Distinct Expression Patterns of Genes Coding for Biological Response Modifiers Involved in Inflammatory Responses and Development of Fibrosis in Chronic Hepatitis C: Upregulation of SMAD-6 and MMP-8 and Downregulation of CAV-1, CTGF, CEBPB, PLG, TIMP-3, MMP-1, ITGA-1, ITGA-2 and LOX.

Background and Objectives: The aim of this study was to analyze the expression of genes on transcriptomic levels involved in inflammatory immune responses and the development of fibrosis in patients with chronic hepatitis C. Materials and Methods: Expression patterns of 84 selected genes were analyzed with real-time quantitative RT PCR arrays in the peripheral blood of treatment-naive patients with chronic hepatitis C and healthy controls. The panel included pro- and anti-fibrotic genes, genes coding for extracellular matrix (EMC) structural constituents and remodeling enzymes, cell adhesion molecules, inflammatory cytokines, chemokines and growth factors, signal transduction members of the transforming growth factor- beta (TGF-ß) superfamily, transcription factors, and genes involved in epithelial to mesenchymal transition. Results: The expression of SMAD-6 coding for a signal transduction TGF-beta superfamily member as well as MMP-8 coding for an ECM protein were significantly increased in CHC patients compared with controls. Conclusions: Chronic hepatitis C was also characterized by a significant downregulation of a set of genes including CAV-1, CTGF, TIMP-3, MMP-1, ITGA-1, LOX, ITGA-2, PLG and CEBPB encoding various biological response modifiers and transcription factors. Our results suggest that chronic hepatitis C is associated with distinct patterns of gene expression modulation in pathways associated with the regulation of immune responses and development of fibrosis.

Frequency of baseline NS5A resistance-associated substitutions in patients infected with genotype 1 of hepatitis C virus in Croatia.

The backbone of current treatment for chronic Hepatitis C virus (HCV) infection are direct-acting antivirals targeting viral nonstructural proteins (NS3, NS4A, NS5A, NS5B). To date, there are six NS5A inhibitors approved for treatment of chronic HCV infection. The presence of drug-associated resistance substitutions is mainly due to fast error-prone replication, showing differential frequency between genotypes and subtypes. The aim of this study was to determine the frequency of baseline resistance to NS5A protein inhibitors in patients with genotype 1 HCV in Croatia. Resistance-associated substitutions (RAS) were detected by Sanger sequencing of HCV NS5A region amplified from 84 patients followed by phylogenetic analysis and analysis with Geno2Pheno algorithm. The frequency of NS5A RAS was 14.3% and highly dependent on viral subtype. The overall frequency of NS5A RAS was higher in patients infected with HCV subtype 1b (24.2%) than in those infected with HCV subtype 1a (7.8%). Overall, three resistance-conferring mutations were detected (Q30R, M28T and Y93H) along with two mutations (M28V and L31I) that cause reduced susceptibility to NS5A inhibitors. Analysis of the sequences showed two distinct subtype 1a clades with RAS detected in 4.3% (1/23) clade I and 10.7% (3/28) clade II sequences. Only a few distinct NS5A RAS were detected suggesting a high degree of homogeneity of the viral population. High frequency of clinically relevant NS5A RAS in Croatia suggest that the analysis of frequency and patterns of resistance mutations in local populations and evaluation of their possible clinical impact could be beneficial.

Chlamydia trachomatis in Cervical Lymph Node of Man with Lymphogranuloma Venereum, Croatia, 20141.

We report an HIV-infected person who was treated for lymphogranuloma venereum cervical lymphadenopathy and proctitis in Croatia in 2014. Infection with a variant L2b genovar of Chlamydia trachomatis was detected in a cervical lymph node aspirate. A prolonged course of doxycycline was required to cure the infection.

Immune-escape mutations and stop-codons in HBsAg develop in a large proportion of patients with chronic HBV infection exposed to anti-HBV drugs in Europe.

