HIV RNA levels are influenced by genetic characteristics of both the host and the virus. Here we applied machine learning techniques to determine if plasma-derived HIV-1 amino acid sequences can be used to predict spontaneous virologic control. We studied the relationship between HIV-1 env genotype and viral load in 20 chronically-infected patients undergoing treatment interruptions (SSITT, Swiss-Spanish Intermittent Treatment Trial) and in 104 primary HIV infected (PHI) patients before antiretroviral therapy (cART) and where applicable also after treatment stop. Extensive longitudinal sampling during the interruptions was performed in nine SSITT patients. Sequences obtained from these nine patients during the first virus rebound were used as a training data set and revealed a strong genetic signature (accuracy 98.6% in cross-validation) associated with control of viremia at levels below 5000 copies/mL of viral RNA maintained for at least two months after the final cART stop. The simple sequence pattern at gp120 positions 268E/358T was confirmed to be predictive of control in the clonal sequences originating from these patients during all subsequent rebounds. Sequences from the remaining 11 SSITT patients with less frequent sampling and from the PHI patients were used for external validation. High sensitivities (71-100%) and negative predictive values (80-100%) but low positive predictive values (12-40%) were achieved in the patient-wise analysis which was based on presence of the genetic pattern in all clones. These results suggest that presence of virus lacking the amino acid pattern 268E/358T is associated with VL >5000 at baseline of PHI and with low probability of spontaneous virologic control after treatment stop. Conversely, however, presence of 268E/358T does not predict control of viremia. These residues in HIV gp120 might affect in vivo HIV-1 fitness either at the level of Env function or influence susceptibility to adaptive or innate immune response.
HIV-1; envelope gp120; evolution; machine learning
Neisseria gonorrhoeae can rapidly develop resistance to antimicrobial agents. Over the last years, decreased gonococcal susceptibility to third-generation cephalosporins, especially cefixime, emerged worldwide. Therefore, current international guidelines recommend dual therapy for gonorrhoea with ceftriaxone plus either azithromycin or doxycycline. Gonococcal susceptibility data in Switzerland are sparse.
We investigated the prevalence of antibiotic susceptibility of N. gonorrhoeae in specimens collected between 1990 and 2012 at the University of Zurich, Switzerland. Minimum inhibitory concentrations (MICs) for cefixime, ceftriaxone, ciprofloxacin, and penicillin were determined by Etests. The European Committee on Antimicrobial Susceptibility Testing (EUCAST) breakpoints were used to define reduced susceptibility.
A total of 320 isolates were tested. Between 1990 and 2006 all tested samples were susceptible to both cephalosporins. Subsequently, the prevalence of elevated MICs for cefixime increased to 10.4% (2007/2008), 11.5% (2009/2010), and 11.4% (2011/2012); and for ceftriaxone to 2.4% (2007/2008), 4.7% (2009/2010), and 0% (2011/2012), respectively. The prevalence of resistance to ciprofloxacin (72.7%) and penicillin (22.7%) was high in 2011/2012.
Decreasing susceptibility of N. gonorrhoeae to third-generation cephalosporins in Switzerland supports treatment recommendations with ceftriaxone plus azithromycin or doxycycline. Health-care providers need to be aware of possible treatment failures with cephalosporins. Continued surveillance of gonococcal antimicrobial resistance is essential.
Neisseria gonorrhoeae; Gonorrhoea; Antimicrobial resistance; Cephalosporins
We studied whether the change in antibiotic susceptibility testing (AST) guidelines from CLSI to EUCAST influenced cumulative antibiograms in a tertiary care hospital in Switzerland.
Antibiotic susceptibilities of non-duplicate isolates collected within a one-year period before (period A) and after (period B) changing AST interpretation from CLSI 2009 to EUCAST 1.3 (2011) guidelines were analysed. In addition, period B isolates were reinterpreted according to the CLSI 2009, CLSI 2013 and EUCAST 3.1 (2013) guidelines.
