Dynamics of two separate but linked HIV-1 CRF01_AE outbreaks among injection drug users in Stockholm, Sweden, and Helsinki, Finland.

Detailed phylogenetic analyses were performed to characterize an HIV-1 outbreak among injection drug users (IDUs) in Stockholm, Sweden, in 2006. This study investigated the source and dynamics of HIV-1 spread during the outbreak as well as associated demographic and clinical factors. Seventy Swedish IDUs diagnosed during 2004 to 2007 were studied. Demographic, clinical, and laboratory data were collected, and the V3 region of the HIV-1 envelope gene was sequenced to allow detailed phylogenetic analyses. The results showed that the Stockholm outbreak was caused by a CRF01_AE variant imported from Helsinki, Finland, around 2003, which was quiescent until the outbreak started in 2006. Local Swedish subtype B variants continued to spread at a lower rate. The number of new CRF01_AE cases over a rooted phylogenetic tree accurately reflected the transmission dynamics and showed a temporary increase, by a factor of 12, in HIV incidence during the outbreak. Virus levels were similar in CRF01_AE and subtype B infections, arguing against differences in contagiousness. Similarly, there were no major differences in other baseline characteristics. Instead, the outbreak in Stockholm (and Helsinki) was best explained by an introduction of HIV into a standing network of previously uninfected IDUs. The combination of phylogenetics and epidemiological data creates a powerful tool for investigating outbreaks of HIV and other infectious diseases that could improve surveillance and prevention.

Host-Epstein-Barr virus relationship affected by immunostimulation in HIV-infected patients representing distinct progressor profile groups.

BACKGROUND: Human immunodeficiency virus (HIV) infection with pronounced immunosuppression disrupts Epstein-Barr virus (EBV)-host balance with increased lymphoma risk. We explored whether different host responses to HIV are reflected in the EBV-host balance. METHODS: Eleven unvaccinated HIV-positive patients and 16 participants in a vaccine trial were included in the study. Blood samples were collected, B cells extracted, and EBV DNA load was determined using a semiquantitative polymerase chain reaction (PCR) method. RESULTS: Treatment-naïve patients with a history of symptomatic primary HIV infection showed non-significant, but higher EBV load compared to untreated long-term non-progressors. A significant difference in HIV RNA titres between these groups correlated weakly to EBV DNA load. Patients in the vaccine trial with recombinant HIV gp160 and/or adjuvant and with a history of symptomatic primary HIV infection, showed a 1-log increase in EBV load compared to patients with long-lasting HIV disease. CONCLUSION: Different host responses to HIV infection, especially in combination with vaccination, can be reflected in the EBV-host balance.

Prophylaxis and treatment of HIV-1 infection in pregnancy: Swedish recommendations 2010.

Prophylaxis and treatment with antiretroviral drugs and the use of elective caesarean section have resulted in a very low mother-to-child transmission of human immunodeficiency virus (HIV) during recent years. The availability of new antiretroviral drugs, updated general treatment guidelines and increasing knowledge of the importance of drug resistance, have necessitated regular revisions of the "Prophylaxis and treatment of HIV-1 infection in pregnancy" recommendations. For these reasons, The Swedish Reference Group for Antiviral Therapy (RAV) updated the 2007 recommendations at an expert meeting that took place on 25 March 2010. The most important revisions from the previous recommendations are: (1) it is recommended that treatment during pregnancy starts at the latest at gestational week 14-18; (2) ongoing efficient treatment at confirmed pregnancy may, with a few exceptions, be continued; (3) lopinavir/r and atazanavir/r are equally recommended protease inhibitors; (4) if maternal HIV RNA is >50 copies/ml close to delivery, a planned caesarean section, intravenous zidovudine, oral nevirapine for the mother and post-exposure prophylaxis for the infant with 3 antiretroviral drugs are recommended; (5) for delivery at <34 gestational weeks, intravenous zidovudine and oral nevirapine for the mother and at 48-72 h for the infant is recommended, in addition to other prophylaxis; (6) intravenous zidovudine is not recommended when HIV RNA is <50 copies/ml and a caesarean section is performed; (7) it is recommended that prophylaxis for the infant is started within 4 h; (8) prophylactic zidovudine for the infant may be administered twice daily instead of 4 times a day, as was the case previously; and (9) the number of sampling occasions for the infant has been decreased.

Regulation of human vitamin D(3) 25-hydroxylases in dermal fibroblasts and prostate cancer LNCaP cells.

