Search

The author reported a case of mycobacterial infection, which was diagnosed after a lengthy process. However, the case is actually not one of tuberculosis in the stricter sense: the confirmed species—Mycobacterium avium intracellulare complex—belongs to the group of non-tuberculous mycobacteria. These are often found in the environment and, in cultures, are possibly only contaminants. However, in this case the diagnosis was firmly established following the criteria of the American Thoracic Society (ATS) – the specimens came from otherwise sterile body fluids (1). Microscopy using different staining methods, as mentioned in the article, is less relevant than cultures and PCR for confirming a diagnosis, since microscopy has lower sensitivity and, in contrast to the other methods, cannot differentiate between species (2). It therefore needs to be emphasized that the diagnostic mistake in the case report consisted mainly in not doing culture and PCR. Non-tuberculous mycobacteria—such as the described species—display sensitivities to anti-mycobacterial chemotherapy different from Mycobacterium tuberculosis complex. After resistance testing, recommended initial treatment should therefore consist of a combination of clarithromycin, ethambutol, and, if required, rifabutin. By contrast to the treatment described in the article, however, isoniazid is not the treatment of choice (3).

HLA, the most genetically diverse loci in the human genome, play a crucial role in host-pathogen interaction by mediating innate and adaptive cellular immune responses. A vast number of infectious diseases affect East Africa, including HIV/AIDS, malaria, and tuberculosis, but the HLA genetic diversity in this region remains incompletely described. This is a major obstacle for the design and evaluation of preventive vaccines. Available HLA typing techniques, that provide the 4-digit level resolution needed to interpret immune responses, lack sufficient throughput for large immunoepidemiological studies. Here we present a novel HLA typing assay bridging the gap between high resolution and high throughput. The assay is based on real-time PCR using sequence-specific primers (SSP) and can genotype carriers of the 49 most common East African class I HLA-A, -B, and -C alleles, at the 4-digit level. Using a validation panel of 175 samples from Kampala, Uganda, previously defined by sequence-based typing, the new assay performed with 100% sensitivity and specificity. The assay was also implemented to define the HLA genetic complexity of a previously uncharacterized Tanzanian population, demonstrating its inclusion in the major East African genetic cluster. The availability of genotyping tools with this capacity will be extremely useful in the identification of correlates of immune protection and the evaluation of candidate vaccine efficacy.

This book has its origins in a seminar series on transitions through the lifecourse which was part of the UK Teaching and Learning Research Programme (TLRP; see www.tlrp.org). It aims to bring together and evaluate insights about educational, life and work transitions from different elds of research and a range of theoretical orientations. The book responds to the injunction that researchers need to chart ‘what individuals actually do and how this is changing’ as a ‘rst step to understanding what it means’ (Bynner quoted by Hayward et al. 2005: 115). In different ways, the chapters that follow explore the concept of transitions and its contemporary importance in policy and educational practices. In doing so they enable the book to address the following questions:What are the main characteristics of transitions depicted in policy, practice • and research? How do different ideas and perspectives about transition, people’s agency, • identity and the effects of structural conditions help us to understand transitions better in research, policy and practice? Why are transitions a problem for some individuals and groups and, • conversely, for whom are transitions not a problem? What interventions, activities or practices are seen as useful in dealing with • transitions? What aspects of transitions are contested from different perspectives?•This book arose from a recognition that the research eld around transitions is fragmented both historically and between disciplines and theoretical orientations. The book does not claim to represent a unitary view about transitions in the lifecourse. Rather, it brings together policy, professional and academic concerns about transitions in the lifecourse through the conceptual lenses of identity, agency and structure. The theorising and ndings about transition, and the questions they raise about new forms of support, management and pedagogy, offered in the book, are meant to provide a basis for further thinking and empirical study.

Background Host restriction factors (HRFs) TRIM5alpha and APOBECs effect post-entry inhibition of HIV-1 replication in vitro but in vivo significance remains unclear. By acting as a first line of defense, HRFs could potentially act synergistically with adaptive/vaccine-induced immunity. HRFs are polymorphic in world populations, but little is known about their variation in East Africa, despite its high rate of HIV-1 infection. This study describes trim5 and apobec polymorphisms in East African populations.

Human immunodeficiency virus-1 subtypes A and C differ in the highly conserved Gag-TL9 epitope at a single amino acid position. Similarly, the TL9 presenting human leukocyte antigen (HLA) class I molecules B42 and B81 differ only at 6 amino acid positions. Here, we addressed the influence of such minor viral and host genetic variation on the TL9-specific CD8 T-cell response. The clonotypic characteristics of CD8 T-cell populations elicited by subtype A or subtype C were distinct, and these responses differed substantially with respect to the recognition and selection of TL9 variants. Irrespective of the presenting HLA class I molecule, CD8 T-cell responses elicited by subtype C exhibited largely comparable TL9 variant cross-recognition properties, expressed T-cell receptors that used almost exclusively the TRBV 12-3 gene, and selected for predictable patterns of viral variation within TL9. In contrast, subtype A elicited TL9-specific CD8 T-cell populations with completely different, more diverse TCRBV genes and did not select for viral variants. Moreover, TL9 variant cross-recognition properties were extensive in B81+ subjects but limited in B42+ subjects. Thus, minor viral and host genetic polymorphisms can dramatically alter the immunologic and virologic outcome of an epitope-specific CD8 T-cell response.

