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      Lymphocyte subsets in healthy Malawians: Implications for immunologic assessment of HIV infection in Africa

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          Abstract

          Background

          CD4 +T lymphocyte measurements are the most important indicator of mortality in HIV-infected individuals in resource-limited settings. There is currently a lack of comprehensive immunophenotyping data from African populations to guide the immunologic assessment of HIV infection.

          Objective

          To quantify variation in absolute and relative lymphocyte subsets with age in healthy Malawians.

          Methods

          Lymphocyte subsets in peripheral blood of 539 healthy HIV-uninfected Malawians stratified by age were enumerated by flow cytometry.

          Results

          B and T–lymphocyte and T-lymphocyte subset absolute concentrations peaked in early childhood then decreased to adult levels, whereas lymphocyte subset proportions demonstrated much less variation with age. Adult lymphocyte subsets were similar to those in developed countries. In contrast, high B-lymphocyte and CD8 +T-lymphocyte levels among children under 2 years, relative to those in developed countries, resulted in low CD4 +T-lymphocyte percentages that varied little between 0 and 5 years (35% to 39%). The CD4 +T-lymphocyte percentages in 35% of healthy children under 1 year and 18% of children age 1 to 3 years were below the World Health Organization threshold defining immunodeficiency in HIV-infected children in resource-limited settings. Thirteen percent of healthy children under 18 months old had a CD4:CD8T-lymphocyte ratio <1.0, which is commonly associated with HIV infection. All immunologic parameters except absolute natural killer lymphocyte concentration varied significantly with age, and percentage and overall absolute CD4 +T-lymphocyte counts were higher in females than males.

          Conclusion

          Although lymphocyte subsets in Malawian adults are similar to those from developed countries, CD4 +T-lymphocyte percentages in young children are comparatively low. These findings need to be considered when assessing the severity of HIV-related immunodeficiency in African children under 3 years.

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          Most cited references26

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          CLSI-Derived Hematology and Biochemistry Reference Intervals for Healthy Adults in Eastern and Southern Africa

          Background Clinical laboratory reference intervals have not been established in many African countries, and non-local intervals are commonly used in clinical trials to screen and monitor adverse events (AEs) among African participants. Using laboratory reference intervals derived from other populations excludes potential trial volunteers in Africa and makes AE assessment challenging. The objective of this study was to establish clinical laboratory reference intervals for 25 hematology, immunology and biochemistry values among healthy African adults typical of those who might join a clinical trial. Methods and Findings Equal proportions of men and women were invited to participate in a cross sectional study at seven clinical centers (Kigali, Rwanda; Masaka and Entebbe, Uganda; two in Nairobi and one in Kilifi, Kenya; and Lusaka, Zambia). All laboratories used hematology, immunology and biochemistry analyzers validated by an independent clinical laboratory. Clinical and Laboratory Standards Institute guidelines were followed to create study consensus intervals. For comparison, AE grading criteria published by the U.S. National Institute of Allergy and Infectious Diseases Division of AIDS (DAIDS) and other U.S. reference intervals were used. 2,990 potential volunteers were screened, and 2,105 (1,083 men and 1,022 women) were included in the analysis. While some significant gender and regional differences were observed, creating consensus African study intervals from the complete data was possible for 18 of the 25 analytes. Compared to reference intervals from the U.S., we found lower hematocrit and hemoglobin levels, particularly among women, lower white blood cell and neutrophil counts, and lower amylase. Both genders had elevated eosinophil counts, immunoglobulin G, total and direct bilirubin, lactate dehydrogenase and creatine phosphokinase, the latter being more pronounced among women. When graded against U.S.-derived DAIDS AE grading criteria, we observed 774 (35.3%) volunteers with grade one or higher results; 314 (14.9%) had elevated total bilirubin, and 201 (9.6%) had low neutrophil counts. These otherwise healthy volunteers would be excluded or would require special exemption to participate in many clinical trials. Conclusions To accelerate clinical trials in Africa, and to improve their scientific validity, locally appropriate reference ranges should be used. This study provides ranges that will inform inclusion criteria and evaluation of adverse events for studies in these regions of Africa.
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            Population-based hematologic and immunologic reference values for a healthy Ugandan population.