In this study, we examined whether 1alpha,25-dihydroxyvitamin D(3) (calcitriol), phenobarbital, and the antiretroviral drug efavirenz, drugs used by patient groups with high incidence of low bone mineral density, could affect the 25-hydroxylase activity or expression of human 25-hydroxylases in dermal fibroblasts and prostate cancer LNCaP cells. Fibroblasts express the 25-hydroxylating enzymes CYP2R1 and CYP27A1. LNCaP cells were found to express two potential vitamin D 25-hydroxylases-CYP2R1 and CYP2J2. The presence in different cells of nuclear receptors vitamin D receptor (VDR), pregnane X receptor (PXR), and constitutive androstane receptor (CAR) was also determined. Phenobarbital suppressed the expression of CYP2R1 in fibroblasts and CYP2J2 in LNCaP cells. Efavirenz suppressed the expression of CYP2R1 in fibroblasts but not in LNCaP cells. CYP2J2 was slightly suppressed by efavirenz, whereas CYP27A1 was not affected by any of the two drugs. Calcitriol suppressed the expression of CYP2R1 in both fibroblasts and LNCaP cells but had no clear effect on the expression of either CYP2J2 or CYP27A1. The vitamin D(3) 25-hydroxylase activity in fibroblasts was suppressed by both calcitriol and efavirenz. In LNCaP cells, consumption of substrate (1alpha-hydroxyvitamin D(3)) was used as indicator of metabolism because no 1alpha,25-dihydroxyvitamin D(3) product could be determined. The amount of 1alpha-hydroxyvitamin D(3) remaining in cells treated with calcitriol was significantly increased. Taken together, 25-hydroxylation of vitamin D(3) was suppressed by calcitriol and drugs. The present study provides new information indicating that 25-hydroxylation of vitamin D(3) may be regulated. In addition, the current results may offer a possible explanation for the impaired bone health after treatment with certain drugs.

pol gene sequence variation in Swedish HIV-2 patients failing antiretroviral therapy.

There is limited knowledge about how to treat and interpret results from genotypic resistance assays in HIV-2 infection. Here, genetic variation in HIV-2 pol gene was studied in 20 of 23 known HIV-2 cases in Sweden. Five patients with signs of virological treatment failure were longitudinally studied. Clinical, virological and immunological data were collected and the protease (PR) and first half of the reverse transcriptase (RT) was amplified and directly sequenced from plasma samples. Moderate to extensive genetic evolution was observed in four of the five patients who failed treatment. Some mutations occurred at positions known to confer resistance in HIV-1, but many occurred at other positions in PR and RT. All patients had been treated with zidovudine alone or in combination with other antiretroviral drugs, but none displayed a mutation at position 215, which is the primary zidovudine resistance site in HIV-1. Instead, a E219D mutation evolved in virus from two patients and a Q151M mutation evolved in two other patients. A M184V mutation indicative of lamivudine resistance was detected in three patients. The virus of one patient who had been treated with ritonavir, nelfinavir, and lopinavir successively acquired nine unusual mutations in the protease gene, most of which are not considered primary or secondary resistance mutations in HIV-1. Our data indicate that the evolutionary pathways that lead to antiretroviral resistance in HIV-2 and HIV-1 exhibit both similarities and differences. Genotypic HIV-2 resistance assays cannot be interpreted using algorithms developed for HIV-1, instead new algorithms specific for HIV-2 have to be developed.

Epstein Barr virus DNA analysis in blood predicts disease progression in a rare case of plasmablastic lymphoma with effusion.

BACKGROUND: In HIV-1-infected patients a long lasting CD4+ cell decline influences the host-EBV balance and thereby increases the risk for EBV related malignancies. In spite of a world-wide access to combination antiretroviral therapy (cART) there are still a considerable number of HIV-1-infected patients who will develop severe immunodeficiency. These undiagnosed HIV-1 infected patients, so called late testers, demonstrate an increased lymphoma risk, compared to patients diagnosed early. Consecutive individual screening for EBV DNA-load in late testers might be a useful predictor of emerging EBV-malignancy. METHODS: Patient biopsies and ascites were analyzed morphologically, by immuncyto-histochemistry and in-situ hybridization. Viral DNA and RNA load were quantified by PCR. Cell lines from primary tumor and from ascites, were established in vitro and further analyzed. RESULT: We here report on a case of EBV-positive lymphoma in an AIDS patient, first presenting with pleural effusion and ascites and was thus initially considered a primary effusion lymphoma (PEL) but was later diagnosed as a plasmablastic lymphoma (PBL). The patient had responded to cART with undetectable HIV-RNA and increased CD4 cell count one year prior to lymphoma presentation. At the time of lymphoma diagnosis the HIV-RNA values were <50 RNA-copies per mL blood (undetectable) and the CD4-positive cell count 170 ×106/L. The lymphoma was CD45-negative and weakly CD22- and CD30-positive. The patient had a history of Kaposi sarcoma and HHV-8 seropositivity. The lymphoma biopsies, and three cell lines derived on different occasions from the tumor cell effusion, were all EBV-positive but HHV-8 negative.A noticeable EBV-DNA load decline was observed during the remission of the lymphoma following CHOP-therapy. The EBV-DNA load increased dramatically at the time of recurrence. CONCLUSION: EBV DNA load might be useful in monitoring the effect of lymphoma treatment as well as in estimating the risk of EBV-associated lymphoma in HIV-1 infected patients with pronounced immunosuppression.

