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Norovirus (NoV) is the leading cause of acute gastroenteritis (AGE) in many countries that have introduced universal rotavirus mass vaccination. This is the first study to report data on NoV strains in Estonia. We recruited 2249 children aged 0-18 years hospitalized for AGE in Estonian hospitals from 01 February 2015 to 31 August 2016. Norovirus gastroenteritis (NoVGE) was diagnosed in 14.5% (n=325) cases. Stool sample for RNA extraction and genotyping was available in 86% (n=280) of NoVGE cases (2015, n= 91; 2016, n=189). Dominant capsid types detected in 75% (n=210) samples were, GII.4 (63.8%, n=134), GII.3 (15.2%, n=32), GII.17 (6.7% n=14) and GII.6 (5.2% n=11). Prevailing RNA polymerase types found in 77% (n= 215) samples were GII.P31 (51.1%, n=110), GII.P21 (17.7%, n=38), GII.P4 (11.2%, n=24) and GII.P7 (6.5%, n=14). Both regions were typeable for 67% (n=189) of samples. Most prevalent strains were GII.4Sydney_2012[P31] (48.7%, n=92), GII.3[P21] (15.3%, n=29), GII.4Sydney_2012[P4] (5.8%, n=11) and GII.17[P17] (5.8%, n=11). Simpson's diversity index showed a significant difference between the age groups 1-4 and 5-9 years: D 0.64 (95% CI 0.55 to 0.73) versus 0.83 (95% CI 0.81 to 0.86), respectively (p=0.03). An accurate understanding of NoV strain diversity is important for control and preventive measures, especially in the post-rotavirus vaccine era. This article is protected by copyright. All rights reserved.

Background Estonia has a typical Eastern European HIV epidemic where the most frequent co-infection is chronic hepatitis C (HCV). We aimed to describe the changes in HCV prevalence, the distribution of HCV genotypes (GT), and HCV treatment in Estonian people living with HIV over 15 years. Methods We used data of subjects included to the Estonian HIV Cohort Study (E-HIV) before 31st of December 2015. We compared two time periods—first, 1st of January 2000 to 31st of December 2008 when the HIV epidemic was mostly spreading among people who inject drugs (PWID) and second, 1st of January 2009 to 31st of December 2015 when HIV started to emerge to the general population. Results Of 4422 HIV positives 3708 (84%) had information about their HCV serostatus; 2706 (61%) were HCV seropositive, of latter 1625 (60%) were HCV RNA positive, 239 (9%) had their HCV GT determined, and 141 (5%) received treatment for HCV. The dominating subtypes were 1b (42%) and 3a (37%) followed by 1a (16%), and the few cases of 2 (1.5%). HCV prevalence was 1.5 times (95% CI 1.4–1.6) higher in subjects diagnosed with HIV in first as compared to those diagnosed in second period (84% vs 56%, respectively). There were more men and the median age at HIV diagnosis was lower in HIV/HCV co-infected than in HIV mono-infected patients (70% vs 47% and 24 years vs. 30 years, respectively; both p Conclusion There is a decrease in HCV prevalence but it remains high among HIV positive PWID, suggesting that there is need for improvement of harm reduction programs among PWID.

The integrase strand transfer inhibitor (INSTI) dolutegravir is commonly used in combination antiretroviral therapy regimens and retains strong potency even with primary resistance mutations to some other INSTIs. Acquisition of accessory mutations to primary mutations results in significant increases in dolutegravir resistance. Previously, we reported that addition of the secondary mutation T97A can result in rapid treatment failure in individuals with INSTI mutations at positions 140 and 148. Here, we conducted a detailed case study of one of these individuals and find that T97A-containing HIV emerged from a large replicating population from only a few (≤4) viral lineages. When combined with primary INSTI resistance mutations, T97A provides a strong selective advantage; the finding that T97A-containing variants spread by replication and recombination, and persisted for months after discontinuing dolutegravir, has important implications as dolutegravir is rolled out worldwide.

Background Estonia implemented the rotavirus (RV) vaccine into its national immunization program in July 2014. We aimed to determine circulating RV genotypes and the clinical profile by genotypes from February 1, 2015, to August 30, 2016, among children 0-18 years hospitalized due to rotavirus gastroenteritis (RVGE). Methods During an observational study in 7 Estonian hospitals, we determined the RV genotypes in stool samples of RVGE patients who met predetermined criteria. Shannon's diversity index (H´) and Simpson's index (D) was used to evaluate genotype diversity by season and age and to compare prevaccine period data (2007-2008) for children 0-4 years of age (n = 77) to corresponding data from the postvaccine period (2015-2016, n = 346). The Vesikari Clinical Severity Scoring System was used for clinical profile evaluation. Results Stool samples of 479 RVGE patients were genotyped. Seventy-seven percent of RVGE infections were caused by G4P[8] (n = 150, 31%), G1P[8] (n = 100, 21%), G9P[8] (n = 79, 16%), G2P[4] (n = 23, 5%), G4P[4] (n = 17, 4%). The prevailing genotypes varied seasonally. Diversity increased during the postvaccine period among age groups 0-4: H´1.42 (95% CI: 1.2-1.7) in the prevaccine era versus 1.8 (95% CI: 1.7-2) in the postvaccine era (P = 0.008), and D 0.6 (95% CI: 0.5-0.7) versus 0.78 (0.75-0.80) (P = 0.01), respectively. The off-season period presented higher diversity compared with in-seasons. G2P[8], G1P[8], G4P[4], G9P[8], and G8P[8] presented with a different clinical profile compared with others. Conclusion Since the introduction of universal mass vaccination in Estonia, the circulating RV genotypes have changed compared with those reported in the prevaccine era. Our study adds to knowledge about RV genotype distribution in Europe and expected dynamics after RV universal mass vaccination and provides insight on the clinical profile of prevailing genotypes.

