Prevalence of drug resistance among HIV-1 treatment-naive patients in Greece during 2003-2015: Transmitted drug resistance is due to onward transmissions.

BACKGROUND: The prevalence of HIV-1 drug resistance among treatment-naïve patients ranges between 8.3% and 15% in Europe and North America. Previous studies showed that subtypes A and B were the most prevalent in the Greek HIV-1 epidemic. Our aim was to estimate the prevalence of resistance among drug naïve patients in Greece and to investigate the levels of transmission networking among those carrying resistant strains. METHODS: HIV-1 sequences were determined from 3428 drug naïve HIV-1 patients, in Greece sampled during 01/01/2003-30/6/2015. Transmission clusters were estimated by means of phylogenetic analysis including as references sequences from patients failing antiretroviral treatment in Greece and sequences sampled globally. RESULTS: The proportion of sequences with SDRMs was 5.98% (n=205). The most prevalent SDRMs were found for NNRTIs (3.76%), followed by N(t)RTIs (2.28%) and PIs (1.02%). The resistance prevalence was 22.2% based on all mutations associated with resistance estimated using the HIVdb resistance interpretation algorithm. Resistance to NNRTIs was the most common (16.9%) followed by PIs (4.9%) and N(t)RTIs (2.8%). The most frequently observed NNRTI resistant mutations were E138A (7.7%), E138Q (4.0%), K103N (2.3%) and V179D (1.3%). The majority of subtype A sequences (89.7%; 245 out of 273) with the dominant NNRTI resistance mutations (E138A, K103N, E138Q, V179D) were found to belong to monophyletic clusters suggesting regional dispersal. For subtype B, 68.1% (139 out of 204) of resistant strains (E138A, K103N, E138Q V179D) belonged to clusters. For N(t)RTI-resistance, evidence for regional dispersal was found for 27.3% and 21.6% of subtype A and B sequences, respectively. CONCLUSIONS: The TDR rate based on the prevalence of SDRM is lower than the average rate in Europe. However, the prevalence of NNRTI resistance estimated using the HIVdb approach, is high in Greece and it is mostly due to onward transmissions among drug-naïve patients.

Estimation of the Direct Cost of HIV-Infected Patients in Greece on an Annual Basis.

OBJECTIVE: HIV infection is currently regarded as a global chronic disease. The purpose of this study was to assess the direct cost of illness per patient per year in Greece. METHODS: A retrospective study for the estimation of the direct cost of HIV infection was performed from the third-party payer perspective. Data from 447 patients monitored in a general hospital of Athens were collected from their medical records. The survey involved all services and treatments that patients (stratified into three health states according to the number of CD4 cells/ml as defined by the Centers for Disease Control and Prevention classification system for HIV infection) received in 1 year, as well as demographic data. RESULTS: The annual direct cost per patient was calculated at €6859 ± €4699. Antiretroviral therapy cost was estimated at €5741, while the annual cost of providing health care services regardless of health state was computed at €1118, with laboratory investigation and imaging studies representing €924 (13.5%), outpatient visits €34 (0.5%), and hospitalization €160 (2.3%) of total cost, respectively. Overall, direct cost per patient was found to increase as the CD4 T lymphocytes decreased, leading to prolonged hospitalization and an increase in the number of laboratory tests. Direct cost for patients with more than 500 CD4 cells/µl was estimated at €6067, whereas for those with 200 to 499 cells/µl and less than 200 cells/µl, it was assessed at €6857 and €7654, respectively. CONCLUSIONS: The direct cost of HIV infection per patient increased as CD4 T lymphocytes decreased. The largest part of expenses was attributed to antiretroviral therapy, followed by laboratory tests/imaging studies, hospitalization, and finally outpatient visits.

Effects of highly active antiretroviral therapy on platelet activating factor metabolism in naive HIV-infected patients: ii) study of the abacavir/lamivudine/efavirenz HAART regimen.