BACKGROUND: HBsAg immune-escape mutations can favor HBV-transmission also in vaccinated individuals, promote immunosuppression-driven HBV-reactivation, and increase fitness of drug-resistant strains. Stop-codons can enhance HBV oncogenic-properties. Furthermore, as a consequence of the overlapping structure of HBV genome, some immune-escape mutations or stop-codons in HBsAg can derive from drug-resistance mutations in RT. This study is aimed at gaining insight in prevalence and characteristics of immune-associated escape mutations, and stop-codons in HBsAg in chronically HBV-infected patients experiencing nucleos(t)ide analogues (NA) in Europe. METHODS: This study analyzed 828 chronically HBV-infected European patients exposed to ≥ 1 NA, with detectable HBV-DNA and with an available HBsAg-sequence. The immune-associated escape mutations and the NA-induced immune-escape mutations sI195M, sI196S, and sE164D (resulting from drug-resistance mutation rtM204 V, rtM204I, and rtV173L) were retrieved from literature and examined. Mutations were defined as an aminoacid substitution with respect to a genotype A or D reference sequence. RESULTS: At least one immune-associated escape mutation was detected in 22.1% of patients with rising temporal-trend. By multivariable-analysis, genotype-D correlated with higher selection of ≥ 1 immune-associated escape mutation (OR[95%CI]:2.20[1.32-3.67], P = 0.002). In genotype-D, the presence of ≥ 1 immune-associated escape mutations was significantly higher in drug-exposed patients with drug-resistant strains than with wild-type virus (29.5% vs 20.3% P = 0.012). Result confirmed by analysing drug-naïve patients (29.5% vs 21.2%, P = 0.032). Strong correlation was observed between sP120T and rtM204I/V (P < 0.001), and their co-presence determined an increased HBV-DNA. At least one NA-induced immune-escape mutation occurred in 28.6% of patients, and their selection correlated with genotype-A (OR[95%CI]:2.03[1.32-3.10],P = 0.001). Finally, stop-codons are present in 8.4% of patients also at HBsAg-positions 172 and 182, described to enhance viral oncogenic-properties. CONCLUSIONS: Immune-escape mutations and stop-codons develop in a large fraction of NA-exposed patients from Europe. This may represent a potential threat for horizontal and vertical HBV transmission also to vaccinated persons, and fuel drug-resistance emergence.

Chemokines CXCL10, CXCL11, and CXCL13 in acute disseminated encephalomyelitis, non-polio enterovirus aseptic meningitis, and neuroborreliosis: CXCL10 as initial discriminator in diagnostic algorithm?

We investigated potential diagnostic usefulness of serum and cerebrospinal fluid (CSF) concentrations of chemokines CXCL10, CXCL11, and CXCL13 in pediatric patients with acute disseminated encephalomyelitis (ADEM) (n = 23), non-polio enterovirus aseptic meningitis (NPEV AM) (n = 20), and neuroborreliosis (NB) (n = 21) and children with acute infectious diseases with neurological symptoms but with excluded neuroinfection/neuroinflammation (controls, n = 20). CSF levels of CXCL10 and CXCL11 were higher in patients with NPEV AM than those in other children, and CXCL10 levels showed a high discriminative potential (area under the receiver operating characteristic curve, ROC, 0.982) with high specificity and sensitivity (both 95%). CSF levels of CXCL13 were higher in NB patients than those in other children; however, discriminative potential (area under ROC curve 0.814) and diagnostic properties were moderate (sensitivity 67%, specificity 97%). Data suggest usefulness of chemokine quantification as a diagnostic aid in children with suspected ADEM, NPEV AM, or NB.

Cytokines and statin therapy in chronic obstructive pulmonary disease patients.