Tuberculosis associated immune reconstitution inflammatory syndrome (TB-IRIS) is a serious complication of starting combination antiretroviral therapy (cART). TB-IRIS emerges early after cART initiation and is characterized by rapid expansions of TB-specific CD4+ T cells and high levels of inflammatory mediators driven by CD4+ T cells. The effects of TB-IRIS on HIV populations are unknown, but could result in profound expansion and elimination of HIV infected cells via cellular activation and acute inflammation. We investigated immediate and long-term effects of TB-IRIS on HIV infected cells with and without TB-IRIS. We measured plasma HIV RNA, cell-associated HIV RNA and HIV DNA levels and compared genetic characteristics of HIV populations after prolonged cART. We found that TB-IRIS was associated with more diverse HIV DNA populations and HIV reservoirs after IRIS were distinct from pre-therapy populations, suggesting that TB-IRIS can shape the HIV reservoir with detrimental implications for HIV remission strategies.

Background Estonia has a typical Eastern European HIV epidemic where the most frequent co-infection is chronic hepatitis C (HCV). We aimed to describe the changes in HCV prevalence, the distribution of HCV genotypes (GT), and HCV treatment in Estonian people living with HIV over 15 years. Methods We used data of subjects included to the Estonian HIV Cohort Study (E-HIV) before 31 st of December 2015. We compared two time periods – first, 1 st of January 2000 - 31 st of December 2008 when HIV epidemic was mostly spreading among people who inject drugs (PWID) and second, 1 st of January 2009 to 31 st of December 2015 when HIV started to emerge to general population. Results Of 4422 HIV positives 3708 (84%) had information about their HCV serostatus; 2706 (61%) were HCV seropositive, of latter 1625 (60%) were HCV RNA positive, 239 (9%) had their HCV GT determined, and 141 (5%) received treatment for HCV. HCV prevalence was 1.5 times (95% CI 1.4-1.6) higher in subjects diagnosed with HIV in first as compared to those diagnosed in second period (84% vs 56%, respectively). There were more men and the median age at HIV diagnosis was lower in HIV/HCV co-infected than in HIV mono-infected patients (70% vs 47% and 24 years vs. 30 years, respectively; both p

We investigated the presence of a single-nucleotide polymorphism designated rs12979860 in the interferon λ4 (IFNλ4) gene among 345 people who inject drugs (PWID) and 495 blood donors to evaluate associations between the rs12979860 genotypes and human immunodeficiency virus/hepatitis C virus (HIV/HCV). The rs12979860 TT genotype was over-represented among HIV+ PWID than HIV- PWID and blood donors (16% vs 8% and 10%, P = 0.03, respectively). PWID with TT genotype had approximately twice the probability of being HIV+ (odds ratio [OR], 2.19; 95% confidence interval [CI], 1.11 to 4.33) than PWID without TT. Every additional year of intravenous drug use (IVDU) decreased the OR 1.16 times (OR, 0.86; 95% CI, 0.75 to 0.98). This suggests that rs12979860 TT increases susceptibility to HIV and this impact decreases with increasing duration of IVDU.

Estonia implemented rotavirus universal mass vaccination (RV UMV) in July 2014. We aimed to describe changes in acute gastroenteritis (AGE) hospitalization during RV seasons before (2007-2013) and after (2015-2018) RV UMV and compare patient profile of hospitalized AGE patients aged 0-18 years during first two consecutive RV seasons 2015 vs 2016.We described AGE hospitalization patterns pre-and post-vaccine era using Estonian Health Insurance Fund (HIF) database. During a two-year observational multicenter study in seven Estonian hospitals from 01st of February 2015 to 30th August 2016 we assessed patient profile of all patients who met pre-determined AGE criteria.In post-vaccine era AGE hospitalization rate decreased from 10 to 8 per 1000 population (RR 0.81, 95% CI 0.79-0.83) compared to pre-vaccine era. Decreased RV seasonal activity, 81% (95% CI 77-84) and 55% (95% CI 52-58) reduction of rotavirus gastroenteritis (RVGE) hospitalization among age groups1 and 1-4, respectively and upsurge of norovirus gastroenteritis (NoVGE) hospitalizations (RR = 1.8; 95% CI 1.6-1.9) was seen. In the multicenter observational study, among 2249 AGE patients hospitalized median age of RVGE patients increased from 2 to 3 years (p 0.01) and duration of hospital stay decreased among RVGE, NoVGE and other GE patients during two consecutive RV seasons. According to Vesikari Clinical Severity Scoring System statistically significant change of severity score distribution in two RV seasons was seen (p 0.001) with trend towards less severe AGE hospitalizations; 82.5% vs 70.5% severe cases in 2015 vs 2016, respectively.RV UMV lead to immediate and sustainable reduction of hospitalizations due to RVGE in children aged4 years and reduction of overall AGE accompanied with the decrease in the severity of hospitalized children.