Human Immunodeficiency Virus (HIV)-infected patients are at increased risk for cardiovascular diseases partly due to chronic inflammation. Some antiretroviral drugs and Highly Active Anti-Retroviral Therapy (HAART) regimens seem to be related and amplify this increased risk, especially the ones containing abacavir. Platelet-Activating-Factor (PAF) is a potent inflammatory mediator that is implicated in both cardiovascular diseases and HIV-related manifestations. Our objective is to study the in vivo effect of the abacavir/lamivudine/efavirenz first-line HAART regimen on PAF metabolism in HIV-infected patients. The specific activities of PAF basic biosynthetic enzymes in leukocytes and platelets, PAF-cholinephosphotransferase (PAF-CPT) and lyso-PAF-acetyltransferase (Lyso-PAF-AT), but also those of PAF-basic catabolic enzymes, PAF acetylhydrolase (PAF-AH) in leukocytes and platelets and Lipoprotein-associated-Phospholipase-A2 (LpPLA2) in plasma, were measured in blood samples of 10 asymptomatic naïve male HIV-infected patients just before and after 1, 3 and 6 months of treatment. CD4 cell counts, viral load and several biochemical markers were also measured in the same blood samples of these patients. The repeated ANOVA measures and the Pearson r criterion were used for studying statistical differences and correlations - partial correlations respectively. Even though viral load was decreased and CD4 cell counts were beneficially increased after treatment with the abacavir/lamivudine/efavirenz regimen, the main enzyme of the remodelling PAF-synthesis that is implicated in pro-atherogenic inflammatory procedures, Lyso-PAF-AT activity, was increased at 3 months of treatment in both leukocytes and platelets, while the main enzyme of PAF-degradation, PAF-AH, was increased as a response only in leukocytes at the 3rd month. Although the abacavir/lamivudine/efavirenz HAART regimen exhibits very efficient antiretroviral activities, on the other hand it induces an in vivo transient increase in the inflammation-related remodeling PAF-biosynthetic pathway. This finding supports the hypothesis of inflammation-mediated increased cardiovascular risk in HIV-infected patients during the first months of abacavir-containing HAART.

HIV-1 outbreak among injecting drug users in Greece, 2011: a preliminary report.

A significant increase (more than 10-fold) in the number of newly diagnosed HIV-1 infections among injecting drug users (IDUs) was observed in Greece during the first seven months of 2011. Molecular epidemiology results revealed that a large proportion (96%) of HIV-1 sequences from IDUs sampled in 2011 fall within phylogenetic clusters suggesting high levels of transmission networking. Cases originated from diverse places outside Greece supporting the potential role of immigrant IDUs in the initiation of this outbreak.

HIV-1 drug-resistance patterns among patients on failing treatment in a large number of European countries.

BACKGROUND: Information about patterns of HIV-1 drug resistance among treatment-exposed patients is crucial for the development of novel effective drugs. Currently no system exists that monitors patterns of resistance in patients failing therapy. METHODS: The study included 1,988 HIV-1 sequences from patients experiencing therapy failure collected between 2000 and 2004 in 15 European countries. Genotypic resistance was interpreted using the ANRS algorithm. Phenotypic resistance was predicted using the Virco geno- to phenotype system. RESULTS: 80.7% of the sequences included at least one drug-resistance mutation. Mutations were found for NRTIs (73.5%), NNRTIs (48.5%), and protease inhibitors (35.8%). Ninety percent of sequences with genotypic resistance harbored M184V, M41L, K103N, D67N, and/or T215Y. Among NRTIs, resistance was most frequently predicted for lamivudine. About half of all sequences had reduced susceptibility for NNRTIs. Resistance to most boosted protease inhibitors was found in < 25%. No sequence had resistance to all currently available drugs. CONCLUSION: Levels of resistance among patients with therapy failure were high. The patterns of resistance reflect resistance to drugs available for a longer time. Fully suppressive regimens can be designed even for the most mutated HIV because boosted protease inhibitors have remained active against most circulating viruses and new drug classes have become available.

Prevalence of resistance-associated mutations in newly diagnosed HIV-1 patients in Greece.

The prevalence of HIV-1 drug resistance mutations in naïve patients has been previously shown to differ greatly with the geographic origin. The purpose of this study was to prospectively estimate the prevalence of HIV-1 drug resistance in Greece by analyzing a representative sample of newly HIV-1 diagnosed patients, as part of the SPREAD collaborative study. Protease (PR) and partial reverse transcriptase (RT) sequences were determined from 101 newly diagnosed HIV-1 patients, in Greece, during the period September 2002--August 2003, representing one-third of the total newly diagnosed HIV-1 patients in the same time period. The prevalence of HIV-1 drug resistance was estimated according to the IAS-USA mutation table taking into account all mutations in RT and only major mutations in PR region. The overall prevalence of resistance was 9% [95% confidence interval (CI): 4.2--16.2%]. The prevalence of mutations associated with resistance to NRTIs was 5% (95% CI: 1.6--11.2%), for NNRTIs was 4% (95% CI: 1.1--9.8%), while no major resistance mutations were found in PR. No multi-class resistance was detected in the study population. The prevalence of resistant mutations in the recent seroconverters was 22%. For two individuals, there was clear evidence for transmitted resistance based on epidemiological information for a known source of HIV-1 transmission. The prevalence of the HIV-1 non-B subtypes and recombinants was 52%.

