephalitis serocomplex. There are three subtypes of TBEV: European, Far-Eastern and Siberian subtypes, which differ in geographical distribution, tick vector and clinical manifestation of disease in humans. TBEV is endemic in a wide geographic area ranging from Central Europe and the Scandinavian Peninsula to Japan. The virus is maintained in nature in so-called natural foci in cycles involving ticks and wild vertebrate hosts (mainly small rodents). The principal vector for the European subtype is Ixodes (I.) ricinus tick, whereas for Far-Eastern and Siberian subtypes it is I. persulcatus. In the Baltic States and Finland, co-circulation of two or all three subtypes was documented. Several animals, principally small rodents, serve as virus reservoirs. In the tick population, TBEV is transmitted by feeding/co-feed ing on the same host, transovarially (from infected females to their eggs) and trans-stadially (from one development stage to the next). An infected tick remains infected for life. While most TBE infections in humans occur following a tick bite, alimentary routes of TBEV transmission (consumption of unpasteurized milk/milk products from infected livestock) have also been described. All three tick stages can transmit the infection to humans. In the last decade, an increase of TBE incidence has been observed in some endemic areas. This could be due to a number of interacting factors such as changes in the climatic conditions affecting tick habitats, improvements in the quality of epidemiological surveillance systems and diagnostics, in landscape resources and their utilization and more outdoor recreation activity. In addition, the endemic area of TBEV has expanded to higher altitudes (up to 1500 m), apparently influenced by climatic changes. The typical clinical picture of infection with European subtype TBEV is characterized by a biphasic course (50%-77%). The first phase is characterized by nonspecific, flu-like symptoms followed by an asymptomatic interval of about one week. In 20%-30% of persons who develop symptoms, the second phase occurs with symptoms of central nervous system involvement (meningitis, encephalitis, myelitis, radiculitis). The mortality rate for European subtype is 1%-2%. Diagnosis is usually based on detection of specific antibodies (enzyme immunoassay, indirect immunofluorescent assay, plaque reduction neutralization test). From 1993 to 2013, a total of 777 cases of TBE were reported in Croatia. Endemicity is highest in north-western counties (mean incidence 3.61-6.78/100,000 inhabitants). The majority of patients were older than 20 years (88%). Most cases (73%) were reported from May to July.

[West nile virus infection: re-emergent disease in Croatia]. / Infekcija virusom zapadnog nila: re-emergentna bolest u hrvatskoj.

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).

A study of the genetic variability of human respiratory syncytial virus in Croatia, 2006-2008.

Human respiratory syncytial virus (HRSV) is a common etiological agent of acute lower respiratory tract disease in infants. The molecular epidemiology of HRSV in Croatia over four consecutive seasons (from 2006 to 2008) was investigated. A total of 72 HRSV samples were chosen from 696 screened cases in a pediatric clinic in Zagreb. Molecular characterization of HRSV revealed the predominance of HRSV group B viruses in the first two epidemic seasons and HRSV group A viruses in the next two seasons. According to the phylogenetic analysis, NA1 and BA9 were the predominant circulating HRSV genotypes detected during the study. Overall, 82.9% of all HRSV A strains belonged to the NA1 genotype. The HRSV B genotype BA9, detected in two consecutive seasons (2006 and 2007), was the predominant circulating HRSV B genotype, accounting for 80.6% of all HRSV B strains. This study provides data on the circulation pattern of HRSV genotypes in Croatia and their molecular characterization.

Analysis of biennial outbreak pattern of respiratory syncytial virus according to subtype (A and B) in the Zagreb region.