The majority of species/drug combinations showed no differences in susceptibility rates comparing periods A and B. However, in some gram-negative bacilli, decreased susceptibility rates were observed when comparing CLSI 2009 with EUCAST 1.3 within period B: Escherichia coli / cefepime, 95.8% (CLSI 2009) vs. 93.1% (EUCAST 1.3), P=0.005; Enterobacter cloacae / cefepime, 97.0 (CLSI 2009) vs. 90.5% (EUCAST 1.3), P=0.012; Pseudomonas aeruginosa / meropenem, 88.1% (CLSI 2009) vs. 78.3% (EUCAST 1.3), P=0.002. These differences were still evident when comparing susceptibility rates according to the CLSI 2013 guideline with EUCAST 3.1 guideline. For P. aeruginosa and imipenem, a trend towards a lower antibiotic susceptibility rate in ICUs compared to general wards turned into a significant difference after the change to EUCAST: 87.9% vs. 79.8%, P=0.08 (CLSI 2009) and 86.3% vs. 76.8%, P=0.048 (EUCAST 1.3).
The change of AST guidelines from CLSI to EUCAST led to a clinically relevant decrease of susceptibility rates in cumulative antibiograms for defined species/drug combinations, particularly in those with considerable differences in clinical susceptibility breakpoints between the two guidelines.
HIV-positive patients receiving combination antiretroviral therapy (cART) frequently experience metabolic complications such as dyslipidemia and insulin resistance, as well as lipodystrophy, increasing the risk of cardiovascular disease (CVD) and diabetes mellitus (DM). Rates of DM and other glucose-associated disorders among HIV-positive patients have been reported to range between 2 and 14%, and in an ageing HIV-positive population, the prevalence of DM is expected to continue to increase. This study aims to develop a model to predict the short-term (six-month) risk of DM in HIV-positive populations and to compare the existing models developed in the general population.
All patients recruited to the Data Collection on Adverse events of Anti-HIV Drugs (D:A:D) study with follow-up data, without prior DM, myocardial infarction or other CVD events and with a complete DM risk factor profile were included. Conventional risk factors identified in the general population as well as key HIV-related factors were assessed using Poisson-regression methods. Expected probabilities of DM events were also determined based on the Framingham Offspring Study DM equation. The D:A:D and Framingham equations were then assessed using an internal-external validation process; area under the receiver operating characteristic (AUROC) curve and predicted DM events were determined.
Of 33,308 patients, 16,632 (50%) patients were included, with 376 cases of new onset DM during 89,469 person-years (PY). Factors predictive of DM included higher glucose, body mass index (BMI) and triglyceride levels, and older age. Among HIV-related factors, recent CD4 counts of<200 cells/µL and lipodystrophy were predictive of new onset DM. The mean performance of the D:A:D and Framingham equations yielded AUROC of 0.894 (95% CI: 0.849, 0.940) and 0.877 (95% CI: 0.823, 0.932), respectively. The Framingham equation over-predicted DM events compared to D:A:D for lower glucose and lower triglycerides, and for BMI levels below 25 kg/m2.
The D:A:D equation performed well in predicting the short-term onset of DM in the validation dataset and for specific subgroups provided better estimates of DM risk than the Framingham.
HIV; combination antiretroviral treatment; diabetes mellitus; risk equation
The serological response to treatment was studied in 264 syphilis patients; it was influenced by syphilis stage but not by human immunodeficiency virus infection and reinfection. Some of the recommendations of current guidelines are critically discussed, and amendments are proposed.
Background. Serology is the mainstay for syphilis diagnosis and treatment monitoring. We investigated serological response to treatment of syphilis according to disease stage and HIV status.
Methods. A retrospective cohort study of 264 patients with syphilis was conducted, including 90 primary, 133 secondary, 33 latent, and 8 tertiary syphilis cases. Response to treatment as measured by the Venereal Disease Research Laboratory (VDRL) test and a specific IgM (immunoglobulin M) capture enzyme-linked immunosorbent assay (ELISA; Pathozyme-IgM) was assessed by Cox regression analysis.
Results. Forty-two percent of primary syphilis patients had a negative VDRL test at their diagnosis. Three months after treatment, 85%–100% of primary syphilis patients had reached the VDRL endpoint, compared with 76%–89% of patients with secondary syphilis and 44%–79% with latent syphilis. In the overall multivariate Cox regression analysis, serological response to treatment was not influenced by human immunodeficiency virus (HIV) infection and reinfection. However, within primary syphilis, HIV patients with a CD4 count of <500 cells/μL had a slower treatment response (P = .012). Compared with primary syphilis, secondary and latent syphilis showed a slower serological response of VDRL (P = .092 and P < .001) and Pathozyme-IgM tests (P < .001 and P = .012).
Conclusions. The VDRL should not be recommended as a screening test owing to lack of sensitivity. The syphilis disease stage significantly influences treatment response whereas HIV coinfection only within primary syphilis has an impact. VDRL test titers should decline at least 4-fold within 3–6 months after therapy for primary or secondary syphilis, and within 12–24 months for latent syphilis. IgM ELISA might be a supplement for diagnosis and treatment monitoring.