            To assess the validity of the reference values for hematologic and immunologic indices currently used in Africa, we evaluated blood samples from 3,311 human immunodeficiency virus (HIV)-negative Ugandans aged 1 week to 92 years. Erythrocyte, hemoglobin, and hematocrit levels and mean corpuscular volume all significantly increased with age (P < 0.001) and were independent of gender until the age of 13 years, after which the levels were higher in males than in females (P < 0.001). White blood cell, neutrophil, lymphocyte, basophil, and monocyte counts significantly declined with age until the age of 13 years (P < 0.001), with no differences by gender, while platelet counts declined with age (P < 0.001) and showed differences by gender only among adults older than age 24 years. CD4+- and CD8+-cell counts declined with age until the age of 18 years; thereafter, females had higher counts than males. The absolute values for many of these parameters differed from those reported for populations outside Africa, suggesting that it may be necessary to develop tables of reference values for hematologic and immunologic indices specific for the African population. This may be particularly important with regard to CD4+-cell counts among children because significant differences in absolute and percent CD4+-cell counts exist between the values for Western populations and the values for the population evaluated in our study. These differences could influence the decision to initiate antiretroviral therapy among children infected with HIV.
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              Reference Ranges for the Clinical Laboratory Derived from a Rural Population in Kericho, Kenya

              The conduct of Phase I/II HIV vaccine trials internationally necessitates the development of region-specific clinical reference ranges for trial enrolment and participant monitoring. A population based cohort of adults in Kericho, Kenya, a potential vaccine trial site, allowed development of clinical laboratory reference ranges. Lymphocyte immunophenotyping was performed on 1293 HIV seronegative study participants. Hematology and clinical chemistry were performed on up to 1541 cohort enrollees. The ratio of males to females was 1.9∶1. Means, medians and 95% reference ranges were calculated and compared with those from other nations. The median CD4+ T cell count for the group was 810 cells/µl. There were significant gender differences for both red and white blood cell parameters. Kenyan subjects had lower median hemoglobin concentrations (9.5 g/dL; range 6.7–11.1) and neutrophil counts (1850 cells/µl; range 914–4715) compared to North Americans. Kenyan clinical chemistry reference ranges were comparable to those from the USA, with the exception of the upper limits for bilirubin and blood urea nitrogen, which were 2.3-fold higher and 1.5-fold lower, respectively. This study is the first to assess clinical reference ranges for a highland community in Kenya and highlights the need to define clinical laboratory ranges from the national community not only for clinical research but also care and treatment.
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                Author and article information

                Journal
                J Allergy Clin Immunol
                J. Allergy Clin. Immunol
                The Journal of Allergy and Clinical Immunology
                Mosby
                0091-6749
                1097-6825
                January 2010
                January 2010
                : 125
                : 1
                : 203-208
                Affiliations
                [a ]Malawi-Liverpool-Wellcome Trust Clinical Research Programme, College of Medicine, University of Malawi, Blantyre, Malawi
                [b ]Department of Microbiology, College of Medicine, University of Malawi, Blantyre, Malawi
                [c ]Liverpool School of Tropical Medicine, University of Liverpool, Liverpool, United Kingdom
                [d ]Division of Medical Microbiology, School of Infection and Host Defence, University of Liverpool, Liverpool, United Kingdom
                [e ]Department of Zoology, University of Oxford, Oxford, United Kingdom
                [f ]Medical Research Council Centre for Immune Regulation and Clinical Immunology Service, School of Immunity and Infection, University of Birmingham, Birmingham, United Kingdom
                Author notes
                []Reprint requests: Calman A. MacLennan, DPhil, FRCPath, Medical Research Council Centre for Immune Regulation, School of Immunity and Infection, College of Medical and Dental Sciences, Institute of Biomedical Research, Wolfson Drive, The University of Birmingham Edgbaston, Birmingham, B15 2TT, United Kingdom. c.maclennan@ 123456bham.ac.uk
                Article
                YMAI7979
                10.1016/j.jaci.2009.10.010
                2887487
                19944455
                f11fcd90-3b4e-46a3-9f28-215c0c9d045d
                © 2010 Mosby, Inc.

                This document may be redistributed and reused, subject to certain conditions.

                History
                : 16 May 2009
                : 8 September 2009
                : 7 October 2009
                Categories
                Immune Deficiencies, Infection, and Systemic Immune Disorders

                Immunology
                nk, natural killer,lymphocyte subsets,immunophenotyping,art, antiretroviral therapy,hiv,africa,flow cytometry,cd4 lymphocyte count,aids,who, world health organization

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