Cytokines are biological response modifiers involved in the pathophysiology of chronic obstructive pulmonary disease (COPD). This study investigated the potential use of cytokines as disease severity biomarkers in COPD patients and the possible effect of statin therapy on cytokine expression. Possible associations between cytokines, body mass index (BMI) and smoking have also been studied. Cytokines IFN-γ, IL-2, IL-12 p70, TNF-α, TNF-ß, IL-4, IL-5, IL-6, IL-10, IL-1ß and IL-8 were measured in the plasma of 100 clinically stable COPD patients using a fluorescent bead immunoassay on a flow cytometer. When patients were grouped according to Global Initiative for Chronic Obstructive Lung Disease (GOLD) stage (A-D), no significant differences in cytokine concentrations were found (p > .05). Significantly decreased concentrations of IL-1ß, IL-2, IL-4, IL-8, IL-10, IL-12p70 and TNF-α were found in COPD patients receiving statin therapy in comparison with COPD patients not receiving statin therapy (p < .05). COPD patients with increased BMI (>25) had decreased IL-2 (p=.038), IL-8 (p = .039) and IL-10 (p = .005) concentrations compared to normal BMI (20-25) patients. Current COPD smokers had increased concentrations of IL-5 (p = .037) compared to former COPD smokers. Hierarchical cluster analysis showed several patterns of measured cytokines in serum of patients with stable COPD. Statin therapy is associated with decreased expression of selected Th1 and Th2 cytokines in COPD, and this effect could be of relevance in COPD patients with increased cardiovascular risk. Concentrations of Th1 and Th2 cytokines in plasma cannot be used as biomarkers of disease severity or progression of COPD.

Pre-vaccination prevalence of high-risk human papillomaviruses (HPV) in women from Kosovo and their related sociodemographic characteristics.

OBJECTIVES: Kosovo's current health care system does not support organized nationwide cervical cancer screening and human papillomavirus (HPV) vaccination programs. To date, no reliable data are available on cervical cancer incidence and mortality in Kosovo, or on high-risk HPV (HR-HPV) prevalence and HPV type distribution. Our aim is to determinate the pre-vaccination prevalence and distribution of HR-HPVs and to assesses the associations between sociodemographic characteristics and increased risk of HPV infection in women from Kosovo. MATERIAL AND METHODS: Detection of HR-HPV DNA in cytologically evaluated cervical smears was performed using a clinically validated Abbott RealTime High Risk HPV test, Roche Linear Array HPV Genotyping Test, HPV52 type-specific real-time PCR and an in-house GP5+/GP6+/68 PCR. RESULTS: The crude overall prevalence of any of the HR-HPVs was estimated at 13.1% (26/199; 95% confidence interval (CI): 9.1-18.5%), with HPV16 being the most common type (7/26, 26.9%), followed by HPV31 and HPV51, each detected in 4/26 (15.4%) cervical specimens, HPV18, detected in 3/26 (11.5%) specimens, HPV52 and HPV66, each detected in 2/26 (7.7%) specimens, and HPV33, HPV45, HPV56, and HPV58, each detected in a single (3.9%) specimen. Women over 40 (OR = 0.36), older than 18 at sexual debut (odds ratio (OR) = 0.28), those that had delivered at least one child (OR = 0.32), and those that had a history of pregnancy termination (OR = 0.39) were at lower risk for HPV infection. CONCLUSION: Because more than 70% of cervical precancerous lesions could have been prevented in Kosovo using nationwide HPV-based cervical cancer screening and HPV vaccination, it is of outmost importance to implement both programs in the national health care system as soon as possible.

Combined Analysis of the Prevalence of Drug-Resistant Hepatitis B Virus in Antiviral Therapy-Experienced Patients in Europe (CAPRE).

BACKGROUND: European guidelines recommend treatment of chronic hepatitis B virus infection (CHB) with the nucleos(t)ide analogs (NAs) entecavir or tenofovir. However, many European CHB patients have been exposed to other NAs, which are associated with therapy failure and resistance. The CAPRE study was performed to gain insight in prevalence and characteristics of NA resistance in Europe. METHODS: A survey was performed on genotypic resistance testing results acquired during routine monitoring of CHB patients with detectable serum hepatitis B virus DNA in European tertiary referral centers. RESULTS: Data from 1568 patients were included. The majority (73.8%) were exposed to lamivudine monotherapy. Drug-resistant strains were detected in 52.7%. The most frequently encountered primary mutation was M204V/I (48.7%), followed by A181T/V (3.8%) and N236T (2.6%). In patients exposed to entecavir (n = 102), full resistance was present in 35.3%. Independent risk factors for resistance were age, viral load, and lamivudine exposure (P < .001). CONCLUSIONS: These findings support resistance testing in cases of apparent NA therapy failure. This survey highlights the impact of exposure to lamivudine and adefovir on development of drug resistance and cross-resistance. Continued use of these NAs needs to be reconsidered at a pan-European level.