The acid-fast bacillus Mycobacterium tuberculosis is often the first manifestation of acquired immunodeficiency syndrome in patients infected with human immunodeficiency virus (HIV). This study was conducted to better understand the mechanism underlying M. tuberculosis-specific pathogenicity early after onset of HIV infection.M. tuberculosis-specific T helper 1 (Th1) cells were studied in HIV negative (n=114) and chronically HIV infected (n=68) Tanzanian subjects by using early secreted antigenic target 6 (ESAT6) protein or tuberculin (purified protein derivative) with interferon-gamma ELISPOT and intracellular cytokine staining. In a longitudinal study, the effect of acute HIV infection on M. tuberculosis-specific Th1 cells was determined by polychromatic flow cytometric analysis in 5 subjects with latent M. tuberculosis infection who became infected with HIV.In tuberculosis (TB)-asymptomatic subjects (i.e., subjects with unknown TB status who did not show clinical signs suggestive of TB), chronic HIV infection was associated with a decreased percentage of subjects with detectable M. tuberculosis-specific Th1 cells (P.001) a decrease which was not observed among subjects with active TB. Acute HIV infection induced a rapid depletion of M. tuberculosis-specific Th1 cells in 4 subjects remained TB asymptomatic, whereas the population of these cells remained stable in subjects who remained HIV negative (P.01).Taken together, these data suggest a mechanism of rapid M. tuberculosis-specific Th1 cell depletion that may contribute to the early onset of TB in individuals with latent M. tuberculosis infection who become HIV infected.

The advent of T-cell assay methodologies that are amenable to high throughput coupled with the availability of large libraries of overlapping peptides have revolutionized the fields of vaccine efficacy testing and cellular immune response assessment. Since T-cell assay performance is critically dependent upon the quality and specificity of the stimulating peptides, assurance of high-quality and reliable input peptides is an important aspect of assay validation. Herein, we demonstrate that individual peptides from large human immunodeficiency virus (HIV)-based peptide library sets obtained directly from two independent custom peptide suppliers contained contaminating peptides capable of giving false-positive results, which were consistent with nominal antigen-specific CD8 + T-cell responses. In-depth investigation of the cellular response in terms of responding CD8 + T-cell frequency and human leukocyte antigen (HLA) restriction led to the conclusion that one set of HIV type 1 (HIV-1)-derived peptides was contaminated with a peptide from human cytomegalovirus (HCMV), which is commonly used in cellular immunology research applications. Analytical characterization of the original stock of the suspect HIV-1 peptide confirmed the presence of ∼1% by weight of the HCMV peptide. These observations have critical implications for quality assurance (QA) and quality control (QC) of peptides used in clinical trials where cellular immune-based assays are important end-point determinants. We propose a simple schema of biological QA/QC protocols to augment the standard biochemical QA/QC analyses as a means to circumvent this and other problems that can affect cellular immune-based assay outcome and interpretation.

OBJECTIVES Southwest Tanzania is affected by an HIV-1 epidemic consisting of subtypes A, C, and D, and their recombinant forms. This study was designed to assess whether the Gag- and Nef-specific T-cell response is biased towards recognizing the infecting subtype. METHODS The infecting subtypes were characterized with a Multi-hybridization assay that discriminates between subtypes A, C and D. The interferon-gamma ELISPOT assay was used to detect the Gag- and Nef-specific T-cell responses in freshly isolated peripheral blood mononuclear cells in 56 seropositive patients. To study the HIV-specific T-cell responses, isolate-based Gag and Nef peptide sets representative of the locally occurring subtypes were used. The results were analysed at the total protein and single peptide level. RESULTS In the study population, 35% were infected with a pure C subtype, 24% and 23% with ACD or AC recombinant forms, respectively. The total magnitude (P < 0.01) and breadth (P < 0.01) of the Gag-specific T-cell response detected with the subtype C-Gag peptide set was significantly greater than that detected with either the subtype A-Gag or D-Gag peptide sets. No significant difference was observed in the Nef-specific response. In 85% of responses targeting the most immunodominant Gag epitopes with subtype-specific sequence differences, the best recognized epitope variant corresponded to the infecting subtype. CONCLUSIONS The Gag-specific T-cell response had a preference for recognizing peptides related to the infecting subtype.

Background In Tanzania, drug-resistant malaria parasites are an increasing public health concern. Because of widespread chloroquine (CQ) resistance Tanzania changed its first line treatment recommendations for uncomplicated malaria from CQ to sulfadoxine-pyrimethamine (SP) in 2001. Loss of SP sensitivity is progressing rapidly. SP resistance is associated with mutations in the dihydrofolate reductase (pfdhfr) and dihydropteroate synthase (pfdhps) genes. Methods In samples from 86 patients with uncomplicated Plasmodium falciparum malaria from Mbeya and Matema, Mbeya region, south-western Tanzania, the occurrence of mutations was investigated in the pfcrt and pfmdr1 genes which are associated with CQ resistance and in pfdhfr and pfdhps, conferring SP resistance, as well in cytb which is linked to resistance to atovaquone. Results Pfcrt T76 occurs in 50% and pfmdr1 Y86 in 51.7%. Pfdhfr triple mutations coexisting with pfdhps double mutations were detected in 64.3% of the P. falciparum isolates. This quintuple mutation is seen as a possible predictive molecular marker for SP treatment failure. Mutations of the cytb gene were not detected. Conclusion These findings of a high prevalence of mutations conferring SP resistance correspond to data of in vivo SP efficacy studies in other regions of Tanzania and underline the recommendation of changing first-line treatment to artemisinin-based combination therapy.