Saline nasal irrigation (SNI) is often recommended as additional nonpharmacologic treatment, having proven its efficacy in acute and chronic rhinosinusitis and for therapy after sinonasal surgery. To date, however, no systematic review or meta-analysis exists showing the influence of SNI on allergic rhinitis (AR). This study aimed to establish the impact of SNI on symptoms of AR in different patient groups.
We conducted a systematic search of Medline, Embase, Cochrane Central Register of Controlled Trials, and ISI Web of Science databases for literature published from 1994 to 2010 on SNI in AR. Prospective, randomized, controlled trials that assessed the effects of SNI on four different outcome parameters were included. The evaluation focused on primary (symptom score) and secondary parameters (medicine consumption, mucociliary clearance, and quality of life).
Three independent reviewers chose 10 originals that satisfied the inclusion criteria (>400 participants total) from 50 relevant trials. SNI performed regularly over a limited period of up to 7 weeks was observed to have a positive effect on all investigated outcome parameters in adults and children with AR. SNI produced a 27.66% improvement in nasal symptoms, a 62.1% reduction in medicine consumption, a 31.19% acceleration of mucociliary clearance time, and a 27.88% improvement in quality of life.
SNI using isotonic solution can be recommended as complementary therapy in AR. It is well tolerated, inexpensive, easy to use, and there is no evidence showing that regular, daily SNI adversely affects the patient's health or causes unexpected side effects.
Adjunctive treatment; allergic rhinitis; hypertonic saline solution; isotonic saline solution; nasal douche; nasal lavage; nasal obstruction; pollinosis; saline nasal irrigation; sneezing
We present the case of an HIV infected male patient with erythema nodosum leprosum and function loss of the peroneus nerve as manifestations of lepromatous leprosy. Since symptoms occurred after initiation of antiretroviral therapy and recovery of the immune system, the clinical picture is suggestive of a rare form of an immune reconstitution inflammatory syndrome.
Long-term benefits of combination antiretroviral therapy (cART) initiation during primary HIV-1 infection are debated.
The evolution of plasma HIV-RNA (432 measurements) and cell-associated HIV-DNA (325 measurements) after cessation of cART (median exposure 18 months) was described for 33 participants from the Zurich Primary HIV Infection Study using linear regression and compared with 545 measurements from 79 untreated controls with clinically diagnosed primary HIV infection, respectively a known date for seroconversion.
On average, early treated individuals were followed for 37 months (median) after cART cessation; controls had 34 months of pre-cART follow-up. HIV-RNA levels one year after cART interruption were −0.8 log10 copies/mL [95% confidence interval −1.2;−0.4] lower in early treated patients compared with controls, but this difference was no longer statistically significant by year three of follow-up (−0.3 [−0.9; 0.3]). Mean HIV-DNA levels rebounded from 2 log10 copies [1.8; 2.3] on cART to a stable plateau of 2.7 log10 copies [2.5; 3.0] attained 1 year after therapy stop, which was not significantly different from cross-sectional measurements of 9 untreated members of the control group (2.8 log10 copies [2.5; 3.1]).
The rebound dynamics of viral markers after therapy cessation suggest that early cART may indeed limit reservoir size of latently infected cells, but that much of the initial benefits are only transient. Owing to the non-randomized study design the observed treatment effects must be interpreted with caution.
In the Zurich Primary HIV infection study (ZPHI), minority drug-resistant HIV-1 variants were detected in some acutely HIV-1-infected patients prior to initiation of early antiretroviral therapy (ART). Here, we investigated the reappearance of minority K103N and M184V HIV-1 variants in these patients who interrupted efficient early ART after 8–27 months according to the study protocol. These mutations are key mutations conferring drug resistance to reverse transcriptase inhibitors and they belong to the most commonly transmitted drug resistance mutations.
Early ART was offered to acutely HIV-1-infected patients enrolled in the longitudinal prospective ZPHI study. Six patients harboring and eleven patients not harboring drug-resistant viruses at low frequencies prior to ART were included in this substudy. Minority K103N and M184V HIV-1 variants were quantified in longitudinal plasma samples after treatment interruption by allele-specific real-time PCR. All 17 patients were infected with HIV-1 subtype B between 04/2003 and 09/2005 and received LPV/r+AZT+3TC during primary HIV-1 infection (PHI). Minority K103N HIV-1 variants reappeared after cessation of ART in two of four patients harboring this variant during PHI and even persisted in one of those patients at frequencies similar to the frequency observed prior to ART (<1%). The K103N mutation did not appear during treatment interruption in any other patient. Minority M184V HIV-1 variants were detected in two patients after ART interruption, one harboring and one not harboring these variants prior to ART.