BACKGROUND: The epidemic pattern of respiratory syncytial virus (RSV) in Croatia is biennial. In order to determine if the circulation of different RSV subtypes affects the outbreak cycle, the aim of the present study was to analyze the epidemic pattern of RSV in children in Croatia (Zagreb region) over a period of 3 consecutive years. METHODS: The study group consisted of 696 inpatients, aged 0-5 years, who were hospitalized with acute respiratory tract infections caused by RSV, in Zagreb, in the period 1 January 2006-31 December 2008. The virus was identified in nasopharyngeal secretions using direct immunofluorescence. The virus subtype was determined on real-time polymerase chain reaction. RESULTS: Of 696 RSV infections identified in children, subtype A virus caused 374 infections, and subtype B, 318. Four patients had a dual RSV infection (subtypes A and B). The period of study was characterized by four epidemic waves of RSV infections: the first, smaller, in the spring of 2006; the second, larger, in December 2006/January 2007; the third in spring 2008, followed by a fourth outbreak beginning in November of 2008. The biennial virus cycles were persistent although the predominant RSV subtype in the first two epidemic waves was subtype B, and in the second two it was subtype A. CONCLUSION: Over a 3 year period of observation, the biennial RSV cycle in Croatia cannot be explained by a difference in the predominant circulating subtype of RSV. Other unknown factors account for the biennial cycle of RSV epidemics in Croatia.

Immunity to varicella-zoster virus in Croatian women of reproductive age targeted for serology testing.

OBJECTIVE: The aim of this study was to determine the immunity to varicella-zoster virus (VZV) in Croatian pregnant and non-pregnant women of reproductive age. METHODS: During 2007-2011, a total of 638 women aged 16-45 years were tested for the presence of VZV IgM and IgG antibodies using commercial enzyme-linked immunosorbent assay. Samples positive for IgG antibodies with positive or equivocal IgM antibodies were tested for IgG avidity. RESULTS: The overall IgG seroprevalence was 84.3 %. There was a significant increase in IgG seropositivity with age (OR = 1.04 for 1-year increase in age; 95 % CI 1.01-1.08). The lowest seroprevalence rate was reported in the 16-20 age groups (78.6 %), and the highest was in the 41-45 age groups (94.3 %). There was no significant difference in seroprevalence among women residing in urban and rural areas (83.6 vs. 87.0 %, OR 0.76, 95 % CI 0.43-1.34). CONCLUSIONS: The results of this study have shown that a high proportion of Croatian childbearing-aged women (15.7 %) who were referred to the laboratory for VZV serology testing are susceptible to VZV and, thus, at risk for contracting varicella during pregnancy. Serology testing of adolescent girls and adult women who do not have a documented history of varicella is encouraged with the aim of vaccinating seronegative girls and women against VZV before pregnancy. In addition, testing of pregnant women is advised to identify susceptible women and vaccinate them after delivery.

High prevalence of Bartonella henselae and Bartonella quintana antibodies in Croatian patients presenting with lymphadenopathy.

Between 2007 and 2010, a total of 268 Croatian patients with lymphadenopathy were tested for IgM/IgG antibodies to Bartonella (B.) henselae and B. quintana. Samples from 44.4% patients showed positive IgG antibodies: 35.8% to B. henselae, 6.7% to B. quintana and 1.9% to both Bartonella species. There was no difference in seropositivity between males and females (47.4% vs. 41.5%). Seroprevalence was high in all age groups (40.4-60.9%). Patients from urban and rural areas showed a similar seroprevalence rate (44.1% vs. 44.8%). Positive IgM antibodies were found in 28.3% patients varying from 17.5% and 37.5% among age groups. Most cases were reported from August to March.

The role of IgG avidity in diagnosis of cytomegalovirus infection in newborns and infants.