Background Rickettsioses are endemic in sub-Sahara Africa. Burden of disease, risk factors and transmission are hitherto sparsely described. Methods From the EMINI (Evaluating and Monitoring the Impact of New Interventions) population cohort, we randomly selected 1,228 persons above the age of 5 years from the nine participating communities in Mbeya region, Southwestern Tanzania, stratified by age, altitude of residence and ownership of domestic mammals, to conduct a cross-sectional seroprevalence study in. The aim was to estimate the seroprevalence of IgG antibodies against Spotted Fever Group (SFG) rickettsiae and to assess socioeconomic and environmental risk factors. Serology (indirect immunofluorescence) was performed at a dilution of 1:64. Results SFG-seropositivity in the cohort was found to be 67.9% (range among nine sites: 42.8–91.4%). Multivariable analysis revealed an association with age (prevalence ratio, PR per 10 years: 1.08; 95% CI 1.06–1.10), warmer temperatures (PR per °C: 1.38; 1.11–1.71), male gender (PR 1.08; 1.00–1.16), and low population density (PR per 1.000 persons/km²increase 0.96; 0.94–0.99). At higher elevations, higher cattle density was associated with higher seroprevalence. Conclusion SFG rickettsial infection seems to be common in the more rural population of Mbeya Region. Spread seems to be further limited by temperature and higher elevation. Examination of the contribution of SFG to febrile illnesses seems warranted in a prospective study to estimate the disease burden in the population. This will also allow determination of the causative pathogens., Author Summary We report a high seroprevalence for antibodies against Spotted Fever Group (SFG) rickettsiae in Southwestern Tanzania, a group of bacteria that is mostly transmitted by ectoparasites such as fleas, lice, mites, and ticks. Serum samples from 1.228 persons were selected, and 67.9% were positive indicating past infection. Seropositivity was clearly associated with warmer temperature, low population density and elevations below 1,500 m. These infections may have been caused by Rickettsia africae, which is transmitted to humans by a cattle tick, and was found in similar settings in Africa. To investigate further on the significance of SFG rickettsioses on disease burden in the area, investigations in acutely febrile patients should be conducted.

HIV-1 diversity, frequency of recombinants, and dual infection were determined in two populations with different HIV risk behavior. A high-risk cohort of 600 female bar workers and a normal-risk population of 1,108 antenatal clinic attendees and blood donors were recruited. Behavioral data were assessed and blood for HIV- 1 diagnosis and genotyping was sampled. HIV-1 subtypes were defined through the multiregion hybridization assay (MHA(acd)). HIV-1 prevalence differed significantly among the two populations. The prevalence was 67.8% in the population of bar workers and 17% in the normal-risk population (antenatal care attendees and blood donors). Within the normal-risk population the HIV-1 prevalence was lowest in the group of volunteer blood donors. The frequency of HIV-1 infection in women was 1.7 times higher than in men. The overall subtype distribution was A (8.5%), C (40.8%), D (3.8%), AC (25.4%), AD (5.4%), CD (8.8%), and ACD (7.3%). In the high-risk population there was a higher percentage of HIV-1 recombinant strains (54% vs. 40%, p0.05) and a higher frequency of dual infections (19% vs. 9%, p0.02) compared to the normal-risk population. High-risk populations may play an important role in the evolution of HIV, as they can provide an opportunity for the virus to coinfect, recombine, and adapt to the host-specific genetic background.

The European Commission (EC) has strong commitments and recognises the need to continue to ensure that HIV/AIDS research efforts receive global attention. The EC is facing this challenge in a global context and has made substantial investments together with European Developing Countries Clinical Trial Partnership (EDCTP) to formulate a program for the accomplishment of a scientific strategic plan promoting the European/African HIV vaccine development approach. The EC and EDCTP has convened a number of meetings by experts in basic and clinical virology, immunology, epidemiology, as well as industrial and regulatory representatives. The remit of the committee of experts was to define (1) objective criteria for selection of HIV candidates; (2) to determine criteria for selection of sites for clinical trials in Europe and Africa. The resulting consensus paper will guide the EC and EDCTP in developing HIV vaccine strategy and recommendations.

Human immunodeficiency virus type 1 (HIV-1) superinfection refers to the acquisition of another strain by an already infected individual. Here we report a comprehensive genetic analysis of an HIV-1 superinfection acquired heterosexually. The infected individual was in a high-risk cohort in Tanzania, was exposed to multiple subtypes, and was systematically evaluated every 3 months with a fluorescent multiregion genotyping assay. The subject was identified in the window period and was first infected with a complex ACD recombinant strain, became superinfected 6 to 9 months later with an AC recombinant, and was monitored for >2.5 years. The plasma viral load exceeded 400,000 copies/ml during the first 9 months of infection but resolved to the set point of 67,000 copies/ml by 3 months after superinfection; the CD4 cell count was 377 cells/μl at 30 months. Viral diversity was evaluated with techniques designed to fully sample the quasispecies, permitting direct observation of the evolution, temporal fluctuation, and intercompartment dynamics of the initial and superinfecting strains and recombinants derived from them. Within 3 months of superinfection, seven different molecular forms were detected in gag and six were detected in env . The proportions of forms fluctuated widely over time in plasma and peripheral blood mononuclear cells, illustrating how challenging the detection of dually infected individuals can be. Strain-specific nested PCR confirmed that the superinfecting strain was not present until the 9 month follow-up. This study further defines the parameters and dynamics of superinfection and will foster appropriate studies and approaches to gain a more complete understanding of risk factors for superinfection and its impact on clinical progression, epidemiology, and vaccine design.

The objective was to characterize HIV-1 strains in a potential vaccine trial cohort (CODE) in the Mbeya region of southwest Tanzania. Study volunteers (n = 3096) were recruited from urban areas in Mbeya Town using two different recruitment strategies and in a nearby rural village. Cryopreserved plasma from 507 HIV-1 prevalent cases was the source of viral RNA for HIV-1 genotyping by the Multi-region Hybridization Assay the MHAacd and selected strains were confirmed by complete genome sequencing. The overall HIV-1 prevalence was 16.6% [95% confidence interval (CI) 15.3– 17.9] within the cohort. HIV-1 prevalence was higher among women and in urban areas. Recruitment through advertisement targeted a high-risk urban male population for HIV-1 infection [adjusted odds ratio (adj. OR) 1.68; 95% CI 1.13–2.51] when compared with men recruited door-to-door. The complexity of the HIV-1 epidemic was also higher in urban areas evidenced by the high-risk of HIV-1 infection with a recombinant strain (adj. OR 2.69; 95% CI 1.08–6.69) and HIV-1 dual infection (adj. OR 5.16; 95% CI 1.07–24.9) mainly driven by urban men recruited through advertisement. Overall the urban epidemic was more genetically complex with higher prevalence and more recombinants and dual infections. Vaccine trials in Mbeya region can assess a complex HIV-1 population dynamic and determine vaccine efficacy in relationship to the genetic diversity of HIV-1 strains that challenge vaccines. (authors)