Minority K103N HIV-1 variants, present in acutely HIV-1 infected patients prior to early ART, can reappear and persist after interruption of suppressive ART containing two nucleoside/nucleotide analogue reverse transcriptase inhibitors and a ritonavir-boosted protease inhibitor.
Nasal packs are indispensable in ENT practice. This study reviews current indications, effectiveness and risks of nasal packs and stents. In endoscopic surgery, nasal packs should always have smooth surfaces to minimize mucosal damage, improve wound healing and increase patient comfort. Functional endoscopic endonasal sinus surgery allows the use of modern nasal packs, since pressure is no longer required. So called hemostatic/resorbable materials are a first step in this direction. However, they may lead to adhesions and foreign body reactions in mucosal membranes. Simple occlusion is an effective method for creating a moist milieu for improved wound healing and avoiding dryness. Stenting of the frontal sinus is recommended if surgery fails to produce a wide, physiologically shaped drainage path that is sufficiently covered by intact tissue.
nasal packing; stenting; wound healing; FESS; occlusive wound care; septoplasty; turbinate surgery; nasal tamponade
The fraction of ambiguous nucleotide calls in bulk sequencing of human immunodeficiency virus type 1 (HIV-1) carries important information on viral diversity and the age of infection. In particular, a fraction of ambiguous nucleotides of >.5% provides evidence against a recent infection event <1 year ago.
Background. The time passed since the infection of a human immunodeficiency virus (HIV)–infected individual (the age of infection) is an important but often only poorly known quantity. We assessed whether the fraction of ambiguous nucleotides obtained from bulk sequencing as done for genotypic resistance testing can serve as a proxy of this parameter.
Methods. We correlated the age of infection and the fraction of ambiguous nucleotides in partial pol sequences of HIV-1 sampled before initiation of antiretroviral therapy (ART). Three groups of Swiss HIV Cohort Study participants were analyzed, for whom the age of infection was estimated on the basis of Bayesian back calculation (n = 3,307), seroconversion (n = 366), or diagnoses of primary HIV infection (n = 130). In addition, we studied 124 patients for whom longitudinal genotypic resistance testing was performed while they were still ART-naïve.
Results. We found that the fraction of ambiguous nucleotides increased with the age of infection with a rate of .2% per year within the first 8 years but thereafter with a decreasing rate. We show that this pattern is consistent with population-genetic models for realistic parameters. Finally, we show that, in this highly representative population, a fraction of ambiguous nucleotides of >.5% provides strong evidence against a recent infection event <1 year prior to sampling (negative predictive value, 98.7%).
Conclusions. These findings show that the fraction of ambiguous nucleotides is a useful marker for the age of infection.
Unnecessary or inappropriate use of antimicrobials is associated with the emergence of antimicrobial resistance, drug toxicity, increased morbidity and health care costs. Antimicrobial use has been reported to be incorrect or not indicated in 9–64% of inpatients. We studied the quality of antimicrobial therapy and prophylaxis in hospitalized patients at a tertiary care hospital to plan interventions to improve the quality of antimicrobial prescription.
Experienced infectious diseases (ID) fellows performed audits of antimicrobial use at regular intervals among all patients—with or without antimicrobials—hospitalized in predefined surgical, medical, haemato-oncological, or intensive care units. Data were collected from medical and nursing patient charts with a standardized questionnaire. Appropriateness of antimicrobial use was evaluated using a modified algorithm developed by Gyssens et al.; the assessment was double-checked by a senior ID specialist.
We evaluated 1577 patients of whom 700 (44.4%) had antimicrobials, receiving a total of 1270 prescriptions. 958 (75.4%) prescriptions were for therapy and 312 (24.6%) for prophylaxis. 37.0% of therapeutic and 16.6% of prophylactic prescriptions were found to be inappropriate. Most frequent characteristics of inappropriate treatments included: No indication (17.5%); incorrect choice of antimicrobials (7.6%); incorrect application of drugs (9.3%); and divergence from institutional guidelines (8%). Characteristics of inappropriate prophylaxes were: No indication (9%); incorrect choice of antimicrobials (1%); duration too long or other inappropriate use (6.7%). Patterns of inappropriate antimicrobial varied widely in the different hospital units; empirical prescriptions were more frequently incorrect than prescriptions based on available microbiological results.