[Recent developments in serologic and molecular diagnosis of hepatitis B and C]. / Novosti u seroloskoj i molekularnoj dijagnostici hepatitisa B i C.

The 2013 Update of the Croatian Guidelines for the Diagnosis and Treatment of Viral Hepatitis summarizes recent developments in the diagnosis of hepatitis B and C. Determination of HBsAg, anti-HBc and anti-HBs is the initial step in the diagnostic workup of acute and chronic hepatitis B. Other hepatitis B serologic markers should be analyzed in the second stage of the diagnostic workup in HBsAg and/or anti-HBc positive patients. A positive anti-HBc finding should be followed by HBV DNA quantification. HBsAg quantification is complimentary to the HBV DNA quantification and is used: (i) to differentiate between inactive HBsAg carriers and active chronic HBeAg-negative hepatitis B in patients with HBV DNA < 2000 IU/mL; and (ii) for treatment monitoring in patients with chronic hepatitis B receiving pegylated interferon-alpha. Real-time PCR remains the method of choice for detection and quantification of HBV DNA. The first step in HCV testing is determination of specific antibodies via screening assays, enzyme immunoassays or point-of-care assays. All persons with positive results of anti-HCV screening assays should be additionally tested for HCV RNA or presence of HCV viral capsid antigen. Confirmatory anti-HCV assays should be used as additional assays for confirmation of reactive results obtained by screening enzyme immunoassays in HCV RNA-negative persons only. Molecular assays with identical lower limit of detection (LLOD) and lower limit of quantification are recommended for monitoring of viral kinetics during chronic hepatitis C triple therapy. HCV resistance testing to protease inhibitors is not part of the recommended diagnostic monitoring of patients receiving triple therapy. HCV subtyping is currently not recommended as part of pretreatment diagnostic algorithm due to currently insufficient evidence on its clinical usefulness. IL-28 genotype is an important predictor of SVR in patients treated with a combination of interferon-alpha and ribavirin as well as in patients with HCV genotype 1 receiving triple therapy. IL-28B genotyping is recommended as part of pretreatment diagnostic workup in patients with chronic hepatitis C and is a particularly important parameter for recommending double versus triple therapy in treatment-naïve patients with chronic hepatitis C.

[Viralhepatitis. Croatian Consensus Statement 2013]. / Virusni hepatitis. Hrvatska konsenzus konferencija 2013.

Croatian Consensus Conferences on Viral Hepatitis took place in 2005 and 2009. Considering the numerous novel concepts on the epidemiology, diagnosis and management of viral hepatitis (chronic hepatitis C genotype 1 in particular) that have emerged in the past four years, a new Croatian Consensus Conference on Viral Hepatitis was held in Zagreb on February 28, 2013. The abridged text of the Croatian Consensus Conference on Viral Hepatitis 2013 presents the new concepts on the epidemiology of viral hepatitis, serologic and molecular diagnosis of viral hepatitis, determination of the IL-28 gene promoter polymorphism, fibrosis grading, algorithm for patient diagnostic follow up, treatment of chronic hepatitis C (genotypes 1-6) and hepatitis B, treatment of special populations (children, dialysis patients, transplanted patients, individuals with HIV/HCV co-infection), and therapy side effects.

Prevalence of HIV and sexually transmitted infections and patterns of recent HIV testing among men who have sex with men in Zagreb, Croatia.