HIV-1 drug resistance mutations (DRMs) and substitutions were assessed after the failure of the first line non-nucleoside reverse transcriptase inhibitors (NNRTIs) + 2 nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs) treatment regimens (efavirenz [EFV] + lamivudine[3TC] + zidovudine [ZDV] vs. EFV + 3TC + ddI) among the HIV-1 CRF06_cpx infected subjects in Estonia. HIV-1 genomic RNA was sequenced; DRMs and amino acid substitutions were compared in 44 treatment naive and 45 first-line NNRTI + 2 NRTI treatment failed patients consisting of EFV + 3TC + ZDV (n = 17) and EFV + 3TC + didanosine[ddI] (n = 21) therapy failed sub-populations. At least one DRM was found in 78% of treatment experienced patients. The most common NRTI mutations were M184V (80%), L74V (31%), L74I (17%), K219E (9%), and M184I (9%), NNRTI mutations were K103N (83%), P225H (14%), L100I (11%), and Y188L (11%), reflecting generally the similar pattern of DRMs to that seen in treatment failed subtype B viruses. Sub-population analysis revealed that EFV + 3TC + ddI failed patients had more DRMs compared to EFV + 3TC + ZDV failed patients, especially the ddI DRM L74IV and several additional NNRTI DRMs. Additionally, CRF06_cpx specific mutation E179V and substitutions R32K, K122E, and V200AE were also detected in treatment experienced population. After the failure of the first-line EFV + 3TC + ddI therapy HIV-1 CRF06_cpx viruses develop additional NRTI and NNRTI mutations compared to EFV + 3TC + ZDV regimen. Therefore the usage of EFV + 3TC + ddI in this subtype decreases the options for next regimens containing abacavir, and NNRTI class agents.

Wohlfahrtiimonas chitiniclastica is a rare human pathogen mostly associated with parasitic flies. This is the report on W. chitiniclastica infection in the soft tissue and bone at a 58°N latitude in the northern temperate climate zone. The importance of correct identification of clinically relevant bacteria is highlighted.

BackgroundWe aimed to determine the genetic relatedness between Staphylococcus epidermidis colonizing breast milk (BM) and BM-fed neonates during the first month of life.MethodsS. epidermidis was isolated from the stool and skin swabs of 20 healthy term and 49 preterm neonates hospitalized in the neonatal intensive care unit and from the BM of mothers once a week and typed by multilocus variable-number tandem-repeat analysis. Virulence-related genes were determined by PCR.ResultsThe gut (95%) and skin (100%) of term neonates were colonized with strains genetically similar to those in BM and carrying mecA and IS256 at low rate (both6.7%). In preterm neonates, colonization with strains genetically similar to those in BM was low on the skin (34.7%) and in the gut in the first week of life (14.3%), but the prevalence of mecA (90.6%) and IS256 (61.7%) was high. By the fourth week, in the gut of preterm neonates the prevalence of mecA (73.8%) and IS256 (18.4%) decreased, but colonization with strains genetically similar to those in BM increased (83.7%).ConclusionDuring early life, the skin and gut of preterm neonates is colonized with S. epidermidis that is distinct from strains found in BM, but gradually the gut is enriched with strains genetically similar to those in BM, as in term neonates.

Background Coagulase-negative staphylococci (CoNS) are the first colonizers of gastrointestinal tract (GIT) and the commonest cause of late-onset sepsis (LOS) in preterm neonates. Intravascular catheters are considered a major source of CoNS bacteremia. However, several cases of LOS remain without an identified source. To elucidate whether GIT could be a potential source of invasive strains, we aimed to assess the molecular similarity between CoNS from blood and GIT in preterm neonates with LOS. Methods Altogether 22 blood and 53 GIT isolates collected from 22 neonates with LOS caused by CoNS (Staphylococcus haemolyticus in 13, Staphylococcus epidermidis in 7 and Staphylococcus hominis in 2 patients) were included. Rectal swabs were collected twice weekly from birth, but only isolates obtained before LOS were analyzed. S. epidermidis isolates were typed by multilocus variable number of tandem repeats analysis and multilocus sequence typing, S. haemolyticus by pulsed-field gel electrophoresis. Results Eighteen of 22 neonates had the same CoNS species in GIT and bloodstream; all these isolates from them (altogether 18 blood and 28 GIT isolates) underwent typing. The genotypic similarity between bloodstream and ≥1 antecedent GIT isolates was observed in 13 of 18 patients-3 of 7 with S. epidermidis and 10 of 11 with S. haemolyticus infection. The concordant GIT isolates were collected 0-7 days before the positive blood culture. Conclusions The similarity between CoNS from GIT and bloodstream indicates that preterm neonates harbour invasive strains in GIT before LOS. Whether there is a causal relationship between GIT colonization and LOS remains to be elucidated in further studies.