Audits of individual patient care provide important data to identify local problems in antimicrobial prescription practice. In our study, antimicrobial prescriptions without indication, and divergence from institutional guidelines were frequent errors. Based on these results, we will tailor education, amend institutional guidelines and further develop the infectious diseases consultation service.
Although combination antiretroviral therapy (cART) initiated in the acute phase of HIV-1 infection may prevent expansion of the latent reservoir, its benefits remain controversial. In the current study, HIV-1 RNA transcription patterns in peripheral blood mononuclear cells (PBMC) were monitored during acute cART to assess the effect of early treatment on cellular viral reservoirs.
Acutely HIV-1 infected patients (n = 24) were treated within 3–15 weeks after infection. Patients elected to cease treatment after ≥1 year of therapy. HIV-1 DNA (vDNA), HIV-1 RNA species expressed both in latently and productively infected cells, unspliced (UsRNA), multiply spliced (MsRNA-tatrev; MsRNA-nef), and PBMC-associated extracellular virion RNA (vRex), expressed specifically by productively infected cells, were quantified in PBMC by patient matched real-time PCR prior, during and post cART. In a matched control-group of patients on successful cART started during chronic infection (n = 15), UsRNA in PBMC and vDNA were measured cross-sectionally. In contrast to previous reports, PBMC-associated HIV-1 RNAs declined to predominantly undetectable levels on cART. After cART cessation, UsRNA, vRex, and MsRNA-tatrev rebounded to levels not significantly different to those at baseline (p>0.1). In contrast, MsRNA-nef remained significantly lower as compared to pretreatment (p = 0.015). UsRNA expressed at the highest levels of all viral RNAs, was detectable on cART in 42% of patients with cART initiated during acute infection as opposed to 87% of patients on cART initiated during chronic infection (Fisher's exact test; p = 0.008). Accordingly, UsRNA levels were 105–fold lower in the acute as compared to the chronic group.
Early intervention resulted in profound depletion of PBMC expressing HIV-1 RNA. This is contrary to chronically infected patients who predominantly showed continuous UsRNA expression on cART. Thus, antiretroviral treatment initiated during the acute phase of infection prevented establishment or expansion of long-lived transcriptionally active viral cellular reservoirs in peripheral blood.
We have repeatedly detected Candidatus Neoehrlichia mikurensis, a bacterium first described in Rattus norvegicus rats and Ixodes ovatus ticks in Japan in 2004 in the blood of a 61-year-old man with signs of septicemia by 16S rRNA and groEL gene PCR. After 6 weeks of therapy with doxycycline and rifampin, the patient recovered.
Candidatus Neoehrlichia mikurensis; septicemia; human infection; 16S rRNA gene PCR; therapy; tick-borne pathogen; bacteria; dispatch
Combination antiretroviral treatment (cART) has been very successful, especially among selected patients in clinical trials. The aim of this study was to describe outcomes of cART on the population level in a large national cohort.
Characteristics of participants of the Swiss HIV Cohort Study on stable cART at two semiannual visits in 2007 were analyzed with respect to era of treatment initiation, number of previous virologically failed regimens and self reported adherence. Starting ART in the mono/dual era before HIV-1 RNA assays became available was counted as one failed regimen. Logistic regression was used to identify risk factors for virological failure between the two consecutive visits.
Of 4541 patients 31.2% and 68.8% had initiated therapy in the mono/dual and cART era, respectively, and been on treatment for a median of 11.7 vs. 5.7 years. At visit 1 in 2007, the mean number of previous failed regimens was 3.2 vs. 0.5 and the viral load was undetectable (<50 copies/ml) in 84.6% vs. 89.1% of the participants, respectively. Adjusted odds ratios of a detectable viral load at visit 2 for participants from the mono/dual era with a history of 2 and 3, 4, >4 previous failures compared to 1 were 0.9 (95% CI 0.4–1.7), 0.8 (0.4–1.6), 1.6 (0.8–3.2), 3.3 (1.7–6.6) respectively, and 2.3 (1.1–4.8) for >2 missed cART doses during the last month, compared to perfect adherence. From the cART era, odds ratios with a history of 1, 2 and >2 previous failures compared to none were 1.8 (95% CI 1.3–2.5), 2.8 (1.7–4.5) and 7.8 (4.5–13.5), respectively, and 2.8 (1.6–4.8) for >2 missed cART doses during the last month, compared to perfect adherence.