OBJECTIVE: To determine the prevalence of HIV and other sexually transmitted infections (STIs) among men who have sex with men (MSM) in Zagreb, Croatia, and assess correlates of testing for HIV in the past 12 months. METHODS: The authors carried out a bio-behavioural survey using respondent-driven sampling (RDS) from September 2010 to February 2011. Participants completed a questionnaire and were asked to provide blood, urine, oropharyngeal and rectal swabs for the detection of infections. Data were analysed using RDS Analysis Tool 6.0.1 and STATA V.8.0. RESULTS: A total of 387 MSM were recruited at the University Hospital for Infectious Diseases. The age range of recruited men was 18-57 years. HIV prevalence was 2.8% (95% CI 1.1% to 5.1%) (3.6%, unadjusted), lower than that found in the first RDS survey carried out in 2006 (4.5%, 95% 2.2% to 7.3%) (4.9%, unadjusted). The seroprevalence of herpes virus type 2 was 5.9% (6.9, unadjusted) and that of syphilis measured by Treponema pallidum haemagglutination assay was 7.6% (6.7%, unadjusted). The authors found urethral and/or rectal infections with Chlamydia trachomatis in 7.2% (8.5%, unadjusted) of men and gonoccocal in 2.7% (2.1%, unadjusted). HIV testing in the past 12 months was reported by 32.7% (38.9%, unadjusted). In the multivariate analysis, significant correlates of recent HIV testing were having more than three partners in the past 12 months and the knowledge of HIV status of a regular partner. CONCLUSIONS: The results indicate that there might have not been a progression of an HIV and STI epidemic in the past 5 years among MSM in Croatia. Prevention should expand by providing better uptake of HIV and STI testing services, thus enabling timely treatment.

The Effect of Treatment-Induced Viral Eradication on Cytokine and Growth Factor Expression in Chronic Hepatitis C.

In this study, we evaluated the effect of hepatitis C virus eradication using direct-acting antivirals (DAA) on the serum cytokine and growth factor profiles of chronic hepatitis C patients (CHC). Serum concentrations of 12 cytokines and 13 growth factors were measured in 56 patients with CHC before, during the DAA treatment and after sustained virological response using bead-based flow cytometry. Cytokine and growth factor levels were also measured in 15 healthy individuals. The majority of the selected cytokines and growth factors exhibited similar concentrations before, during and after successful DAA treatment, the exceptions being IL-10, EGF, HGF and VEGF. Significantly lower concentrations of IL-10, IL-13, IL-4, IL-4, IL-9, TNF- α and higher levels of Ang-2, HGF and SCF were observed in patients with CHC before and after DAA treatment compared with healthy individuals. Patients with severe fibrosis stages exhibited higher levels of Ang-2 and lower levels of EGF, PDGF-AA and VEGF. Furthermore, IL-4, IL-5 and SCF were characterized as potential biomarkers of DAA treatment using random forest. Additionally, logistic regression characterized EGF as a potential biomarker of severe CHC. Our results suggest inhibition of pro-inflammatory processes and promotion of liver regeneration in CHC patients during DAA treatment.

Distinct Cytokine Profiles in Severe COVID-19 and Non-Alcoholic Fatty Liver Disease.

Non-alcoholic fatty liver disease (NAFLD) is identified as a risk factor for developing severe COVID-19. While NAFLD is associated with chronic low-grade inflammation, mechanisms leading to immune system hyperactivation remain unclear. The aim of this prospective observational study is to analyze cytokine profiles in patients with severe COVID-19 and NAFLD. A total of 94 patients with severe COVID-19 were included. Upon admission, clinical and laboratory data were collected, a liver ultrasound was performed to determine the presence of steatosis, and subsequently, 51 were diagnosed with NAFLD according to the current guidelines. There were no differences in age, sex, comorbidities, and baseline disease severity between the groups. Serum cytokine concentrations were analyzed using a multiplex bead-based assay by flow cytometry. Upon admission, the NAFLD group had higher C-reactive protein, procalcitonin, alanine aminotransferase, lactate dehydrogenase, and fibrinogen. Interleukins-6, -8, and -10 and CXCL10 were significantly higher, while IFN-γ was lower in NAFLD patients. Patients with NAFLD who progressed to critical illness had higher concentrations of IL-6, -8, -10, and IFN-ß, and IL-8 and IL-10 appear to be effective prognostic biomarkers associated with time to recovery. In conclusion, NAFLD is associated with distinct cytokine profiles in COVID-19, possibly associated with disease severity and adverse outcomes.

Human papillomavirus genotype specificity of hybrid capture 2 low-risk probe cocktail.