We aimed to determine the rate of GBV-C viremia, seropositivity, and genotypes among people who inject drugs (PWID) and healthy volunteers in Estonia and to evaluate associations between GBV-C and sociodemographic factors, intravenous drug use, co-infections. The study included 345 Caucasian PWID and 118 healthy volunteers. The presence of GBV-C RNA (viremia) was determined by reverse transcriptase-nested PCR in 5' long terminal repeat. PCR products were sequenced and genotyped by phylogenetic analysis. GBV-C seropositivity was determined by ELISA. One third of PWID (114/345) and 6% (7/118) of healthy volunteers (OR = 7.8, 95% CI = 3.5-20.5, P < 0.001) were GBV-C viremic. In PWID group, 79% of sequences belonged to subtype 2a, 19% to subtype 2b, and two remained unclassified. In healthy volunteers, six out of seven sequences belonged to subtype 2a and one to subtype 2b. We found HIV+ PWID to have two times increased odds of being GBV-C viremic compared to HIV- PWID (62% vs. 38%; OR = 2.13, 95% CI = 1.34-3.36, P = 0.001). In addition, odds of being GBV-C viremic decreased with increasing age (OR = 0.94, 95% CI = 0.90-0.98, P = 0.001). HIV positivity remained associated with GBV-C viremia in multivariate analysis after adjustment for age (OR = 2.23, 95% CI = 1.39-3.58, P = 0.001). GBV-C seropositivity was similar among PWID and healthy volunteers (2.3% vs. 1.7%, respectively; OR = 1.4, 95% CI =0.3-13.5, P = 1). In an Eastern European country we demonstrated that GBV-C viremia is common among PWID, but uncommon among healthy volunteers, and GBV-C seropositivity is infrequent among both groups. Similarly to other European countries and USA, GBV-C 2a is the most common genotype in Estonia. J. Med. Virol. 89:632-638, 2017. © 2016 Wiley Periodicals, Inc.

Late-onset sepsis (LOS) in preterm neonates is increasingly reported to be associated with gut-colonizing Staphylococcus epidermidis. We aimed to describe the molecular epidemiology of S. epidermidis colonizing the gut of neonates hospitalized in two neonatal intensive care units. S. epidermidis from rectal swabs were typed by multilocus variable-number tandem-repeat analysis (MLVA), randomly chosen isolates of predominant MLVA types additionally by multilocus sequence typing. Antimicrobial susceptibility, the presence of icaA, IS256, arginine catabolic mobile element (ACME), agr type, and SCCmec type were determined. Of 276 neonates (38.4%), 106 were colonized with S. epidermidis, yielding a total of 139 isolates (62 in one unit and 77 in another unit). Of the 55 MLVA types identified, the five predominant detected in both units corresponded to sequence type (ST) 2, ST5, and ST59 or its single locus variant ST81 and formed three major MLVA clonal complexes accounting for 74.8% of all isolates. Overall, the prevalence of mecA, icaA, IS256, and ACME was 91.4%, 28.1%, 64%, and 77%, respectively. Of the mecA-positive isolates (n = 127), 43.9% carried SCCmec type IV. Of eight episodes of LOS, four were caused by ST2 and two by ST5. Preventing gut colonization with nosocomial epidemic S. epidermidis in hospitalized neonates could contribute to the prevention of LOS.

In Europe, human T-lymphotropic virus (HTLV) type 2 mainly occurs among intravenous drug users (IDUs) with prevalence up to 15% and HTLV-1 among general population with prevalence1%. However, there is no data regarding the prevalence of HTLV-1 or HTLV-2 in Eastern European IDUs population where HIV prevalence is relatively high. We aimed to determine the prevalence and genotypes of HTLV-1/HTLV-2 among IDUs and healthy volunteers in Estonia.The study included 345 IDUs and 138 healthy volunteers. The presence of HTLV-1/HTLV-2 was determined by nested PCR; positive and negative controls were used in every PCR run.The analysed IDUs resembled the IDUs of HIV epidemic in Estonia: mainly male (79%) with median age of 30years (interquartile range [IQR] 25-34), and prolonged duration of intravenous drug usage (11years; IQR 7-14). The prevalence exposure to blood-borne viral infections was high - 50% were HIV positive, 88% hepatitis C positive, 67% hepatitis B positive. Of IDUs, 64% reported receptive needle sharing in the past and 18% at least once a month during last six months. None of the IDUs carried HTLV-1 but there was a case of HTLV-2 (prevalence 0.3%; 95% CI 0.1-1.6). All healthy volunteers were HTLV-1 and HTLV-2 PCR negative.This is the first study investigating the prevalence of HTLV-1/HTLV-2 among high risk population and healthy volunteers in Eastern European region. Our results suggest that despite other widely spread blood-borne infections (e.g. HIV, HBV, HCV) HTLV-1 and HTLV-2 are rare among IDUs in Estonia.