A higher number of previous virologically failed regimens, and imperfect adherence to therapy were independent predictors of imminent virological failure.
The aims of this study were to determine the incidence of diabetes among HIV-infected patients in the Data Collection on Adverse Events of Anti-HIV Drugs (D:A:D) cohort, to identify demographic, HIV-related, and combination antiretroviral therapy (cART)-related factors associated with the onset of diabetes, and to identify possible mechanisms for any relationships found.
RESEARCH DESIGN AND METHODS
D:A:D is a prospective observational study of 33,389 HIV-infected patients; diabetes is a study end point. Poisson regression models were used to assess the relation between diabetes and exposure to cART after adjusting for known risk factors for diabetes, CD4 count, lipids, and lipodystrophy.
Over 130,151 person-years of follow-up (PYFU), diabetes was diagnosed in 744 patients (incidence rate of 5.72 per 1,000 PYFU [95% CI 5.31–6.13]). The incidence of diabetes increased with cumulative exposure to cART, an association that remained significant after adjustment for potential risk factors for diabetes. The strongest relationship with diabetes was exposure to stavudine; exposures to zidovudine and didanosine were also associated with an increased risk of diabetes. Time-updated measurements of total cholesterol, HDL cholesterol, and triglycerides were all associated with diabetes. Adjusting for each of these variables separately reduced the relationship between cART and diabetes slightly. Although lipodystrophy was significantly associated with diabetes, adjustment for this did not modify the relationship between cART and diabetes.
Stavudine and zidovudine are significantly associated with diabetes after adjustment for risk factors for diabetes and lipids. Adjustment for lipodystrophy did not modify the relationship, suggesting that the two thymidine analogs probably directly contribute to insulin resistance, potentially through mitochondrial toxicity.
Although guidelines in individuals not infected with the human immunodeficiency virus (HIV) consider diabetes mellitus (DM) to be a coronary heart disease (CHD) equivalent, there is little information on its association with CHD in those infected with HIV. We investigated the impact of DM and preexisting CHD on the development of a new CHD episode among 33 347 HIV-infected individuals in the Data Collection on Adverse Events of Anti-HIV Drugs (D:A:D Study).
Methods and Results
Over 159 971 person-years, 698 CHD events occurred. After adjustment for gender, age, cohort, HIV transmission, ethnicity, family history of CHD, smoking, and calendar year, the rate of a CHD episode was 7.52 times higher (Poisson regression, 95% CI 6.02 to 9.39, P=0.0001) in those with preexisting CHD than in those without preexisting CHD, but it was only 2.41 times higher (95% CI 1.91 to 3.05, P=0.0001) in those with preexisting DM compared with those without DM. No statistical interactions were apparent between either diagnosis and sex; although older people with DM had an increased CHD rate compared with younger people, older people with preexisting CHD had a lower event rate. A statistically significant interaction between preexisting DM and CHD (P=0.003) suggested that the CHD rate in those with preexisting CHD and DM is lower than expected on the basis of the main effects alone.
DM and preexisting CHD are both important risk factors for CHD events in HIV-infected individuals. There is a need for targeted interventions to reduce the risk of CHD in both high-risk groups of HIV-infected individuals.
coronary disease; diabetes mellitus; risk factors; human immunodeficiency virus; epidemiology
Mathematical models based on kinetics of HIV-1 plasma viremia after initiation of combination antiretroviral therapy (cART) inferred HIV-infected cells to decay exponentially with constant rates correlated to their strength of virus production. To further define in vivo decay kinetics of HIV-1 infected cells experimentally, we assessed infected cell-classes of distinct viral transcriptional activity in peripheral blood mononuclear cells (PBMC) of five patients during 1 year after initiation of cART
In a novel analytical approach patient-matched PCR for unspliced and multiply spliced viral RNAs was combined with limiting dilution analysis at the single cell level. This revealed that HIV-RNA+ PBMC can be stratified into four distinct viral transcriptional classes. Two overlapping cell-classes of high viral transcriptional activity, suggestive of a virion producing phenotype, rapidly declined to undetectable levels. Two cell classes expressing HIV-RNA at low and intermediate levels, presumably insufficient for virus production and occurring at frequencies exceeding those of productively infected cells matched definitions of HIV-latency. These cells persisted during cART. Nevertheless, during the first four weeks of therapy their kinetics resembled that of productively infected cells.