Epstein-Barr virus genome load is increased by therapeutic vaccination in HIV-l carriers, and further enhanced in patients with a history of symptomatic primary infection.

OBJECTIVE: Epstein-Barr virus (EBV) infection is an established risk factor for B-cell lymphomas in Human Immunodeficiency virus (HIV)-1 infected patients. A disturbed EBV-host relationship is seen in patient groups with a high risk for EBV-associated lymphomas. We have analysed this relationship by measuring EBV-DNA in the blood of HIV-1 carriers. METHOD: EBV-DNA load in B-cells was monitored by PCR in non- or insufficiently antiretroviral treated and rgp160-vaccinated HIV-patients. RESULTS: Both asymptomatic HIV-infected and AIDS-patients showed a 25-40-fold increase in the number of B cell associated EBV-DNA copies compared to healthy controls. Patients included in a vaccine trial with recombinant HIV gp160 showed a 5-fold increase of EBV load compared to non-immunised patients and a 50-fold increase compared to healthy controls. There was no difference whether they received vaccine or "placebo". Vaccinated patients with a history of symptomatic primary HIV-1 infection (PHI) had a 280-fold increase in median EBV load compared to healthy controls, thus suggesting a synergistic effect between the vaccination and PHI, which hypothetically could affect lymphoma risk. CONCLUSIONS: We recommend analysis of EBV-load and long term follow up of lymphoma risk in all therapeutic HIV-1 vaccination trials.

Characterization of HIV-specific CD4+ T cell responses against peptides selected with broad population and pathogen coverage.

CD4+ T cells orchestrate immunity against viral infections, but their importance in HIV infection remains controversial. Nevertheless, comprehensive studies have associated increase in breadth and functional characteristics of HIV-specific CD4+ T cells with decreased viral load. A major challenge for the identification of HIV-specific CD4+ T cells targeting broadly reactive epitopes in populations with diverse ethnic background stems from the vast genomic variation of HIV and the diversity of the host cellular immune system. Here, we describe a novel epitope selection strategy, PopCover, that aims to resolve this challenge, and identify a set of potential HLA class II-restricted HIV epitopes that in concert will provide optimal viral and host coverage. Using this selection strategy, we identified 64 putative epitopes (peptides) located in the Gag, Nef, Env, Pol and Tat protein regions of HIV. In total, 73% of the predicted peptides were found to induce HIV-specific CD4+ T cell responses. The Gag and Nef peptides induced most responses. The vast majority of the peptides (93%) had predicted restriction to the patient's HLA alleles. Interestingly, the viral load in viremic patients was inversely correlated to the number of targeted Gag peptides. In addition, the predicted Gag peptides were found to induce broader polyfunctional CD4+ T cell responses compared to the commonly used Gag-p55 peptide pool. These results demonstrate the power of the PopCover method for the identification of broadly recognized HLA class II-restricted epitopes. All together, selection strategies, such as PopCover, might with success be used for the evaluation of antigen-specific CD4+ T cell responses and design of future vaccines.

The Effect of Antiretroviral Combination Treatment on Epstein-Barr Virus (EBV) Genome Load in HIV-Infected Patients.

We evaluated the effect of combination anti-retroviral treatment (cART) on the host control of EBV infection in moderately immunosuppressed HIV-1 patients. Twenty HIV-1 infected individuals were followed for five years with repeated measurements of EBV DNA load in peripheral blood lymphocytes in relation to HIV-RNA titers and CD4+ cell counts. Individuals with optimal response, i.e. durable non-detectable HIV-RNA, showed a decline of EBV load to the level of healthy controls. Individuals with non-optimal HIV-1 control did not restore their EBV control. Long-lasting suppression of HIV-replication after early initiation of cART is a prerequisite for re-establishing the immune control of EBV.