Coinfections with more than one human immunodeficiency virus type 1 (HIV-1) subtype appear to be the source of new recombinant strains and may be commonplace in high-risk cohorts exposed to multiple subtypes. Many potential dual infections have been identified during the HIV Superinfection Study in Mbeya, Tanzania, where 600 female bar workers who are highly exposed to subtypes A, C, and D have been evaluated every 3 months for over 3 years by use of the MHAacd HIV-1 genotyping assay. Here we describe an in-depth, longitudinal analysis of the viral quasispecies in a woman who was triply infected with HIV-1 and who developed AIDS and passed away 15 months after enrollment. The MHA results obtained at 0, 3, 6, 9, and 12 months revealed dual-probe reactivities and shifts in subtype over time, indicating a potential dual infection and prompting further investigation. The multiple infection was confirmed by PCR amplification of three genome regions by a multiple primer approach, followed by molecular cloning and sequencing. A highly complex viral quasispecies was found, including several recombinant forms, with vpu /gp120 being the most diverse region. A significant fluctuation in molecular forms over time was observed, showing that the serial sample format is highly desirable, if not essential, for the identification of multiple infections. In a separate experiment, we confirmed that the detection of coinfections is more efficient with the use of multiple amplification primers to overcome the primer bias that results from the enormous diversity in the HIV-1 genome.

BACKGROUND: Pilot studies suggest that a single, 2-g oral dose of azithromycin may be an alternative to a 2.4-MU intramuscular dose of penicillin G benzathine in the prevention and treatment of syphilis. We evaluated the efficacy of treatment with azithromycin in a developing country. METHODS: A total of 328 subjects, 25 with primary and 303 with high-titer (a titer of at least 1:8 on a rapid plasmin reagin [RPR] test) latent syphilis, were recruited through screening of high-risk populations in Mbeya, Tanzania, and randomly assigned to receive 2 g of azithromycin orally (163 subjects) or 2.4 million units of penicillin G benzathine intramuscularly (165 subjects). The primary outcome was treatment efficacy, with cure defined serologically (a decline in the RPR titer of at least two dilutions by nine months after treatment) and, in primary syphilis, by epithelialization of ulcers within one or two weeks. RESULTS: The average age of participants was 27.0 years, 235 (71.6 percent) were female, and 171 (52.1 percent) were seropositive for human immunodeficiency virus. Cure rates were 97.7 percent (95 percent confidence interval, 94.0 to 99.4) in the azithromycin group and 95.0 percent (95 percent confidence interval, 90.6 to 97.8) in the penicillin G benzathine group (95 percent confidence interval for the difference, -1.7 to 7.1 percent), achieving prespecified criteria for equivalence. Cure rates were also similar three and six months after treatment in the two groups and in all subgroups. Cure rates at three months were 59.4 percent (95 percent confidence interval, 51.8 to 67.1) in the azithromycin group and 59.5 percent (95 percent confidence interval, 51.8 to 67.3) in the penicillin G benzathine group and at six months were 85.5 percent (95 percent confidence interval, 79.4 to 90.6) and 81.5 percent (95 percent confidence interval, 74.8 to 87.4), respectively. CONCLUSIONS: Single-dose oral azithromycin is effective in treating syphilis and may be particularly useful in developing countries in which the use of penicillin G benzathine injections is problematic. However, recent reports of azithromycin-resistant Treponema pallidum in the United States indicate the importance of continued monitoring for resistance.

Objectives: To determine baseline prevalence of sexually transmitted infections (STI) and other reproductive tract infections (RTI) and their association with HIV as well as sociodemographic and behavioural characteristics in a newly recruited cohort of female bar workers in Mbeya Region, Tanzania Methods: 600 female bar workers were recruited from 17 different communities during September to November 2000 and underwent gynaecological examination, laboratory testing for HIV/STI, and interviews using structured questionnaires. Results: HIV-1 seroprevalence was 68%. Prevalences of STI/RTI were high titre syphilis (TPPA/RPR ⩾1/8), 9%; herpes simplex virus 2 antibodies, 87%; chlamydia, 12%; gonorrhoea, 22%; trichomoniasis, 24%; and bacterial vaginosis, 40%. HIV infection was associated with TPPA and HSV-2 seropositivity, bacterial vaginosis and clinically diagnosed genital ulcers, blisters, and warts. Reported high risk sexual behaviour during the past year (having multiple casual partners) was associated with prevalent STI. Conclusion: Female bar workers in Mbeya are at high risk of STI and HIV infection. Targeted STI/HIV prevention interventions for these women and their sexual partners need to be reinforced. Methods should be sought to improve healthcare seeking and to provide easily accessible and affordable STI care services.

Although the development of novel drugs and combination regimens for tuberculosis has accelerated in recent years, the pipeline remains thin and major challenges remain to be addressed in efficiently evaluating newer drugs to improve treatment outcomes, shorten duration of therapy and tackle drug resistance.