A genotyping study of 285 Hybrid Capture 2 low-risk probe cocktail-positive specimens showed cross-reactivity with several untargeted human papillomavirus genotypes. Cross-reactivity was often clinically beneficial due to the detection of untargeted low-risk genotypes. A total of 8.4% of positive results, usually weak, were due to cross-reactivity with high-risk genotypes. Establishment of a gray zone is recommended.

Seroprevalence, risk factors, and hepatitis C virus genotypes in groups with high-risk sexual behavior in Croatia.

The seroprevalence, risk factors and genotypes of hepatitis C virus (HCV) in groups with high-risk sexual behavior (persons with multiple sexual partners, men who have sex with men, commercial sex workers and their clients and persons with sexually transmitted diseases) in seven Croatian cities were analyzed. A total of 821 participants without history of injecting drug use were included in the study. Anti-HCV prevalence among risk groups varied from 2.9% to 8.5% with an overall prevalence of 4.6% (95% CI = 3.2-6.1) compared with 0.5% (95% CI = 0.0-1.5) in controls (pregnant females; OR = 9.66; 95% CI = 1.32-70.7). HCV-RNA was detected in 73.1% anti-HCV positive patients. Three of the seronegative cases (2.1%) were also found to be HCV-RNA positive ("window period"). Genotype 1 was most commonly detected (55.6%). The most prevalent subtypes were 1a (38.9%) and 3a (38.9%). Sociodemographic characteristics (age, gender, marital status and level of education) were not associated with anti-HCV seropositivity. Among sexually transmitted disease markers, a higher seroprevalence of HCV infection was found in subjects with a history of HBV infection (10.5% vs. 3.8%, P = 0.002) and gonorrhea (13.2% vs. 4.2%, P = 0.011). No other factors reflecting risk sexual behavior such as sexual orientation, number of sexual partners and number of risk behaviors were associated with HCV seroprevalence.

High prevalence of non-B HIV-1 subtypes in seamen and their sexual partners in Croatia.

Travel influences the spread of non-B subtypes of HIV in Europe. The aim of this study was to assess whether the spread of non-B subtypes occurred in Croatia, a country with a low level HIV infection. Phylogenetic analysis of the reverse transcriptase and protease genes was undertaken in 145 Croatian patients with the following risks for HIV-infection: men who have sex with men (n = 64), heterosexual contact (n = 54), intravenous drug use (n = 21), bisexual contact (n = 4) and intravenous drug use plus heterosexual exposure (n = 2). The sample included 25 seamen and 13 steady female sexual partners of seamen. Overall, 38 (26%) of HIV-patients were infected with non-B subtypes. Non-B subtype infections were found only in patients with a possible heterosexual exposure. Non-B infections were most frequent in seamen and their regular female partners (in 27 of 38 seamen and their partners, 71%). The relative distribution of non-B subtypes was as follows: A, 8%; C, 26%; F, 3%; CRF01_AE, 8%; CRF02_AG, 34%; CRF05_DF, 5%; CRF06_cpx, 3%; CRF10_CD 8%; and subtype U, 5%. There were altogether 19 heterosexual couples, phylogenetic analysis confirmed that the paired samples were related in all cases. Non-B subtypes were found only in individuals with a heterosexual risk, mainly in seamen and their partners.

Prevalence of sexually transmitted infections among men who have sex with men in Zagreb, Croatia.

We used respondent-driven sampling among men who have sex with men (MSM) in Zagreb, Croatia in 2006 to investigate the prevalence of HIV, other sexually transmitted infections and sexual behaviours. We recruited 360 MSM. HIV infection was diagnosed in 4.5%. The seroprevalence of antibodies to viral pathogens was: herpes simplex virus type-2, 9.4%; hepatitis A, 14.2%; hepatitis C, 3.0%. Eighty percent of participants were susceptible to HBV infection (HBs antigen negative, and no antibodies to HBs and HBc antigen). Syphilis seroprevalence was 10.6%. Prevalence of Chlamydia and gonorrhoea was 9.0%, and 13.2%, respectively. Results indicate the need for interventions to diagnose, treat and prevent sexually transmitted infections among this population.