We have observed biphasic decays of latently HIV-infected cells of low and intermediate viral transcriptional activity with marked decreases in cell numbers shortly after initiation of therapy and complete persistence in later phases. A similar decay pattern was shared by cells with greatly enhanced viral transcriptional activity which showed a certain grade of levelling off before their disappearance. Thus it is conceivable that turnover/decay rates of HIV-infected PBMC may be intrinsically variable. In particular they might be accelerated by HIV-induced activation and reactivation of the viral life cycle and slowed down by the disappearance of such feedback-loops after initiation of cART.
To evaluate the contribution of complement-mediated lysis to the in vivo activities of neutralizing antibodies, we analyzed the influence of complement activation on treatment success in a recent passive immunization trial with the neutralizing monoclonal antibodies 2G12, 2F5, and 4E10. Administration of monoclonal antibodies led to an immediate, high activation of the complement system even in the absence of viremia in the 14 participating human immunodeficiency virus-infected individuals. Lysis activity measured in patient plasma increased during passive immunization; however, the increases were modest and only partially attributable to the administration of antibodies. We found that unlike neutralization activity, lysis activity was not associated with treatment success in this trial. Compared to complement lysis mounted by the polyclonal antibody response in vivo, monoclonal antibodies were weak inducers of this activity, suggesting that polyclonal responses are more effective in reaching the required threshold of complement activation. Importantly, strong neutralization activity of the monoclonal antibodies did not predict complement lysis activity against patient and reference viruses, suggesting that these activities are not linked. In summary, our data support the notion that the in vivo activities of 2G12, 2F5, and 4E10 are likely due to direct neutralization or Fc receptor-mediated mechanisms such as phagocytosis and antibody-dependent cellular cytotoxicity.
The definition of plasma neutralizing antibody titers capable of controlling human immunodeficiency virus (HIV) infection in vivo is considered a critical step in vaccine development. Here we provide estimates for effective neutralization titers by assessing samples from a recent passive immunization trial with the neutralizing monoclonal antibodies (MAbs) 2G12, 2F5, and 4E10 using an analytic strategy that dissects the contributions of these MAbs to the total neutralization activity in patient plasma. Assessment of neutralization activities for six responding patients with partial or complete control of viremia during the MAb treatment and for the eight nonresponding patients revealed a significant difference between these groups: Among responders, MAb-mediated activity exceeded the autologous neutralization response by 1 to 2 log units (median difference, 43.3-fold), while in the nonresponder group, the autologous activity prevailed (median difference, 0.63-fold). In order to reach a 50% proportion of the responders in our study cohort, MAb neutralizing titers higher than 1:200 were required based on this analysis. The disease stage appears to have a significant impact on the quantities needed, since titers above 1:1,000 were needed to reach the same effect in chronic infection. Although our analysis is based on very small sample numbers and thus cannot be conclusive, our data provide a first estimate on how in vitro-measured neutralizing antibody activity can relate to in vivo efficacy in controlling HIV infection and may therefore provide valuable information for vaccine development. Interestingly, lower neutralizing antibody levels showed an effect in acute compared to chronic infection, suggesting that in early disease stages, therapeutic vaccination may show promise. Equally, this raises hopes that a preventive vaccine could become effective at comparatively lower neutralizing antibody titers.
Human immunodeficiency virus type 1 (HIV-1) transcription is subject to substantial fluctuation during the viral life cycle. Due to the low frequencies of HIV-1-infected cells, and because latently and productively infected cells collocate in vivo, little quantitative knowledge has been attained about the range of in vivo HIV-1 transcription in peripheral blood mononuclear cells (PBMC). By combining cell sorting, terminal dilution of intact cells, and highly sensitive, patient-specific PCR assays, we divided PBMC obtained from HIV-1-infected patients according to their degree of viral transcription activity and their cellular phenotype. Regardless of a patient's treatment status, the bulk of infected cells exhibited a CD4+ phenotype but transcribed HIV-1 provirus at low levels, presumably insufficient for virion production. Furthermore, the expression of activation markers on the surface of these CD4+ T lymphocytes showed little or no association with enhancement of viral transcription. In contrast, HIV-infected T lymphocytes of a CD4−/CD8− phenotype, occurring exclusively in untreated patients, exhibited elevated viral transcription rates. This cell type harbored a substantial proportion of all HIV RNA+ cells and intracellular viral RNAs and the majority of cell-associated virus particles. In conjunction with the observation that the HIV quasispecies in CD4+ and CD4−/CD8− T cells were phylogenetically closely related, these findings provide evidence that CD4 expression is downmodulated during the transition to productive infection in vivo. The abundance of viral RNA in CD4−/CD8− T cells from viremic patients and the almost complete absence of viral DNA and RNA in this cell type during antiretroviral treatment identify HIV+ CD4−/CD8 T cells as the major cell type harboring productive infection in peripheral blood.
