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 ::  Abstract
 :: Introduction
 ::  Materials and Me...
 :: Results
 :: Discussion
 ::  References
 ::  Article Tables

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  Table of Contents     
ORIGINAL ARTICLE
Year : 2012  |  Volume : 58  |  Issue : 3  |  Page : 176-179

Reliability of absolute lymphocyte count as a marker to assess the need to initiate antiretroviral therapy in HIV-infected children


Department of Pediatrics, Pediatric HIV Clinic, B. J. Wadia Hospital for Children, Mumbai, Maharashtra, India

Date of Submission12-Aug-2011
Date of Decision12-Nov-2011
Date of Acceptance06-Jan-2012
Date of Web Publication26-Sep-2012

Correspondence Address:
I Shah
Department of Pediatrics, Pediatric HIV Clinic, B. J. Wadia Hospital for Children, Mumbai, Maharashtra
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0022-3859.101375

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 :: Abstract 

Background: CD4 counts are a standard laboratory measure of disease progression in HIV-infected children. However, CD4 counting is done by flow cytometry and may not always be possible in every centre treating HIV-infected children in resource-limited countries. Absolute Lymphocyte Count (ALC) can be derived easily by performing a routine white blood cell count. The World Health Organization (WHO) in 2006 had recommended ALC to identify HIV-infected children in need of ART in resource-limited settings, when CD4 cell count is not available. Aims: This study aims to assess the reliability of using ALC as a marker for starting antiretroviral therapy (ART) in HIV-infected children in a tertiary hospital setting. Settings and Design: Retrospective analysis of 46 HIV-infected children who presented at a pediatric HIV clinic at a tertiary referral centre from 2002-2005. Materials and Methods: Using WHO 2006 guidelines for cutoff values of ALC and 2008 guidelines for CD4% as a comparative standard, a retrospective analysis was done on ART-naοve HIV-infected children who underwent baseline CD4% and ALC, and sensitivity and specificity of ALC was calculated. Statistical Analysis: Fischer exact two-tailed analysis was used to correlate ALC and CD4 and need for starting ART. Results: Sensitivity of ALC was 27.6% (72.4% were false negatives), specificity was 70.6%, with positive predictive value of 61.5%. On comparison across all clinical stages of disease, only 13/46 children (28.2%) would have been started on ART according to ALC cutoffs versus 29/46 children (63.04%) using CD4 criteria (P value=0.0015). In children with WHO clinical Stage 1 or 2 of disease, only 1/11 (9.1%) children were identified by ALC as requiring ART as opposed to 6/11 (54.5%) children by CD4% (P=0.0635). Conclusions: ALC is an unreliable marker to determine the need for starting ART in HIV-infected children.


Keywords: Absolute lymphocytosis count, CD4, antiretroviral therapy, children


How to cite this article:
Shah I, Parikh S. Reliability of absolute lymphocyte count as a marker to assess the need to initiate antiretroviral therapy in HIV-infected children. J Postgrad Med 2012;58:176-9

How to cite this URL:
Shah I, Parikh S. Reliability of absolute lymphocyte count as a marker to assess the need to initiate antiretroviral therapy in HIV-infected children. J Postgrad Med [serial online] 2012 [cited 2019 Nov 17];58:176-9. Available from: http://www.jpgmonline.com/text.asp?2012/58/3/176/101375



 :: Introduction Top


Currently there are more than 33.2 million adults and children living with HIV and AIDS. [1],[2] The number of people living with HIV/AIDS in India is secondary only to Nigeria and South Africa. CD4 estimation has been accepted as the most important laboratory parameter for deciding when to start antiretroviral therapy (ART) and to study progression of disease in children. [3],[4],[5] However, CD4 counting is done by flow cytometry and may not always be possible in every center treating HIV-infected children in resource-limited countries. There is need for a marker which is less resource-demanding on such countries as the greatest burden of the pandemic is borne by them. Absolute Lymphocyte Count (ALC) can be derived easily in resource-poor countries by performing a routine white blood cell count. The World Health Organization (WHO) in 2006 had recommended use of total lymphocyte count (TLC) to identify HIV-infected children in need of ART in resource-limited settings, [3] when CD4 cell count is not available. (In 2008 the WHO subsequently withdrew its recommendation on using ALC as a parameter for deciding initiation of therapy and the matter continues to be a subject of debate.) [6] Pediatric studies have found ALC as a powerful predictor of disease progression and mortality, however, its correlation with CD4 percentage is inconsistent. [7],[8] In contrast, a recent study in Africa found ALC as an unreliable marker in identifying immune-suppressed children. [9] Numerous papers with contrasting results in the adult population have been published. [10],[11],[12] However, sufficient research, especially focusing on the Indian pediatric population is unavailable. It is also known that the value of lymphocyte subsets obtained in Indian children is different from those of the other populations. [13] This study aims to test the reliability of using ALC as a parameter for deciding when to start ART in HIV-1 infected children in an Indian tertiary hospital setting.