Surveillance for HIV infection among people at increased risk was conducted in five countries in South America. Seroprevalence studies were conducted in more than 36,000 people in Ecuador, Peru, Boliva, Uruguay, and Argentina, along with genetic analysis of the HIV-1 strains. In all countries, the prevalence of HIV-1 among men who have sex with men (MSM) was high (3-30%), whereas the prevalence among female commercial sex workers (FCSMs) was low (0.3-6%). By envelope heteroduplex mobility assay, subtype B predominated in MSM communities and in FCSWs in Ecuador, Bolivia, and Peru. A new genetic screening assay, the multiregion hybridization assay for subtypes B and F (MHA-bf), was developed to improve large-scale genetic screening in South America. MHA-bf can screen four regions of the genome for subtype B or subtype F, and thus can detect most recombinants. The sensitivity of MHA-bf when applied to a panel of pure subtypes and CRF12_BF was 100%, and 88% of unique recombinants were also detected as recombinant. Using MHA-bf, more than 80% of samples from Ecuador, Peru, and Bolivia were classified as pure subtype B, whereas in Uruguay and Argentina this proportion was only 30 to 40%. BF recombinants were the most prevalent form of HIV-1 in Uruguay and Argentina. Subtype B is the most common subtype in countries lacking injecting drug use (IDU) epidemics, whereas BF recombinants are more common in countries where extensive IDU epidemics have been documented, suggesting the ontogeny of recombinant strains in particular risk groups in South America.

Background HIV-1 entry into host cells is mediated by interactions between the virus envelope glycoprotein (gp120/gp41) and host-cell receptors. N-glycans represent approximately 50% of the molecular mass of gp120 and serve as potential antigenic determinants and/or as a shield against immune recognition. We previously reported that N-glycosylation of recombinant gp120 varied, depending on the producer cells, and the glycosylation variability affected gp120 recognition by serum antibodies from persons infected with HIV-1 subtype B. However, the impact of gp120 differential glycosylation on recognition by broadly neutralizing monoclonal antibodies or by polyclonal antibodies of individuals infected with other HIV-1 subtypes is unknown. Methods Recombinant multimerizing gp120 antigens were expressed in different cells, HEK 293T, T-cell, rhabdomyosarcoma, hepatocellular carcinoma, and Chinese hamster ovary cell lines. Binding of broadly neutralizing monoclonal antibodies and polyclonal antibodies from sera of subtype A/C HIV-1-infected subjects with individual gp120 glycoforms was assessed by ELISA. In addition, immunodetection was performed using Western and dot blot assays. Recombinant gp120 glycoforms were tested for inhibition of infection of reporter cells by SF162 and YU.2 Env-pseudotyped R5 viruses. Results We demonstrated, using ELISA, that gp120 glycans sterically adjacent to the V3 loop only moderately contribute to differential recognition of a short apex motif GPGRA and GPGR by monoclonal antibodies F425 B4e8 and 447-52D, respectively. The binding of antibodies recognizing longer peptide motifs overlapping with GPGR epitope (268 D4, 257 D4, 19b) was significantly altered. Recognition of gp120 glycoforms by monoclonal antibodies specific for other than V3-loop epitopes was significantly affected by cell types used for gp120 expression. These epitopes included CD4-binding site (VRC03, VRC01, b12), discontinuous epitope involving V1/V2 loop with the associated glycans (PG9, PG16), and an epitope including V3-base-, N332 oligomannose-, and surrounding glycans-containing epitope (PGT 121). Moreover, the different gp120 glycoforms variably inhibited HIV-1 infection of reporter cells. Conclusion Our data support the hypothesis that the glycosylation machinery of different cells shapes gp120 glycosylation and, consequently, impacts envelope recognition by specific antibodies as well as the interaction of HIV-1 gp120 with cellular receptors. These findings underscore the importance of selection of appropriately glycosylated HIV-1 envelope as a vaccine antigen.

Background To date, Alphavirus infections and their most prominent member, chikungunya fever, a viral disease which first became apparent in Tanzania in 1953, have been very little investigated in regions without epidemic occurrence. Few data exist on burden of disease and socio-economic and environmental covariates disposing to infection. Methods A cross-sectional seroprevalence study was undertaken in 1,215 persons from Mbeya region, South-Western Tanzania, to determine the seroprevalence of anti-Alphavirus IgG antibodies, and to investigate associated risk factors. Results 18% of 1,215 samples were positive for Alphavirus IgG. Seropositivity was associated with participant age, low to intermediate elevation, flat terrain and with IgG positivity for Rift Valley fever, Flaviviridae, and rickettsiae of the spotted fever group. When comparing the geographical distribution of Alphavirus seropositivity to that of Rift Valley fever, it was obvious that Alphaviruses had spread more widely throughout the study area, while Rift Valley fever was concentrated along the shore of Lake Malawi. Conclusion Alphavirus infections may contribute significantly to the febrile disease burden in the study area, and are associated with several arthropod-borne infections. Their spread seems only limited by factors affecting mosquitoes, and seems less restricted than that of Rift Valley fever., Author Summary The origin of febrile disease is often difficult to diagnose. In tropical countries, viral infections that are transmitted by arthropods include, among others, Alphavirus infections (e.g. chikungunya fever), dengue, West Nile, Yellow Fever and Rift Valley fever. In malaria endemic areas, these diseases are often mis-diagnosed and treated as malaria. Our study examined serum samples from 1,215 participants of a population survey from the Mbeya region, south-western Tanzania, for antibodies against Alphaviruses of the Semliki forest group as a sign of past infection. We found 18% of study participants positive, a surprisingly high number which points to a hitherto undetected circulation of Alphaviruses in the region. Among examined risk factors, even terrain, low to moderate elevation and participant age were associated with antibody positivity. Comparison with the distribution of Rift Valley fever seropositivity showed that Alphaviruses are more widely distributed throughout the study area, while Rift Valley fever seems to occur in a limited area close to Lake Malawi only.