The effect of baseline CD4 cell count and HIV-1 viral load on the efficacy and safety of nevirapine or efavirenz-based first-line HAART.

BACKGROUND: A substantial number of patients start their first-line antiretroviral therapy at an advanced stage of an HIV-1 infection. Potential differences between specific drug regimens in antiviral efficacy and safety in these patients are of major importance. METHODS: A post-hoc analysis within the randomized controlled 2NN trial comparing efficacy between regimes containing nevirapine (NVP), efavirenz (EFV), or both, in addition to stavudine and lamivudine. PRIMARY OUTCOME: risk of virologic failure in different strata of baseline CD4 T-lymphocyte counts and plasma HIV-1 RNA concentrations (pVL). Virologic failure: never reaching a pVL < 400 copies/ml, or a rebound to two consecutive values > 400 copies/ml. RESULTS: The risk of virologic failure was increased at very low CD4 counts (< 25 x 10(6) cells/l) compared to CD4 counts > 200 x 10(6) cells/l [hazard ratio (HR), 1.28; 95% confidence interval (CI), 0.93-1.77]. The same was seen for a pVL > or = 100,000 copies/ml compared to a lower pVL (HR, 1.20; CI, 0.96-1.50). There were no statistically significant differences between NVP and EFV in risk of virologic failure within any of the CD4 or pVL strata, although EFV performed slightly better in the low CD4 stratum. The incidence of rash in the NVP group was significantly higher in female patients with higher CD4 cell counts, while adverse events in the EFV group were not associated with CD4 cell count. CONCLUSIONS: Initial antiretroviral therapy including NVP or EFV is effective in patients with an advanced HIV-1 infection. A high baseline CD4 cell count is associated with the occurrence of rash in female patients using NVP.

Body composition changes in protease inhibitor-naive HIV-infected patients treated with two nucleoside reverse transcriptase inhibitors.

OBJECTIVES: In the era of highly active antiretroviral treatment (HAART), there are insufficient data regarding lipodystrophy syndromes in HIV-1-infected patients treated with regimens that do not include protease inhibitors (PIs). We studied changes in body composition in HIV-1-infected patients before and 2 years after starting a non-PI-containing antiretroviral treatment regimen. METHODS: We studied retrospectively the whole body dual energy X-ray absorptiometry (DEXA) scans of 23 PI-naive HIV-1-infected patients (17 males, six females), aged 37.4 +/- 9.3 years with mean CD4 count 401 +/- 130 cells/microL. Thirteen patients were on zidovudine (ZDV) + lamivudine (3TC) and 10 on ZDV + didanosine (ddI). Subjects were evaluated before the beginning of antiretroviral treatment and approximately 24 months later. For each patient body weight, CD4 T-cell counts, bone mineral content (BMC), bone mineral density (BMD) and whole body as well as regional fat and lean body mass were evaluated. RESULTS: A significant decrease in BMC was observed, although the T scores remained within the normal limits. Our patients also exhibited a significant decrease in body weight due almost exclusively to fat loss, while lean mass was minimally affected. Fat loss was statistically significant in the arms and legs, but not in the trunk. The above changes were most prominent in the ZDV + 3TC treatment group; in this group of patients, fat loss was also evident in the trunk. Patients on ZDV + ddI, on the other hand, only showed a significant increase in their legs' lean mass; they preserved their total fat mass and exhibited no other significant changes between the two assessments. CONCLUSIONS: Dual NRTI therapy contributes to fat loss and reduction of bone mineral content in otherwise healthy, clinically stable, PI-naive HIV-infected adults. Compared with patients on ZDV + ddI, patients on ZDV + 3TC had a more prominent fat loss in all body regions.

Prevalence study of hospital-acquired infections in 14 Greek hospitals: planning from the local to the national surveillance level.