Recently, passive immunization of human immunodeficiency virus (HIV)-infected individuals with monoclonal antibodies (MAbs) 2G12, 2F5, and 4E10 provided evidence of the in vivo activity of 2G12 but raised concerns about the function of the two membrane-proximal external region (MPER)-specific MAbs (A. Trkola, H. Kuster, P. Rusert, B. Joos, M. Fischer, C. Leemann, A. Manrique, M. Huber, M. Rehr, A. Oxenius, R. Weber, G. Stiegler, B. Vcelar, H. Katinger, L. Aceto, and H. F. Gunthard, Nat. Med. 11:615-622, 2005). In the light of MPER-targeting vaccines under development, we performed an in-depth analysis of the emergence of mutations conferring resistance to these three MAbs to further elucidate their activity. Clonal analysis of the MPER of plasma virus samples derived during antibody treatment confirmed that no changes in this region had occurred in vivo. Sequence analysis of the 2G12 epitope relevant N-glycosylation sites of viruses derived from 13 patients during the trial supported the phenotypic evaluation, demonstrating that mutations in these sites are associated with resistance. In vitro selection experiments with isolates of four of these individuals corroborated the in vivo finding that virus strains rapidly escape 2G12 pressure. Notably, in vitro resistance mutations differed, in most cases, from those found in vivo. Importantly, in vitro selection with 2F5 and 4E10 demonstrated that resistance to these MAbs can be difficult to achieve and can lead to selection of variants with impaired infectivity. This remarkable vulnerability of the virus to interference within the MPER calls for a further evaluation of the safety and efficacy of MPER-targeting therapeutic and vaccination strategies.
Cellular immune responses are an important correlate of hepatitis C virus (HCV) infection outcome. These responses are governed by the host's human leukocyte antigen (HLA) type, and HLA-restricted viral escape mutants are a critical aspect of this host-virus interaction. We examined the driving forces of HCV evolution by characterizing the in vivo selective pressure(s) exerted on single amino acid residues within nonstructural protein 3 (NS3) by the HLA types present in two host populations. Associations between polymorphisms within NS3 and HLA class I alleles were assessed in 118 individuals from Western Australia and Switzerland with chronic hepatitis C infection, of whom 82 (69%) were coinfected with human immunodeficiency virus. The levels and locations of amino acid polymorphisms exhibited within NS3 were remarkably similar between the two cohorts and revealed regions under functional constraint and selective pressures. We identified specific HCV mutations within and flanking published epitopes with the correct HLA restriction and predicted escaped amino acid. Additional HLA-restricted mutations were identified that mark putative epitopes targeted by cell-mediated immune responses. This analysis of host-virus interaction reveals evidence of HCV adaptation to HLA class I-restricted immune pressure and identifies in vivo targets of cellular immune responses at the population level.
Travelers who visited friends or relatives were more likely to receive a diagnosis of malaria or viral hepatitis than those who traveled for other reasons.
We evaluated the epidemiologic factors of patients seeking treatment for travel-associated illness from January 2004 through May 2005 at the University Hospital of Zurich. When comparing persons whose purpose of travel was visiting friends and relatives (VFR travelers; n = 121) with tourists and other travelers (n = 217), VFR travelers showed a distinct infectious disease and risk spectrum. VFR travelers were more likely to receive a diagnosis of malaria (adjusted odds ratio [OR] = 2.9, 95% confidence interval [CI] 1.2–7.3) or viral hepatitis (OR = 3.1, 95% CI 1.1–9) compared with other travelers but were less likely to seek pre-travel advice (20% vs. 67%, p = 0.0001). However, proportionate rates of acute diarrhea were lower in VFR (173 vs. 364 per 1,000 ill returnees). Travel to sub-Saharan Africa contributed most to malaria in VFR travelers. In countries with large migrant populations, improved public health strategies are needed to reach VFR travelers.
visiting friends and relatives; travel; traveler; infectious disease; malaria; diarrhea; pre-travel advice; research