Group A rotaviruses (RVA) are a major cause of acute gastroenteritis in children ≤ 5 y worldwide which could be prevented with two recently introduced vaccines - monovalent Rotarix (live-attenuated G1P[8] strain) and pentavalent RotaTeq (human-bovine reassortant containing serotypes G1, G2, G3, G4 and P[8]). Prior to implementation of vaccines into national immunization program we aimed to describe RVA genotype distribution in hospitalized children aged5 y in Estonia during 2007-2008. A total of 671 children with confirmed RVA gastroenteritis from three major pediatric hospitals were prospectively enrolled. G- and P-genotypes were detected from 124 stool samples by semi-nested reverse transcription-PCR. Severity of disease was assessed using Clark scoring system. The majority of cases (65%) occurred in infants aged 7 to 24 mo and were of moderate severity (mean Clark score 12.1 (SD 3.2)). The prevailing strain was G2P[4] (34.7%), causing significantly more cases than G4P[8] (12.9%), G1P[8] or G9P[8] (both 4.0%), G3P[8] (1.6%). Yearly differences in genotype distribution occurred, as G2P[4] (52.8%) dominated in 2007, but G4P[8] (26.9%) in 2008. One third of strains remained non-typeable. The distribution of RVA genotypes in Estonia differs from that seen in other Central and Eastern European countries, although one should bear in mind the large proportion of P-untypeable strains and natural fluctuations of dominating RVA genotypes. Nevertheless, considering the high genotype-independent efficacy of the vaccines, introduction of national immunization should be considered.

HIV-1 drug resistance mutations (DRMs) and substitutions were assessed after the failure of the first line non-nucleoside reverse transcriptase inhibitors (NNRTIs) + 2 nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs) treatment regimens (efavirenz [EFV] + lamivudine[3TC] + zidovudine [ZDV] vs. EFV + 3TC + ddI) among the HIV-1 CRF06_cpx infected subjects in Estonia. HIV-1 genomic RNA was sequenced; DRMs and amino acid substitutions were compared in 44 treatment naïve and 45 first-line NNRTI + 2 NRTI treatment failed patients consisting of EFV + 3TC + ZDV (n = 17) and EFV + 3TC + didanosine[ddI] (n = 21) therapy failed sub-populations. At least one DRM was found in 78% of treatment experienced patients. The most common NRTI mutations were M184V (80%), L74V (31%), L74I (17%), K219E (9%), and M184I (9%), NNRTI mutations were K103N (83%), P225H (14%), L100I (11%), and Y188L (11%), reflecting generally the similar pattern of DRMs to that seen in treatment failed subtype B viruses. Sub-population analysis revealed that EFV + 3TC + ddI failed patients had more DRMs compared to EFV + 3TC + ZDV failed patients, especially the ddI DRM L74IV and several additional NNRTI DRMs. Additionally, CRF06_cpx specific mutation E179V and substitutions R32K, K122E, and V200AE were also detected in treatment experienced population. After the failure of the first-line EFV + 3TC + ddI therapy HIV-1 CRF06_cpx viruses develop additional NRTI and NNRTI mutations compared to EFV + 3TC + ZDV regimen. Therefore the usage of EFV + 3TC + ddI in this subtype decreases the options for next regimens containing abacavir, and NNRTI class agents.

Some individuals remain uninfected despite repeated exposure to HIV. This protection against HIV has been partly associated with altered T cell subset distributions and CCR5 expression levels. However, the majority of studies have been conducted in sexually exposed subjects. We aimed to assess whether HIV infection and intravenous drug use were associated with differences in CCR5 expression, immune activation on the CD4+ and CD8+ T cells and T cell distribution among Caucasian persons who inject drugs (PWIDs). Analyses of the data from 41 HIV-positive PWIDs, 47 HIV-exposed seronegative PWIDs (ESNs) and 47 age- and gender-matched HIV-negative non-drug users are presented. Of all of the study subjects, 111 (82 %) were male, and the median age was 29 years. T cell phenotyping was performed in peripheral blood mononuclear cells with multicolour flow cytometry using anti-CD3, CD4, CD8, CD45RA, CD45RO, HLA-DR and CCR5 antibodies. The ESNs exhibited greater levels of immune activation and higher percentages of CD4+ CD45RA+RO+ and CD8+ CD45RA+RO+ cells compared to the controls but not the HIV-positive people. The CCR5 expression on the CD4+ T cell subsets in the ESNs was lower than that in the controls but similar to that the HIV positives. The percentages of CCR5+ T cells were similar in all study groups and in most of the studied cell populations. Intravenous drug use was similarly associated with differences in T cell subset distributions and CCR5 expression among both the HIV-positive and HIV-negative PWIDs compared with the controls.