A prevalence study of hospital-acquired infections (HAI) was carried out in 14 of 112 Greek hospitals (15.7%), scattered throughout Greece. Five of seven Greek university hospitals and nine regional hospitals participated in the one-day study, and 3925 hospitalized patients (10.5% of the total hospital beds in Greece) were recorded. The aim of this project was to organize a surveillance of HAI with the participation of the greatest possible number of Greek hospitals, transferring the experience from the local Cretan infection control network in an effort to create a nationwide network. Special attention was paid to recruit all Greek university hospitals in our attempt to expand the study base. Co-ordination of the participating centres, education of the infection control teams on surveillance methods, preparation of agreed definitions, and elaboration of the protocol for the collection of the data were the major objectives of this study. The difficulties, however, were limited resources and the lack of skilled personnel. The overall prevalence of HAI was found to be 9.3%. The most common HAI recorded involved lower respiratory tract infections (30.3%), followed by urinary tract infections (22.7%), bloodstream infections (15.8%), and surgical site infections (14.8%). The greatest prevalence rate was found in the adult ICU (48.4%), followed by the neonatal ICU (30.3%). The duration of hospitalization, the number of operations, the total number of used devices and invasive procedures were significantly correlated with HAI. Positive cultures were found in 51.5% of the cases. The most frequently isolated micro-organisms were: Pseudomonas aeruginosa (16.6%), Escherichia coli (10.8%), Klebsiella pneumoniae (10.3%), Staphylococcus epidermidis (8.1%) and Staphylococcus aureus (7.6%). The administration of antibiotics was also recorded. The prevalence of antibiotic use was 51.4%.

Mutations associated with genotypic resistance to antiretroviral therapy in treatment naïve HIV-1 infected patients in Greece.

The widespread use of antiviral drugs against HIV has increased the prevalence of HIV-1 resistant strains among naïve individuals due to transmission of resistant strains. The purpose of this study was to investigate the presence of HIV-1 strains harboring resistance mutations in naïve patients in Greece. Blood samples were collected from 25 individuals. The DNA sequence of protease and partial reverse transcriptase regions (codons 41-223) were obtained by direct sequencing. Our results showed the absence of any primary resistance mutations in the study population. However, we were able to identify high prevalence of sequence polymorphisms at positions in reverse transcriptase region associated mainly with resistance to NNRTIs. Moreover, in protease region several secondary mutations were detected, suggesting the higher genetic variability of this region. The clinical significance of the polymorphisms associated with reduced susceptibility to NNRTIs remains to be clarified.

Low-dose IFN-alpha monotherapy in treatment-naive individuals with HIV-1 infection: evidence of potent suppression of viral replication.

To evaluate the safety and antiviral action of interferon-alpha (IFN-alpha) in HIV-1 infection, we undertook a proof of concept study in 27 treatment-naive patients. Eligible patients comprised two groups: the IFN-alphaT group (n = 17), which received 5 MIU IFN-alpha s.c. daily for 32 consecutive days, and the IFN-alphaNT group (n = 10), which did not receive IFN-alpha prior to highly active antiretroviral therapy (HAART), which was commenced on day 28 in both groups. IFN-alphaTreatment was well tolerated in 14 of the 17 patients of the IFN-alphaT group who completed the study. The mean HIV RNA reduction in the IFN-alphaT group on day 14 was 1.1 log(10). Viral load suppression was inversely associated with baseline viral load (p = 0.031). Four weeks after initiation of HAART, IFN-alphaT and IFN-alphaNT group patients had 2.40 and 1.82 log(10) HIV RNA reduction from baseline, respectively (p < 0.001). There was no evidence of cross-resistance with existing antiretrovirals in patients with HIV-RNA rebound after initial plasma viral load decline > or = 1 log(10) during IFN-alpha monotherapy. Thus, low daily IFN-alpha exhibits potent anti-HIV-1 activity in vivo without serious adverse effects. These properties render IFN-alpha an attractive candidate for further assessment as a constituent of HAART.

Virological and immunological response to HAART therapy in a community-based cohort of HIV-1-positive individuals.

PURPOSE: To determine virological and immunological response to highly active antiretroviral therapy (HAART) and to investigate factors influencing response in a community-based setting. METHOD: Plasma HIV RNA levels and CD4 cell counts were studied in 168 unselected individuals starting HAART including indinavir or ritonavir or hard-gel saquinavir-containing regimens. RESULTS: Overall, 60% of the patients reduced their HIV RNA to below 500 Eq/mL, but half of them experienced a subsequent virologic rebound. Patients with higher baseline HIV RNA, higher baseline CD4 cell count, and simultaneous initiation of combination therapy and patients on indinavir or ritonavir regimen were more likely to have virologic response within 6 months since HAART initiation. Patients with lower baseline CD4 cell count and with lower rates of viral clearance had a higher probability of a subsequent virologic rebound. Forty percent of the patients had increased their CD4 cell counts by more than 100 cells/microL (immunologic response). The probability of immunologic response was independent of baseline HIV RNA levels and CD4 cell count; however, the more complete the virologic suppression, the higher the probability of immunologic response. Thirty percent of the patients had discordance between virologic and immunologic responses. CONCLUSION: The rate of virologic failure in this unselected group of patients was higher than that observed in randomized clinical trials, but only a minority (11%) of the patients were treatment naïve. Starting combination therapy simultaneously and initiating antiretroviral therapy before advanced HIV disease has developed predict virologic response, whereas the magnitude of viral suppression predicts mid to long immunological response.