To evaluate the value of IgG avidity in diagnosis of congenital cytomegalovirus (CMV) infection in newborns and infants we collected serum samples from 40 infants under 12 months of age with suspected congenital CMV infection. Sera were tested for IgM, IgG and IgG avidity. For 25 of them, virus isolation and/or polymerase chain reaction (PCR) on urine specimens were performed. Thirteen (32.5%) patients showed the presence of CMV IgM antibodies, 3 (7.5%) had equivocal IgM result, and 24 (60.0%) patients had IgG antibodies only. Using IgG avidity, CMV infection (low avidity index-AI) was documented in 61.5% IgM positive and 54.2% IgM negative patients. Eight of nine (88.8%) IgM positive patients were positive either on virus isolation or PCR. In IgM negative patients, 46.6% urine cultures were positive for CMV and 66.6% were PCR positive. According to age, IgG avidity demonstrated acute/recent primary CMV infection in 58.8% patients younger than three months compared with 91.7% and 81.8% in 3-6 and 6-12 months old babies, respectively. In conclusion, IgG avidity is useful in diagnosis of CMV infection either in IgM positive or IgM negative children older than 3 months of age. In infants less than 3 months, transplacentally derived maternal IgG antibodies of high avidity influence on the IgG avidity result. In these children, CMV infection should be confirmed by direct virologic methods such as virus isolation or PCR.

[Virology diagnosis of re-emergent infections: dengue virus]. / Viroloska dijagnostika reemergentnih infekcija: virus dengue.

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).

Clinical and molecular characterization of a parechovirus type 1 outbreak in neonates in Croatia.

During July 2009 an outbreak in neonates represented with gastrointestinal and respiratory symptoms was observed at the Neonatal Postintensive Care Unit, Clinical Hospital Center, Zagreb. Human parechovirus type 1 (HPeV1) was isolated from seven patients, one of whom was asymptomatic. All but one were premature neonates with serious underlying conditions, and all recovered fully after several days. In order to characterize the HPeV1s, sequencing of the VP1/2A region was conducted on six isolates from the outbreak and four isolates detected in Croatia in 2008 and 2007. The analysis of sequence similarity showed that the nucleotide identity between the prototype strain (Harris) and HPeV1 isolated in Croatia was 76.5-77.5%. Croatian strains from 2007 and 2009 clustered together with strains from the Netherlands and Germany detected in 2003 and 2006, respectively, while strains from 2008 clustered with the strain from Finland detected in 2000. Change of the dominant strains each year may suggest antigenic variation as a result of viral response to specific immunity of the target population.

Herpes simplex virus infection in the Croatian population.

BACKGROUND: Herpes simplex virus (HSV) infections are caused by 2 types of virus, HSV-1 and HSV-2. Both viruses are endemic worldwide. There are marked variations in the seroprevalences of HSV-1 and HSV-2 in Europe. The aim of this study was to determine the seroprevalence of HSV infections in Croatia. METHODS: During a 3-y period (2008-2010), a total of 1672 patients were tested for the presence of HSV-1 and HSV-2 antibodies using an enzyme-linked immunosorbent assay. RESULTS: The overall immunoglobulin G (IgG) seroprevalence rates were 72.5% for HSV-1 and 9.9% for HSV-2. There was no significant difference in seropositivity between males and females for HSV-1 (72.0% vs 73.0%) or HSV-2 (8.9% vs 10.7%). HSV-1 seroprevalence increased from 26.4% in those aged 6 months-9 y to 89.9% in those aged 40?49 y, and remained stable thereafter, ranging from 87.4% to 91.5% (p < 0.001). HSV-2 IgG seropositivity increased progressively from 5.7% in participants aged 20-29 y to 26.5% in participants aged ≥ 60 y (p < 0.001). HSV-1 seroprevalence did not differ between participants residing in urban and rural areas (72.5% vs 72.6%). Urban place of residence was a significant factor for HSV-2 seroprevalence in univariate analysis, but after standardization for age, it was no longer significant. CONCLUSIONS: Multiple logistic regression showed that age was a significant predictor of both HSV-1 and HSV-2 seropositivity, while female gender was a significant predictor of HSV-2 seropositivity. In pregnant women, obstetric history was not a significant predictor of either HSV-1 or HSV-2 seroprevalence.