Estonia is experiencing the new Eastern Europe human immunodeficiency virus (HIV) epidemic, with the highest incidence of new infections in the EU. We describe demographic changes, HIV-related laboratory parameters and co-infections during the concentrated HIV epidemic using the Estonian HIV Cohort Study (E-HIV) database, founded in 2009.All 3750 subjects in the E-HIV database on December 31, 2013 were included. Subjects were divided into risk groups: people who inject drugs (PWIDs), sexual transmission (heterosexual/homosexual), and other (perinatal) or unknown risk group. Subjects diagnosed before 2009 (first period) and after (second period) were analyzed separately.The mean age at diagnosis has increased from 22.8 years (interquartile range (IQR) = 19.5-27.2) to 29.7 years (IQR = 25.3-36.2) (p0.001) between the first and second periods. PWIDs were younger than other transmission groups (23.2 vs 27.1; p0.001). There is a statistical difference in the route of transmission among genders, with overall increasing sexual transmission. The most common AIDS-defining illness was tuberculosis (0.5%). HIV/hepatitis C (HCV) co-infection was diagnosed in 42% of cases. The population median CD4 + cell count at diagnosis has declined over the years; in total 53% have been late presenters. Half of the patients are receiving antiretroviral treatment (cART). The most common combinations are nucleoside reverse transcriptase inhibitor (NRTI) backbone plus protease inhibitors (PIs) (57%) or NRTI backbone + non-NRTIs (42%).The E-HIV enables us to fill the gap in the lack of data on the course of the new Eastern European HIV epidemic. These data demonstrate that the HIV epidemic in Estonia is moving from PWIDs to the general population, suggesting that prevention measures and testing guidelines should be revised.

The aim of this study was to compare the prevalence of different virulence factor (VF) genes in extended-spectrum beta-lactamase (ESBL) producingEscherichia colistrains isolated from the Baltic Sea region. A total of 432 strains of phenotypically ESBL positiveE. coliwere collected from 20 institutions located in Estonia, Latvia, Lithuania, and the region of St. Petersburg in Russia from January to May 2012 and analyzed for phylogenetic group and prevalence of 23 VF genes. The strains were collected from clinical material (urine, blood, wound, and respiratory tract). Bacterial isolates were compared according to phylogenetic group, clinical material, and geographical origin. Most of the VF genes were concentrated within phylogenetic group B2 and/or D. When comparing strains isolated from different countries, it was found that strains originating from Estonia and Latvia belonged mainly to group B2 and strains from Lithuania and Russia mainly to groups B2 and D. The P-fimbrial adhesin genepapEFwas more prevalent in Russian strains, colicin genecvaCin Lithuanian strains, and capsular genekpsMTIIin Latvian strains; serum resistant genetraTwas less prevalent in Estonian strains. The regional differences of VF genes remained statistically significant after taking into account the phylogenetic distribution in the countries.

TLR3 recognizes dsRNA and triggers immune responses against RNA and DNA viruses. A polymorphism in TLR3, rs3775291 (Leu412Phe), has been associated with the increased susceptibility to enteroviral myocarditis, protection against tick-borne encephalitis virus and HIV-1 infection. We investigated Caucasian intravenous drug users (IDUs) and blood donors in order to evaluate the associations between TLR3 genotypes and susceptibility to HIV infection.A total of 345 Caucasian IDUs were recruited, 50% of them were HIV positive, 89% HCV and 77% HBV positive. Based on their history of needle sharing, 20 of the HIV negative IDUs were classified as highly exposed HIV seronegatives (HESNs), 68 as non-HESNs and 85 as unexposed. The control group consisting of 497 blood donors tested negative for all three viruses. TLR3 rs3775291 were determined by using TaqMan Allelic Discrimination Assay.The TLR3 rs3775291 T allele frequency was similar among the HIV negative and HIV positive IDUs and blood donors - 36%, 31% and 34%, respectively. The frequency of persons possessing at least one TLR3 rs3775291 T allele was significantly higher in HESNs compared with blood donors and HIV positive IDUs (80% vs. 55%; p=0.037 and 80% vs. 53%; p=0.031, respectively). In the univariate analysis, persons who possessed at least one T allele had reduced odds of being HIV seropositive (OR=0.29, 95% CI=0.09-0.90). This association remained significant (OR=0.25, 95% CI=0.07-0.87) after the adjustment for other co-variates (HCV, HBV serostatus and duration of intravenous drug use).The TLR3 rs3775291 T allele has a protective effect against HIV infection among HESNs IDUs.