Background It has been hypothesized that helminth infections increase HIV susceptibility by enhancing systemic immune activation and hence contribute to elevated HIV-1 transmission in sub-Saharan Africa. Objective To study systemic immune activation and HIV-1 co-receptor expression in relation to different helminth infections and in response to helminth treatment. Methods HIV-negative adults with (n = 189) or without (n = 57) different helminth infections, as diagnosed by Kato-Katz, were enrolled in Mbeya, Tanzania. Blinded to helminth infection status, T cell differentiation (CD45RO, CD27), activation (HLA-DR, CD38) and CCR5 expression was determined at baseline and 3 months after Albendazole/Praziquantel treatment. Plasma cytokine levels were compared using a cytometric bead array. Results Trichuris and Ascaris infections were linked to increased frequencies of “activated” CD4 and/or CD8 T cells (p, Author Summary Helminth infections are common in sub-Saharan Africa where about half of the population may be infected with one or more helminth species. HIV infection is also highly prevalent in this region. Because of the geographic overlap of helminth and HIV infections, it has been hypothesized that helminth infections may increase susceptibility to HIV by increasing systemic immune activation, which has been linked to increased HIV susceptibility. We therefore investigated the profile of T cell activation in individuals infected with different helminth species before and after helminth treatment within the WHIS cohort in Mbeya, Tanzania. Our study shows that systemic T cell activation differs between infections with different helminths. Particularly Trichuris but also Ascaris and S. mansoni infections were linked to increased frequencies of activated, HLA-DR+ T cells with relatively little effect of helminth treatment. Hookworm infection was associated with a trend towards decreased frequencies of activated, HLA-DR+ CD8+ T cells. Our study supports the concept that helminth infections, which are linked to systemic immune activation, could potentially also contribute to increased HIV transmission.

Background Tuberculosis (TB) causes significant morbidity and mortality globally. High efficacy in clinical trials of the current 4-drug 6-month regimen does not always translate into practice due to a heavy pill burden and long duration. New drugs are in late-phase clinical development, so trials that evaluate novel and existing drugs in combination are urgently needed. A multi-arm multi-stage (MAMS) design, with planned interim analyses allowing stopping of arms not showing benefit, has been developed and used in oncology. We have adapted this approach to create the PanACEA MAMS-TB 5-arm 3-stage phase II trial evaluating new treatments for TB.

Background: A high burden of tuberculosis (TB) occurs in sub-Saharan African countries and many cases of active TB and drug-resistant TB remain undiagnosed. Tertiary care hospitals provide an opportunity to study TB co-morbidity with non-communicable and other communicable diseases (NCDs/CDs). We evaluated the burden of undiagnosed pulmonary TB and multi-drug resistant TB in adult inpatients, regardless of their primary admission diagnosis, in a tertiary referral centre. Methodology/Principal Findings: In this prospective study, newly admitted adult inpatients able to produce sputum at the University Teaching Hospital, Lusaka, Zambia, were screened for pulmonary TB using fluorescent smear microscopy and automated liquid culture. The burden of pulmonary TB, unsuspected TB, TB co-morbidity with NCDs and CDs was determined. Sputum was analysed from 900 inpatients (70.6% HIV infected) 277 (30.8%) non-TB suspects, 286 (31.8%) TB suspects and 337 (37.4%) were already receiving TB treatment. 202/900 (22.4%) of patients had culture confirmed TB. TB co-morbidity was detected in 20/275 (7.3%) NCD patients, significantly associated with diabetes (P = 0.006, OR 6.571, 95% CI: 1.706-25.3). 27/202 (13.4%) TB cases were unsuspected. There were 18 confirmed cases of MDR-TB, 5 of which were unsuspected. Conclusions/Significance: A large burden of unsuspected pulmonary TB co-morbidity exists in inpatients with NCDs and other CDs. Pro-active sputum screening of all inpatients in tertiary referral centres in high TB endemic countries is recommended. The scale of the problem of undiagnosed MDR-TB in inpatients requires further study.

The level of bacterial activity is only poorly defined during asymptomatic Mycobacterium tuberculosis (MTB) infection. The objective was to study the capacity of a new biomarker, the expression of the T cell maturation marker CD27 on MTB-specific CD4 T cells, to identify active tuberculosis (TB) disease in subjects from a MTB and HIV endemic region. The frequency and CD27 expression of circulating MTB-specific CD4 T cells was determined in 96 study participants after stimulation with purified protein derivative (PPD) using intracellular cytokine staining for IFNgamma (IFNγ). Subjects were then stratified by their TB and HIV status. Within PPD responders, a CD27(-) phenotype was associated with active TB in HIV(-) (p = 0.0003) and HIV(+) (p = 0.057) subjects, respectively. In addition, loss of CD27 expression preceded development of active TB in one HIV seroconverter. Interestingly, in contrast to HIV(-) subjects, MTB-specific CD4 T cell populations from HIV(+) TB-asymptomatic subjects were often dominated by CD27(-) cells. These data indicate that down-regulation of CD27 on MTB-specific CD4 T cell could be used as a biomarker of active TB, potentially preceding clinical TB disease. Furthermore, these data are consistent with the hypothesis that late, chronic HIV infection is frequently associated with increased mycobacterial activity in vivo. The analysis of T cell maturation and activation markers might thus be a useful tool to monitor TB disease progression.

To address an existing controversy concerning the presence of HIV-1-specific antibodies of the IgA isotype in the female genital tract secretions of highly-exposed but persistently seronegative (HEPSN) women, 41 samples of plasma and cervicovaginal lavage (CVL) fluid were distributed to six laboratories for their blinded evaluation using ELISA with 10 different HIV-1 antigens, chemiluminescence-enhanced Western blots (ECL-WB), and virus neutralization. HIV-specific IgG or IgA antibodies in plasma samples from HEPSN women were absent or detectable only at low levels. In CVL, 11/41 samples displayed low levels of reactivity in ELISA against certain antigens. However, only one sample was positive in two of five laboratories. All but one CVL sample yielded negative results when analyzed by ECL-WB. Viral neutralizing activity was either absent or inconsistently detected in plasma and CVL. Plasma and CVL samples from 26 HIV-1-infected women were used as positive controls. Irrespective of the assays and antigens used, the results generated in all laboratories displayed remarkable concordance in the detection of HIV-1-specific antibodies of the IgG isotype. In contrast, IgA antibodies to HIV-1 antigens were not detected with consistency, and where present, IgA antibodies were at markedly lower levels than IgG. Although HIV-neutralizing activity was detected in plasma of all HIV-1-infected women, only a few of their CVL samples displayed such activity. In conclusion, frequent HIV-1 sexual exposure does not stimulate uniformly detectable mucosal or systemic HIV-1-specific responses, as convincingly documented in the present blindly performed study using a broad variety of immunological assays. Although HIV-1-infection leads to vigorous IgG responses in plasma and CVL, it does not stimulate sustained IgA responses in either fluid.