 :: Materials and Methods Top


This is a retrospective analytical study done after ethical approval from the institutional academic committee. It aims to study the reliability of using ALC as a parameter to indicate the need for starting ART in children with HIV-1 infection. CD4% is used as a standard reference test. ART-naïve HIV-infected children less than or equal to eight years of age who presented to our pediatric and perinatal HIV clinic in the previous three years from 2002-2005 and underwent baseline CD4 and absolute lymphocyte testing from the same sample of blood were included in the study. Prior to 2002, CD4 and ALC were not done on the same blood sample and thus children before 2002 were not included in the study. Patients who could not afford CD4 estimation were not included in the study. Patients who did not have simultaneous CD4 and ALC from same blood sample were also excluded from the study. Thus, a total of 46 children who fit the inclusion criteria were included in this study. Age limits of the sample were decided thus since WHO had defined ALC values for decision-making only up to eight years of age. CD4 counting was done by Beckman EPICS XL-MCL flow cytometer and the cytostat tetraCHROME reagent containing CD45/CD8/CD4/CD3 monoclonal antibodies. The absolute lymphocyte count was obtained using the Coulter STK-S fully automated hematology analyzer. Age, sex and WHO clinical staging of each patient was also recorded. Need for initiating ART according to WHO clinical staging, ALC (WHO 2006 update) and CD4% (WHO 2008 update) was determined independently. WHO clinical staging 3 or 4, recommended thresholds of ALC, namely, a TLC of <4000 cells/mm3 for children aged <11 months, <3000 cells/mm3 for children aged 12-35 months, <2500 cells/mm3 for children aged 3-5 years, <2000/mm3 for children aged 5-8 years), and threshold CD4 levels for severe immunodeficiency (i.e. <20% for children aged 1 year to less than 5 years or <15% for children aged 5 years and above) were used as criteria for initiation of ART therapy. Also, according to 2008 guidelines all infants less than 1 year of age with proven HIV infection irrespective of CD4 levels are to be started on ART. (Note: The WHO 2008 update changed the CD4% criteria for starting ART in children aged 36 months to 59 months from <15% to <20%. The updated limits for CD4% were followed in this study.)

The children were divided into four age groups on the basis of WHO ART treatment recommendations, where Group 1 = <12 months, Group 2 = ≥1 year and <3 years, Group 3 = ≥3 years and <5 years and group 4 = ≥5 years and <8 years.

Statistical analysis

Sensitivity and specificity of ALC with CD4 estimation was calculated and association between ALC counts and CD4 estimation and need for ART were done by Fischer's Exact test. P value of <0.05 was taken as significant.


 :: Results Top


Male: Female ratio was 25:21. The mean age of patients was 4.22±2.37 years with 47.83% of children between five to eight years of age. The mean ALC was 3281±2128 mm3 and the mean CD4 count and CD4% were 660±562 mm3 and 17.9±12.5% respectively. Need for ART on the basis of immunological markers as per WHO 2008 guidelines, and therapy initiation guidelines for Absolute Lymphocyte count on the basis of WHO 2006 guidelines are depicted in [Table 1]. Sensitivity of ALC as compared to CD4 as reference standard was 0.276 (with 95% CI of (0.147 to 0.457); specificity was 0.706 (with 95% CI of (0.469 to 0.867) and positive predictive value was 0.615 (with 95% CI of (0.355 to 0.823) [Table 2]. Thus out of the 29 children that needed ART as per recommended standard CD4 test, the ALC test missed 21(72.4%) children, who were declared false negatives.
Table 1: Need for starting ART as per CD4%, ALC and WHO clinical staging

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Table 2: ALC predictive value when compared with CD4% and CD4 counts as gold standard

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Comparing data from all the children (n=46) included in our study, it was seen that only 13/46 children (28.2%) would have been started on ART according to ALC cutoffs versus 29/46 children (63.04%) would have benefited from ART using CD4 criteria (P value=0.0015) [Table 3].
Table 3: Comparing CD4 and ALC as parameters to decide initiation of ART across all WHO clinical stages

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In children with WHO clinical Stage 2 or 1 and data available of both CD4% and ALC values (n=11), it was seen that ALC identified only 1/11 (9.1%) children as requiring ART, whereas according to CD4 percentage 6/11 (54.5%) children should have received ART (P value=0.0635) Though not statistically significant, more than 45% of children were missed by using ALC alone in identifying immunosuppressed children with few clinical symptoms.