BACKGROUND Up to 90% HIV-1 positive intravenous drug users (IDUs) are co-infected with HCV. Although best recognized for its function as a major co-receptor for cell entry of HIV, CC chemokine receptor 5 (CCR5) has also been implicated in the pathogenesis of HCV infection. Here, we investigated whether CCR5 haplotypes influence HIV-1 and HCV seropositivity among 373 Caucasian IDUs from Estonia. METHODS Of these IDUs, 56% and 44% were HIV and HCV seropositive, respectively, and 47% were coinfected. 500 blood donors seronegative for HIV and HCV were also evaluated. CCR5 haplotypes (HHA to HHG*2) were derived after genotyping nine CCR2-CCR5 polymorphisms. The association between CCR5 haplotypes with HIV and/or HCV seropositivity was determined using logistic regression analysis. Co-variates included in the models were length of intravenous drug use, HBV serostatus and copy number of CCL3L1, the gene encoding the most potent HIV-suppressive chemokine and ligand for CCR5. RESULTS Compared to IDUs seronegative for both HCV and HIV (HCV-/HIV-), IDUs who were HCV+/HIV- and HCV+/HIV+were 92% and 82%, respectively, less likely to possess the CCR5-HHG*1 haplotype, after controlling for co-variates (P(adjusted) = 1.89 × 10(-4) and 0.003, respectively). This association was mostly due to subjects bearing the CCR5 HHE and HHG*1 haplotype pairs. Approximately 25% and

Human immunodeficiency virus (HIV)-1 transmitted drug resistance in the drug-naïve population is of growing relevance in Estonia, where the number of antiretroviral (ARV) treatment-experienced subjects has been exponentially increasing during the last 10 y. The aim of this study was to estimate the rate of transmitted drug resistance among newly diagnosed subjects in Estonia in 2008. Genotypic resistance testing for viral genomic RNA was conducted for 201 subjects tested HIV-positive between 1 April and 30 November 2008. Of 145 genotyped viral strains in newly diagnosed patients, 123 were CRF06_cpx, 2 were subtype A1 and 3 were subtype B; in 17 cases viral sequences revealed recombinant structures similar to CRF06_cpx, subtype A1 and CRF02_AG. Resistance mutations were found in 8 (5.5%) virus strains, and 3 strains were resistant to at least 2 ARV classes. A total of 2.8% of sequences harboured mutations indicating nucleoside/nucleotide reverse transcriptase inhibitor resistance (M41L, M184V, M184I, T215C and T215D), 2.1% non-nucleoside reverse transcriptase inhibitor resistance (K103N, P225H) and 2.8% protease inhibitor resistance (M46I, L90M). These data suggest the need to extend genotypic HIV-1 drug resistance testing to newly diagnosed HIV-positive subjects to prevent potential ARV treatment failure.

Natural polymorphisms of HIV-1, often associated with drug resistance, are widely described in protease and reverse transcriptase regions but data on their presence in the integrase region, especially in non-B subtypes, are still very limited. We aimed to characterize naturally occurring polymorphisms in the integrase region in 104 treatment-naive and 10 treatment-experienced patients infected predominantly with HIV-1 CRF06_cpx and its recombinant with subtype A1 and/or CRF03_AB viruses. No primary drug resistance mutations against integrase inhibitors were found, but resistance-associated polymorphisms such as V72I, L74I, V201I, and T206S were seen in more than 90% of viruses. Substitutions E157Q and E157K, associated with raltegravir resistance, were found in only two CRF06_cpx strains. We conclude that similar to other HIV-1 non-B subtypes, the CRF06_cpx and its recombinants with subtype A1 and CRF03_AB are rich in integrase region natural polymorphisms, which may impact the development of resistance against integrase inhibitors.

BACKGROUND A high copy number of CCL3L1, the most potent human immunodeficiency virus (HIV)-suppressive chemokine, associates with reduced HIV susceptibility. Whether CCL3L1 influences acquisition of multiple blood-borne infections (eg, hepatitis C virus [HCV], HIV, and hepatitis B virus [HBV] infections), which occur commonly among injection drug users (IDUs), is unknown. METHODS We determined CCL3L1 copy number by real-time polymerase chain reaction among 374 Caucasian IDUs from Estonia; 285 were HCV positive, 208 were HIV positive, 177 were HCV and HIV positive, and 57 were HCV and HIV negative. RESULTS In univariate and multivariate analyses, HCV and HBV seropositivity and duration of IDU each strongly predicted HIV seropositivity. A high CCL3L1 copy number (>2) was associated with an 80% reduced risk of acquiring HIV infection after adjusting for age, sex, HCV and HBV status, CCR5-Delta32 polymorphism, and IDU duration (odds ratio, 0.20; 95% confidence interval, 0.09-0.45). By contrast, CCL3L1 gene dose did not influence HCV seropositivity. Among HCV-positive IDUs, there was a 3.5-fold overrepresentation and 65% underrepresentation of a high CCL3L1 copy number among HCV-positive, HIV-negative subjects and HCV-positive, HIV-positive subjects, respectively. CONCLUSION Among IDUs with extensive exposure to HCV and HIV, CCL3L1 copy number is a major determinant of HIV seropositivity but not of HCV seropositivity. The contrasting distribution of a protective high CCL3L1 copy number among HCV-positive, HIV-negative IDUs versus HCV-positive, HIV-positive IDUs may reflect that HIV preferentially selects for subjects with a low CCL3L1 gene dose.