An optical minimum configuration of a laser interferometer for noncooperative scattering objects is investigated. An optimization of the SNR (signal-to-noise ratio) of the homodyne receiver is achieved by beam design. Scattering measurement object surfaces generate a speckle pattern on the detector surface which influences the SNR. Within the whole operating range of the optics, the transmitted light spot on the surface under investigation should therefore be small. The receiving optics should image the light spot onto the detector surface. The detection of a noncooperative object in a distance of 26 m is proven experimentally.

Background. The accurate diagnosis of TB in HIV-infected patients, particularly with advanced immunosuppression, is difficult. Recent studies indicate that a lipoarabinomannan (LAM) assay (Clearview-TB®-ELISA) may have some utility for the diagnosis of TB in HIV-infected patients; however, the precise subgroup that may benefit from this technology requires clarification. The utility of LAM in sputum samples has, hitherto, not been evaluated. Methods. LAM was measured in sputum and urine samples obtained from 500 consecutively recruited ambulant patients, with suspected TB, from 2 primary care clinics in South Africa. Culture positivity for M. tuberculosis was used as the reference standard for TB diagnosis. Results. Of 440 evaluable patients 120/387 (31%) were HIV-infected. Urine-LAM positivity was associated with HIV positivity (p = 0.007) and test sensitivity, although low, was significantly higher in HIV-infected compared to uninfected patients (21% versus 6%; p200 cells/mm3 (37% versus 0%; p = 0.003). Urine-LAM remained highly specific in all 3 subgroups (95%–100%). 25% of smear-negative but culture-positive HIV-infected patients with a CD4 3 were positive for urine-LAM. Sputum-LAM had good sensitivity (86%) but poor specificity (15%) likely due to test cross-reactivity with several mouth-residing organisms including actinomycetes and nocardia species. Conclusions. These preliminary data indicate that in a high burden primary care setting the diagnostic usefulness of urine-LAM is limited, as a rule-in test, to a specific patient subgroup i.e. smear-negative HIV-infected TB patients with a CD4 count 3, who would otherwise have required further investigation. However, even in this group sensitivity was modest. Future and adequately powered studies in a primary care setting should now specifically target patients with suspected TB who have advanced HIV infection.

Background: Molecular diagnosis using urine is established for many sexually transmitted diseases and is increasingly used to diagnose tumours and other infectious diseases. Storage of urine prior to analysis, whether due to home collection or bio-banking, is increasingly advocated yet no best practice has emerged. Here, we examined the stability of DNA in stored urine in two populations over 28 days. Methodology: Urine from 40 (20 male) healthy volunteers from two populations, Italy and Zambia, was stored at four different temperatures (RT, 4uC, 220u C& 280uC) with and without EDTA preservative solution. Urines were extracted at days 0, 1, 3, 7 and 28 after storage. Human DNA content was measured using multi-copy (ALU J) and single copy (TLR2) targets by quantitative real-time PCR. Zambian and Italian samples contained comparable DNA quantity at time zero. Generally, two trends were observed during storage; no degradation, or rapid degradation from days 0 to 7 followed by little further degradation to 28 days. The biphasic degradation was always observed in Zambia regardless of storage conditions, but only twice in Italy. Conclusion: Site-specific differences in urine composition significantly affect the stability of DNA during storage. Assessing the quality of stored urine for molecular analysis, by using the type of strategy described here, is paramount before these samples are used for molecular prognostic monitoring, genetic analyses and disease diagnosis.