On comparison across age groups, it was seen that the specificity of the ALC test was 75% in the age group ≤3 years and sensitivity was 23%; whereas in the age group >3 years and ≤8 years, specificity was 77.8% and sensitivity 37.5%.


 :: Discussion Top


The reliability and use of ALC as a surrogate marker of immunity, comparable to CD4% and CD4 count has been debated. [10] The use of TLC as a predictor of CD4-cell count and mortality has been observed among HIV-infected children living in the US [8] and Europe [7] as well as among HIV-infected adults residing in the US, [14] India [15] and Africa. [16] The validity of these recommendations among children residing in resource-limited settings is limited, a recently published meta-analysis of approximately 2500 untreated HIV-infected children in resource-limited settings suggests TLC is a poor mortality predictor. [17] ALC measures both CD4 cells and other cells that provide functional immunity. Thus theoretically, it should be able to assess the disease progression in HIV-1 infection. In our study we found that compared to the recommended standard of CD4 testing, ALC had low sensitivity and moderate specificity. A large number of children were declared false negatives. These children would have missed out on necessary therapy if ALC had been used as an indicator of disease progression. In another pediatric study from northern Tanzania, according to WHO-recommended advance disease ALC cutoffs: sensitivity was 23.9% and specificity was 78.2%. [9] These results corroborate our findings. Also, when comparing data of all children across all WHO clinical stages it was seen that overall only 28.2% would have benefitted from ART according to ALC cutoffs versus 63.04% using CD4 cutoffs, which is a substantial difference.

According to the 2006 guidelines, in the absence of CD4 counts in children with WHO clinical Stage 1 disease, treatment was to be withheld and in children with WHO clinical Stage 2 of disease ALC could be used as a surrogate marker to decide when to start ART. In our study, percentage of false negatives in children with WHO clinical Stage 2 was a greatest at 80%. Thus ALC was not helpful in identifying immunosuppressed children with mild clinical presentations. Also, in the children in whom the value of the assays would be most critical (WHO clinical Stage 1 and 2 of disease) only 9% would have been assigned to start ART based on ALC as opposed to 54.5% if based on CD4%, a notable difference. However, since the total number of children in our study with Stage 1 and 2 disease was only 11, further evaluation is necessary.

Because ALC is generally high in children <18 months of age, its reliability as a prognostic indicator in younger children is low. [8] Sensitivity in our study was also higher in children greater than three years of age (37.5%) than in children less than three years of age (23%).

ALC estimates both T-cells and B-cells. In developed countries, T-lymphocyte percentages and B-lymphocyte percentages are stable in the first five years of life at around 65% and 24%, respectively, whereas in a study from Malawi, Africa T-lymphocyte percentages rose from 58% to 68%, and B-lymphocyte percentages fell from 30% to 22% during this period. Similar data in Indian children is lacking and the higher ALC in Indian children and non-correlation with CD4 may also be due to high B-lymphocyte subset and higher CD8 values. [13]

In conclusion, according to our study, low sensitivity makes ALC an unreliable marker of immune suppression in children. ALC does not corroborate with CD4% and does not improve the ability to identify mild clinical cases of children in need of ART. However, our study had limitations as it was a retrospective study and a very small number of patients were included in the study. Besides only those patients who could afford to do CD4 estimation were included and thus there was a selection bias. Thus, further studies in the Indian pediatric population are required.

 
 :: References Top

1.Rodrigo C, Rajapakse S. Current status of HIV/AIDS in South Asia. J Glob Infect Dis 2009;1:93-101.  Back to cited text no. 1
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2.UNAIDS/WHO AIDS Epidemic Update. Available from: http://www.unaids.org/en/KnowledgeCentre/HIVData/EpiUpdate/EpiUpdArchive/2007default.asp. [Last accessed on 2010 May 15].  Back to cited text no. 2
    
3.World Health Organization. Antiretroviral Therapy of HIV infection in infants and children: Towards Universal Access. Recommendations for a Public Health Approach (2006 Revision) Geneva: World Health Organization, 2006. Available from: http://www.who.int/hiv/pub/guidelines.pdf. [Last accessed on 2010 May 15].  Back to cited text no. 3
    