All non-B HIV-1 subtypes and circulating recombinant forms (CRFs) are characterized by several polymorphisms in protease (PR) region. In addition, in recent years the increasing use of antiretroviral treatment (ART) has rapidly raised the spread of transmitted drug resistance. We aimed to determine the presence of naturally occurring polymorphisms and transmitted drug resistance mutations (DRMs) in ART naive HIV-1-positive subjects in Estonia. A total of 115 drug-naive HIV-1-infected subjects (mean age 27 years; 70% male; 65% infected via intravenous drug use and 34% by heterosexual contact) were enrolled. Viral genomic RNA from plasma was directly sequenced in PR, revertase (RT), and envelope (env) regions. Phylogenetic analysis of RT and env regions revealed that 89% and 3% of sequenced viruses belonged to CRF06_cpx and subtype A1, respectively, and 6% were described as unique recombinants (signed A1-06) between CRF06_cpx and subtype A1 viruses. No primary DRMs were found in PR or RT regions indicating the absence of transmitted drug resistance. The most common polymorphisms in the PR region were K14R, M36I, H69K, and L89M seen in 96%, 100%, 99%, and 100%, respectively. The clinical relevance of these polymorphisms in terms of success of ART has to be monitored in future clinical studies.

Background HTLV-1/2 and HIV likely share the same transmission routes and thus in Europe HTLV occurs mainly among IDUs, interests in clinical implications of HTLV-1/2 coinfection on HIV disease progression have risen. So far the available evidence suggests a protective role of HTLV-2 and adverse effect of HTLV-1 on HIV infection. In Europe, the overall prevalence of HTLV infection is up to 18% among IDUs and less than 1% among blood donors. However, there is no data regarding the overall prevalence of HTLV-1/2 in Eastern-European IDU population where the HIV epidemic mainly lies. We aimed to determine the prevalence of HTLV among IDUs in Estonia and compare it with the rate in healthy volunteers.

“Eastern-European type” of HIV-1 epidemic started among Estonian intravenous drug users (IDU) population in 2000. Unlike in other former Soviet Union countries the Estonian HIV-1 epidemic was not caused by subtype A1 viruses but recombinant form CRF06_cpx and its next generation recombinants with subtype A1. A high variety of different recombinant forms has also been described in other IDU driven HIV-1 epidemics in other countries. Aim: to sequence and describe the subtypic structure of near-full genome Estonian HIV-1 inter-subtype recombinant forms in Estonian IDU population. The study included plasma samples from 10 HIV positive subjects collected in 2000-2010. Samples were selected based on discordant subtypes or recombinant sequences in different genomic regions in recent Estonian studies. Patients’ viral genomic RNA was reverse transcribed, amplified, cloned and Sanger sequenced in four overlapping genomic regions. Phylogenetic trees were constructed using the Maximum-Likelihood method and recombination pattern was assessed using Simplot software (similarity and bootscanning blots). Drug resistance mutations and tropism were determined by Stanford HIV Drug Resistance Database and geno2pheno analysis tool using a false positive rate of 10%. According to bootscanning analysis the majority of strains (80%) indicated unique recombination structure between Estonian CRF06_cpx and Russian subtypes A1 sequences, whereas prevailing proportions of regions belong to the CRF06_cpx clade. Phylogenetic tree analysis confirmed these results indicating monophyletic clustering with Estonian CRF06_cpx or Russian subtype A1 sequences in corresponding regions. Of 10 sequences in one NNRTI DRM K103N and V179E was found and in 9 INI accessory mutations L74I was found. Of seven gag region sequences in three cases protease inhibitor associated substitution L449F, and in one case V128I, P452S and maturation inhibitor substitutions V370M were present. Of 9 env sequences one strain was R4 tropic, the others were R5 tropic. Analyzed sequences revealed high recombinational diversity of Estonian inter-subtype HIV-1 recombinant strains, which is also characteristic to other IDU HIV-1 epidemics. Nearly full-length genome sequences generated by this study provide reference material for high-throughput sequencing and for tracking the molecular epidemics in different risk groups and in Estonia and neighboring countries.

Estonian concentrated HIV epidemic started in August 2000 when rare HIV-1 CRF06_cpx was introduced into the population of intravenous drug users (IDUs). The majority of these HIV-positive subjects are likely co-infected with HCV but the prevalence of HCV and its genotypes during HIV concentrated epidemic in Estonia is largely unknown. The Estonian HIV database collects clinical and laboratory data (including HCV) of HIV-positive patients on medical care. Of 4500 patients on medical care 3500 are entered into Estonian HIV database (in total 8664 diagnosed). Aim: to describe the prevalence of HCV infection and the distribution of the HCV genotypes among HIV-positive subjects infected during HIV concentrated epidemic in Estonia. Data for present analyses was extracted from Estonian HIV database on 2nd of January 2014 and it comprised subjects diagnosed HIV-positive from 2000. In total 2,420 of 3,476 (70%) HIV-positive subjects were HCV antibody positive and 1,184 (64%) were HCV RNA positive. More than half of HCV-positives were men (66%) and the median age was 32 years (inter quartile range 30-36 y). The prevalence of HCV was higher in subjects verified to be HIV-positive between 2002 and 2010 as compared to between 2011 and 2013 (90% – 60% in 2002 – 2010 vs 50% – 45% in 2011 – 2013; p