Prophylaxis and treatment of HIV-1 infection in pregnancy: Swedish Recommendations 2007.

Prophylaxis and treatment with antiretroviral drugs, a consequent low viral load, and the use of elective Caesarean section, are factors that radically decrease the risk of HIV transmission from mother to child during pregnancy and delivery. The availability of new antiretroviral drugs, updated general treatment guidelines and increasing knowledge of the importance of drug resistance, have necessitated recurrent revisions of the recommendations for 'Prophylaxis and treatment of HIV-1 infection in pregnancy'. For these reasons, The Swedish Reference Group for Antiviral Therapy (RAV) has, at an expert meeting on May 4, 2007, once more updated the treatment recommendations of 1999, 2002 and 2005, which were defined in cooperation with the Swedish Medical Products Agency (Läkemedelsverket). This new text takes the recently updated general HIV treatment recommendations into account. Furthermore, the very low risk of HIV transmission when the mother is treated with combination antiretroviral therapy, has undetectable levels of viraemia and no obstetric risk factors, has been considered in the recommendations concerning the mode of delivery. Finally, the recommendations for monitoring of infants born to HIV-infected mothers have been modified. The recommendations are evidence graded in accordance with the Oxford Centre for Evidence Based Medicine, 2001 (see http://www.cebm.net/levels_of_evidence.asp#levels).

Children born to HIV-1-infected women in Sweden in 1982-2003: trends in epidemiology and vertical transmission.

To describe the HIV-1 epidemic among childbearing women and their children in Sweden, a population-based analysis of data on all known mother-child pairs in Sweden with perinatal exposure to HIV-1 1982-2003 was conducted. The mother-to-child transmission (MTCT) rate in children prospectively followed from birth decreased from 24.7% in 1985-1993 to 5.7% in 1994-1998 and 0.6% in 1999-2003. The use of antiretroviral treatment of the mother during pregnancy and/or prophylactic antiretroviral intervention increased from 2.3% to 91.6% during the same period, and the elective cesarean delivery rate increased from 8.0% to 80.3%. No MTCT of HIV-1 occurred in Sweden after 1999.Fifty-one vertically HIV-1-infected children aged 2.7 to 17.6 years were living in Sweden by 31 December 2003, 71% being treated with antiretroviral agents. No HIV-1-related child death has been reported in Sweden after 1996. The conclusion is that MTCT of HIV-1 can be almost eliminated when appropriate resources are available. A national pregnancy screening program for HIV-1 running since 1987 with a high acceptance rate and the implementation of measures to prevent MTCT since 1994 have resulted in a significant decrease in the number of infected children. Inasmuch as knowledge of the infection status of the mother is crucial for reduction in MTCT of HIV-1, continued antenatal screening is important even in a low-prevalence country such as Sweden.

Evolution of human immunodeficiency virus type 2 coreceptor usage, autologous neutralization, envelope sequence and glycosylation.

To investigate why human immunodeficiency virus type 2 (HIV-2) is less virulent than HIV-1, the evolution of coreceptor usage, autologous neutralization, envelope sequence and glycosylation was studied in sequentially obtained virus isolates and sera from four HIV-2-infected individuals. Neutralization of primary HIV-2 isolates was tested by a cell line-based assay and IgG purified from patients' sera. Significant autologous neutralization was observed for the majority (39 of 54) of the HIV-2 serum-virus combinations tested, indicating that neutralization escape is rare in HIV-2 infection. Furthermore, sera from 18 HIV-2 patients displayed extensive heterologous cross-neutralization when tested against a panel of six primary HIV-2 isolates. This indicates that HIV-2 is intrinsically more sensitive to antibody neutralization than HIV-1. In line with earlier reports, HIV-2 isolates could use several alternative receptors in addition to the major coreceptors CCR5 and CXCR4. Intrapatient evolution from CCR5 use to CXCR4 use was documented for the first time. Furthermore, CXCR4 use was linked to the immunological status of the patients. Thus, all CXCR4-using isolates, except one, were obtained from patients with CD4 counts below 200 cells microl(-1). Sequence analysis revealed an association between coreceptor usage and charge of the V3 loop of the HIV-2 envelope, as well as an association between the rate of disease progression and the glycosylation pattern of the envelope protein. Furthermore, HIV-2 isolates had fewer glycosylation sites in the V3 domain than HIV-1 (two to three versus four to five). It is proposed here that HIV-2 has a more open and accessible V3 domain than HIV-1, due to differences in glycan packing, and that this may explain its broader coreceptor usage and greater sensitivity to neutralizing antibodies.