Molecular characterization of a complex, recombinant human immunodeficiency virus type 1 (HIV-1) isolate (A/G/J/K/?): evidence to support the existence of a novel HIV-1 subtype.

Recombination is one of several factors that contribute to the great genetic diversity of human immunodeficiency virus type 1 (HIV-1). In the current study, analysis of the full-length genome of a novel complex mosaic HIV-1 isolate (99GR303) from a Greek sailor who was possibly infected in Sierra Leone, Africa is presented. The 99GR303 isolate was found to comprise genomic regions belonging to subtypes A, G, J and K as well as of regions of a subtype that remains unclassified. For a partial region of env as well as vpr, no apparent similarity to the known HIV-1 subtypes or to any of the circulating recombinant forms was found. In fact, in the partial env gene, including the C2-V3 region, the 99GR303 isolate formed a new clade, suggesting the existence of an additional HIV-1 subtype. Thus, novel recombinants embody partial genomic regions which may have originated either from subtypes that existed in the past and became extinct or from contemporary subtypes that are extremely rare.

Comparative evaluation of the QUANTIPLEX HIV-1 RNA 2.0 and 3.0 (bDNA) assays and the AMPLICOR HIV-1 MONITOR v1.5 test for the quantitation of human immunodeficiency virus type 1 RNA in plasma.

HIV-1 RNA measurements from 84 plasma specimens obtained with the QUANTIPLEX HIV-1 RNA 2.0 and 3.0 (bDNA) assays (Chiron Diagnostics, Emeryville, CA) and with the AMPLICOR HIV-1 MONITOR Test, version 1.5 with ultra-sensitive specimen preparation (Roche Diagnostic Systems, Inc., Branchburg, NJ) were compared. The absolute RNA values of tested specimens differed significantly between bDNA 2.0 and bDNA 3.0 or Monitor v1.5 measurements (Wilcoxon signed-rank test P<0.001). Results generated with bDNA 3.0 or with Monitor v1.5 were approximately twofold greater than those generated with bDNA 2.0, with smaller differences at higher HIV-1 RNA levels and greater differences at RNA levels below 1000 copies per ml. Although highly correlated (r=0.92 and 0.86, respectively), viral load data generated with bDNA 2.0 and either bDNA 3.0 or Monitor v1.5 were in poor agreement. Concordant results (difference in log(10) copies per ml <0.5) were found at frequencies of 80% for bDNA 2.0 and bDNA 3.0 and only at 58.5% for bDNA 2.0 and Monitor v1.5. In contrast, bDNA 3.0 and Monitor v1.5 measurements were highly correlated (r=0.96) and in good agreement (92.7%).

Genotypic characterization of human immunodeficiency virus type 1 in Greece. Multicentre Study on HIV-1 Heterogeneity.

The HIV-1 subtype distribution in 83 HIV-1-seropositive individuals living in Greece was investigated by using the heteroduplex mobility assay (HMA), DNA sequencing, and phylogenetic analysis. The results revealed that partial HIV-1 gp120 sequences from 71 (86%) patients were subtype B, 5 (6%) were subtype A, 4 were subtype D (5%), 2 (2%) were subtype C, and 1 (1%) was subtype I. The subtype I isolate was documented in an intravenous drug user. A high prevalence (90-100%) of B isolates among intravenous drug users, hemophiliacs, and homosexual men was observed, in contrast to heterosexuals, among whom non-B subtypes seemed to be common (42.9%, p < 0.001). Among the Greek population subtype B is the most frequent (94%), in contrast to the high prevalence (57%) of non-B isolates found in emigrants living in Greece (p < 0.001). A heterosexual transmission case of subtype D in a Greek individual not traveling abroad was also documented. The broad HIV-1 diversity in Greece may be explained by population movements, such as migration and traveling.