Summary Background Over 1 year since the first reported case, the true COVID-19 burden in Ethiopia remains unknown due to insufficient surveillance. We aimed to investigate the seroepidemiology of SARS-CoV-2 among front-line hospital workers and communities in Ethiopia. Methods We did a population-based, longitudinal cohort study at two tertiary teaching hospitals involving hospital workers, rural residents, and urban communities in Jimma and Addis Ababa. Hospital workers were recruited at both hospitals, and community participants were recruited by convenience sampling including urban metropolitan settings, urban and semi-urban settings, and rural communities. Participants were eligible if they were aged 18 years or older, had provided written informed consent, and were willing to provide blood samples by venepuncture. Only one participant per household was recruited. Serology was done with Elecsys anti-SARS-CoV-2 anti-nucleocapsid assay in three consecutive rounds, with a mean interval of 6 weeks between tests, to obtain seroprevalence and incidence estimates within the cohorts. Findings Between Aug 5, 2020, and April 10, 2021, we did three survey rounds with a total of 1104 hospital workers and 1229 community residents participating. SARS-CoV-2 seroprevalence among hospital workers increased strongly during the study period: in Addis Ababa, it increased from 10·9% (95% credible interval [CrI] 8·3–13·8) in August, 2020, to 53·7% (44·8–62·5) in February, 2021, with an incidence rate of 2223 per 100 000 person-weeks (95% CI 1785–2696); in Jimma Town, it increased from 30·8% (95% CrI 26·9–34·8) in November, 2020, to 56·1% (51·1–61·1) in February, 2021, with an incidence rate of 3810 per 100 000 person-weeks (95% CI 3149–4540). Among urban communities, an almost 40% increase in seroprevalence was observed in early 2021, with incidence rates of 1622 per 100 000 person-weeks (1004–2429) in Jimma Town and 4646 per 100 000 person-weeks (2797–7255) in Addis Ababa. Seroprevalence in rural communities increased from 18·0% (95% CrI 13·5–23·2) in November, 2020, to 31·0% (22·3–40·3) in March, 2021. Interpretation SARS-CoV-2 spread in Ethiopia has been highly dynamic among hospital worker and urban communities. We can speculate that the greatest wave of SARS-CoV-2 infections is currently evolving in rural Ethiopia, and thus requires focused attention regarding health-care burden and disease prevention. Funding Bavarian State Ministry of Sciences, Research, and the Arts; Germany Ministry of Education and Research; EU Horizon 2020 programme; Deutsche Forschungsgemeinschaft; and Volkswagenstiftung.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Beta variant of concern (VOC) resists neutralization by major classes of antibodies from COVID-19 patients and vaccinated individuals. In this study, serum of Beta-infected patients revealed reduced cross-neutralization of wild-type virus. From these patients, we isolated Beta-specific and cross-reactive receptor-binding domain (RBD) antibodies. The Beta-specificity results from recruitment of VOC-specific clonotypes and accommodation of mutations present in Beta and Omicron into a major antibody class that is normally sensitive to these mutations. The Beta-elicited cross-reactive antibodies share genetic and structural features with wild type–elicited antibodies, including a public VH1-58 clonotype that targets the RBD ridge. These findings advance our understanding of the antibody response to SARS-CoV-2 shaped by antigenic drift, with implications for design of next-generation vaccines and therapeutics.

4 Mbeya Medical Research Program, Mbeya, Tanzania Here we explore associations between HLA variation and human immunodeficiency virus type 1 (HIV-1) acquisition and disease progression in a community cohort in Mbeya, Tanzania, a region that, despite harboring high rates of HIV- 1 infection, remains understudied. African-specific allele HLA-A*74:01 was associated with decreased risk of infection (odds ratio (OR), 0.37; 95% confidence interval (CI), 0.14- 0.80; ) and with protection from CD4 + cell counts P p .011 !200 cells/uL in women (OR, 0.31; 95% CI, 0.07-0.91; P p ) and men (OR, 0.15; 95% CI, 0.01-0.78; ). These .032 P p .020 associations remained significant after adjustment for link- age disequilibrium with HLA-B and HLA-C alleles. This ob- servation calls for additional investigation of mechanisms by which HLA-A*74:01 may influence HIV-1 acquisition and control of the infection.

Background The successful cure of tuberculosis (TB) is dependent on adherence to treatment. Various factors influence adherence, however, few are easily modifiable. There are limited data regarding correlates of psychological distress and their association with non-adherence to anti-TB treatment. Methods In a trial of a new TB test, we measured psychological distress (K-10 score), TB-related health literacy, and morbidity (TBscore), prior to diagnosis in 1502 patients with symptoms of pulmonary TB recruited from clinics in Cape Town (n = 419), Harare (n = 400), Lusaka (n = 400), Durban (n = 200), and Mbeya (n = 83). Socioeconomic, demographic, and alcohol usage-related data were captured. Patients initiated on treatment had their DOTS cards reviewed at two-and six-months. Results 22 %(95 % CI: 20 %, 25 %) of patients had severe psychological distress (K-10 ≥ 30). In a multivariable linear regression model, increased K-10 score was independently associated with previous TB [estimate (95 % CI) 0.98(0.09-1.87); p = 0.0304], increased TBscore [1(0.80, 1.20); p

Background: The World Health Organisation (WHO) estimates that 3 million cases of tuberculosis (TB) are missed every year. Identification and treatment of these are critical to achieving global TB control. Patients with sub-clinical TB, extra-pulmonary TB, and drug-resistant TB are difficult to diagnose and may be missed at all points of healthcare. An autopsy study was conducted to ascertain the burden of TB at post-mortem in adults who died in the inpatient general medical wards at a tertiary care referral center in Lusaka, Zambia. Methods: Complete whole body autopsies were performed on 125 adult inpatients. Pathological examination involved two stages: (1) Gross pathology was recorded, and samples were taken from all organs for histopathology and cryopreservation; and (2) Histopathological examination of tissue after appropriate staining. Specific pathology and diseases identified on examination were recorded. Lung tissues were processed using the GeneXpert MTB/RIF Assay. Primary outcome measures were specific diseases stratified by HIV status. Secondary outcomes were missed TB and drug-resistant TB cases. Findings: Of 125 adults, median age 35. years (IQR: 29-43), 80 (64%) were male and 101 (80.8%) were HIV-positive. Tuberculosis was the most common finding at autopsy with 78/125 cases (62.4%), of which 66/78 (84.6%) were HIV-infected. There were 35/78 cases (44.9%) with extra-pulmonary TB, the odds of which were higher among HIV-infected cases (aOR 5.14 (95% CI: 1.04-25.4), p=. 0.045); 25.6% (20/78) of the TB cases were not diagnosed ante-mortem; and 13/78 (16.7%) of the TB cases had undiagnosed MDR-TB. Other autopsy findings included: pyogenic pneumonia 36.8% (46/125); bacterial meningitis 7.2% (9/125); cardiac failure 7.2% (9/125); and malignancies 8.8% (11/125). Prevalence of HIV did not differ between TB and non-TB cases (84.6% vs. 74.5%: p=. 0.163). Interpretation: TB remains an important cause of death in adult inpatients. A substantial number of inpatients with TB and MDR-TB are not diagnosed by the current cascade of healthcare. Inpatient settings in high TB endemic countries should be included in WHO 'high risk' groups, and heightened clinical awareness and more proactive screening for TB and MDR-TB in all inpatients should be required.