4.U.S. Department of Health and Human Services. Guidelines for the Use of Antiretroviral Agents in Pediatric HIV Infection. November 3, 2005. Available from: http://www.aidsinfo.nih.gov/Guidelines/GuidelineDetail.aspx?MenuItem=Guidelines&Search=Off&GuidelineID=8&ClassID=1. [Last accessed on 2010 May 15].  Back to cited text no. 4
    
5.Sharland M, di Zub GC, Ramos JT, Blanche S, Gibb DM, PENTA Steering Committee. PENTA guidelines for the use of antiretroviral therapy in pediatric HIV infection. Pediatric European Network for Treatment of AIDS. HIV Med 2002;3:215-26.  Back to cited text no. 5
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6.World Health Organization. Report of the WHO Technical Reference Group, Paediatric HIV/ART Care Guideline Group Meeting. Geneva: World Health Organization April 2008. Available from: http://www.who.int/hiv/pub/paediatric/WHO_Paediatric_ART_guideline_rev_mreport_2008.pdf. [Last accessed on 2010 July 22].  Back to cited text no. 6
    
7.Mofenson LM, Harris DR, Moye J, Bethel J, Korelitz J, Read JS, et al. Alternatives to HIV-1 RNA concentration and CD4 count to predict mortality in HIV-1-infected children in resource-poor settings. Lancet 2003;362:1625-7.  Back to cited text no. 7
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8.HIV Paediatric Prognostic Markers Collaborative Study. Use of total lymphocyte count for informing when to start antiretroviral therapy in HIV-infected children: A meta-analysis of longitudinal data. Lancet 2005;366:1868-74.  Back to cited text no. 8
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9.Johnson OO, Benjamin DK, Benjamin DK Jr, Schimana W, Tillekeratne GL, Crump JA, et al. Total lymphocyte count and World Health Organization pediatric clinical stage as markers to assess need to initiate antiretroviral therapy among human immunodeficiency virus-infected children in Moshi, Northern Tanzania. Pediatr Infect Dis J 2009;28:493-7.  Back to cited text no. 9
    
10.Ammann A, Burrowes S. Use of Total Lymphocyte Count vs CD4 Cell Count as a Marker of Immunity in HIV-Infected Adults and Children: Women, Children, and HIV. Available from: http://www.womenchildrenhiv.org/wchiv?page=tp-02-03. [Last accessed on 2010 May 15].  Back to cited text no. 10
    
11.Stebbing J, Sawleshwarkar S, Michailidis C, Jones R, Bower M, Mandalia S et al. Assessment of the efficacy of total lymphocyte counts as predictors of AIDS defining infections in HIV-1 infected people. Postgrad Med J 2005;81:586-8.  Back to cited text no. 11
    
12.Daka D, Loha E. Relationship between Total Lymphocyte count (TLC) and CD4 count among peoples living with HIV, Southern Ethiopia: A retrospective evaluation. AIDS Res Ther 2008;5:26.  Back to cited text no. 12
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13.Mandala WL, MacLennan JM, Gondwe EN, Ward SA, Molyneux ME, MacLennan CA. Lymphocyte subsets in healthy Malawians: Implications for immunologic assessment of HIV infection in Africa. J Allergy Clin Immunol 2010;125:203-8.  Back to cited text no. 13
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14.Jacobson MA, Liu L, Khayam-Bashi H, Deeks SG, Hecht FM, Kahn J, et al. Absolute or total lymphocyte count as a marker for the CD4 T lymphocyte criterion for initiating antiretroviral therapy. AIDS 2003;17:917-9.  Back to cited text no. 14
    
15.Kumarasamy N, Mahajan AP, Flanigan TP, Hemalatha R, Mayer KH, Carpenter CC, et al. Total lymphocyte count (TLC) is a useful tool for the timing of opportunistic infection prophylaxis in India and other resource-constrained countries. J Acquir Immune Defic Syndr 2002;31:378-83.  Back to cited text no. 15
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16.Mekonnen Y, Dukers NH, Sanders E, Dorigo W, Wolday D, Schaap A, et al. Simple markers for initiating antiretroviral therapy among HIV-infected Ethiopians. AIDS 2003;17:815-9.  Back to cited text no. 16
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17.Cross Continents Collaboration for Kids (3Cs4Kids) Analysis and Writing Committee. Markers for predicting mortality in untreated HIV-infected children in resource limited settings: A meta analysis. AIDS 2008;22:97-105.  Back to cited text no. 17
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    Tables

  [Table 1], [Table 2], [Table 3]



 

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