Analysis of HIV-1 diversity, primary drug resistance and transmission networks in Croatia.

Molecular epidemiology of HIV-1 infection in treatment-naive HIV-1 infected persons from Croatia was investigated. We included 403 persons, representing 92.4% of all HIV-positive individuals entering clinical care in Croatia in 2014-2017. Overall prevalence of transmitted drug resistance (TDR) was estimated at 16.4%. Resistance to nucleoside reverse transcriptase inhibitors (NRTIs), non-nucleoside RTI (NNRTIs) and protease inhibitors (PIs) was found in 11.4%, 6.7% and 2.5% of persons, respectively. Triple-class resistance was determined in 2.2% of individuals. In addition, a single case (1.0%) of resistance to integrase strand-transfer inhibitors (InSTIs) was found. Deep sequencing was performed on 48 randomly selected samples and detected additional TDR mutations in 6 cases. Phylogenetic inference showed that 347/403 sequences (86.1%) were part of transmission clusters and identified forward transmission of resistance in Croatia, even that of triple-class resistance. The largest TDR cluster of 53 persons with T215S was estimated to originate in the year 1992. Our data show a continuing need for pre-treatment HIV resistance testing in Croatia. Even though a low prevalence of resistance to InSTI was observed, surveillance of TDR to InSTI should be continued.

Seroprevalence of herpes simplex virus type 2 in adult HIV-infected patients and blood donors in Croatia.

The present study estimates herpes simplex virus type 2 (HSV-2) seroprevalence and evaluates its association with age, sex, human herpesvirus type 8 (HHV-8) and human immunodeficiency virus (HIV) among adults in Croatia. A cross-sectional survey included 166 HIV-infected patients and 219 blood donors. Antibodies against HSV-2 were determined by enzyme immunoassays based on gG2 recombinant glycoprotein. HSV-2 seroprevalence was 45.8% in HIV-infected patients and 8.7% in blood donors (p < 0.0001; OR 8.8; 95% CI 5.05-15.49). Independent predictors of HSV-2 seropositivity were HIV infection (OR 11.0; 95% CI 5.93-20.41), female gender (OR 2.28; 95% CI 1.22-4.26), older age (OR 3.93; 95% CI 2.74-7.11), and HHV-8 seropositivity (OR 2.72; 95% CI 1.09-6.75). Understanding the epidemiology of HSV-2 is a critical first step in designing interventions to decrease HSV-2 and HIV transmission. The association of HSV-2 with HIV infection and HHV-8 antibodies suggests a similar transmission route.

Follicular fluid vascular endothelial growth factor is associated with type of infertility and interferon alpha correlates with endometrial thickness in natural cycle in vitro fertilization.

The aim of this study was to analyse the presence of vascular endothelial growth factor (VEGF) and interferon alpha (IFN-α) in the follicular fluid (FF) and their possible influence, as pro-angiogenic or anti-angiogenic factors, on in vitro fertilization outcome. The concentrations of VEGF and IFN-α were correlated with oocyte and embryo quality, concentrations of hormones in the serum, perifollicular blood flow and endometrial thickness. VEGF was detected in all FF samples (median 706.6 pg/ml, range 182.9-6638 pg/ml). IFN-α was detected in 60% of the samples (median 6.5 pg/ml, range 0-79.4 pg/ml), while in 40% of the samples its levels were below the test detection limit. VEGF and IFN-α concentrations did not correlate with the cause of infertility, concentrations of FSH, LH, E2 and prolactin, oocyte or embryo quality. Significantly higher concentrations of VEGF have been found in women with primary compared with secondary infertility (p = 0.011, Mann Whitney test). The concentrations of VEGF and IFN-α did not correlate with the resistance index (RI) on days of hCG administration, follicular aspiration and embryo transfer. However, the concentrations of IFN-α correlated with endometrial thickness on the day of embryo transfer (Spearman correlation coefficient ρ = 0.4107; P < 0.05) but not on days of hCG administration and follicular aspiration. The mechanism of VEGF association with the previous ability of having a child needs to be clarified in future studies. The results of this study indicate a possible role of IFN-α in pathways of endometrial remodelling.