id kits: Determine and Unigold. CD4+ve T cell counts of patients were determined with the Cyflow counter method (Partec, Germany). After an informed consent, 2 - 3 millilitres of whole venous blood was collected from the patient into an ethylenediaminetetraacetic acid (EDTA) containing bottle. The sample was allowed to settle in the refrigerator at the laboratory of the HIV clinic of the Lagos State University Teaching Hospital (LASUTH), South west Nigeria. Plasma was pipette from samples into cryovials and screened for CrAg using WHO recommended Lateral Flow Assay (IMMY, USA) which is a rapid point of care dipstick assay for the diagnosis of cryptococcal disease. The simple to use assay was used for running this screening while strictly observing the manufacturer’s instructions. The point of care Lateral Flow Assay (LFA) has 100% sensitivity and 94% specificity. The LFA was performed in 5 easy steps and objective qualitative results were obtained within 10 minutes. The presence of two visible bands on the dipstick indicates a positive result on the test plasma which is in keeping with the manufacturer’s positive control. The qualitative test result was interpreted as nonreactive (negative) if it showed only one visible band on the dipstick. Inclusion criteria for subjects in this study were age 18 years and above, freshly diagnosed and confirmed positive for HIV, just enrolled into care and treatment naive. The exclusion criteria were age less than 18 years, not confirmed for HIV and treatment experienced. Obtained data was analysed using the Statistical package for Social Sciences (SPSS) version 15.0 software to generate frequencies percentages. P value ≤ 0.05 was considered statistically significant in group comparisons. Results were presented in tables.
Four hundred and thirty two confirmed HIV patients were tested for cryptococcal antigen. There were 184 (42.6%) men and 248 (57.4%) women studied. Seven (1.6%) of the subjects were positive for cryptococcal antigen. The WHO clinical stage of studied patients was; stage I 163 (37.7%), stage II 132 (30.6%) stage III 95 (22.0%) and stage IV 42 (9.7%). The age of the subjects ranged from 18 - 75 years with a mean of 38.15 ± SD 9.53. Majority 196 (45.4%) of the subjects were in the age group 29 - 39 years. Two hundred and sixty six (62%) had CD4+ve T cell count less than 100 cells/ul, 100 (23.3%) between 100 and 199 cells and 63 (14.7%) had above 200 cells/ul as in Table 1. In Table 2, majority 4 (57.1%) of cryprococcal antigen positive subjects were in the age group 29 - 39 years and were all females. Most 5 (71.4%) of the CrAg+ve subjects had secondary education. Six (85.7%) of CrAg positive patients had CD4+ve T cell count below 100 cells/ul while one (14.3%) had count above 200 cells. Three (42.9%) of the CrAg positive patients were in WHO clinical stage 1 while 4 (57.1%) were in stage II with none in stages III and IV. None of the CrAg positive patients had active TB as in Table 3. Mean CD4 count was112 (range 06 - 1264) cells/ul and median was 74.
Our study examined the prevalence of cryptococcal antigenaemia and its relation to CD4+ve T cell counts and WHO clinical staging among treatment naïve HIV positive Nigerian adults. We observed a cryptococcal antigenaemia rate of 1.6%. Our observed overall low cryptococcal antigenaemia of 1.6% among treatment naïve HIV positive patients is comparable with previous reports from Shika in Nigeria 2.2 %  , Tanzania 3%  and USA 2.9%  . However our overall prevalence value in this study is lower than some reports from Nnewi 13.1%  South east, and Benin12.7%  South south part of Nigeria as well as Kenya 11%  , Uganda 5.7%  , Indonesia 7.1%  and Thailand 9.2%  . The observed differences in prevalence rates of cryptoccocal antigen positivity among HIV infected persons in the various studies cited may be explained by differences in sample
Table 1. Sociodemographic characteristics, WHO clinical staging, CD4+ve T cell levels and cyrptococcal antigen status of patients.
Table 2. Age group and demographic distribution of cryptococcal antigen among HIV positive patients.
Table 3. CD4+ve T cell count, WHO clinical stage and TB status of cryptococcal antigen positive patients.
sizes as well as varying sensitivities and specificities with differing methodologies of used kits. We observed in our study that the majority 6 (85.7%) of the CrAg positive patients had CD4+ve T cell count less than 100 cells/ul. This finding of 85.7% CrAg positivity among HIV patients who were severely immunocomprised with CD+veT cell ≤ 100 cells/ul corroborates previous research reports  - . Low CD4+ve T cell level has been found to be the only factor significantly associated with positive serum cryptococcal antigenaemia   . The median CD4+ve T cell count among the subjects in our study was 74 cells/ul. This finding is comparable with previous figure of 62  and 60  in previous researches. A previous study revealed that the median CD4+ve T cell count was also reportedly lower in CrAg positive HIV infected patients compared with the CrAg negative  . It has also been found that the lower the CD4+ve T cell count, the higher the occurrence of cryptoccocal meningitis in HIV infected patients  . Our study is significant in the prevention of cryptococcal meningitis, as it has been shown that the severely immunocompromised HIV patient with CD4+ve T cells ≤ 100 needs to be screened for CrAg and subsequently placed on pre-emptive treatment depending on the outcome of the tests. We also found out in our study that the WHO clinical stages of CrAg positive patients were 1 and 11 with 3 (42.9%) and 4 (57.1%) respectively. None of the CrAg positive HIV infected patients in this study had active tuberculosis. In Uganda, tuberculosis has been reported to be a frequent co-pathogen in patients with cryptococcemia with a 31% concurrent infection among CrAg positive HIV infected patients  . Also in Cape Town, South Africa, a substantial proportion of patients with cryptococcal meningitis were noted to have a history of tuberculosis  . We observed a mean age of 38.1 years among the patients, this is compatible with 38.5 years from a previous study  . More female patients 248 (57.4%) than males 184 (42.6%) were enrolled for our study and all the CrAg positive subjects were female 7 (100%). This is contrary to previous reports from studies where more males were enrolled and also found CrAg positive   . Most of the patients in our study were in age group 29 - 39 years and this observation is similar to findings in a previous research with 24 - 40 years  . The detection of cryptococcal antigen in the serum (CrAg) is clinically relevant and precedes the onset of overt disease as previously documented  . Early diagnosis of CM would translate into a reduction in CM-related deaths because patients who receive early antifungal treatment have been shown to have better outcomes than those who receive delayed treatment  . Considering the substantial mortality and morbidity associated with CM, preventive interventions should be prioritized. CRAG screening directed at all newly diagnosed HIV-positive patients with CD4+ T-cell counts < 100 cells/μl is likely to detect most positive cases who are at risk of developing cryptococcal meningitis with high mortality  . Treatment of patients with isolated cryptococcal antigenemia has been shown to prevent the development of CM and CM-related death  . Prevention of symptomatic CM through early cryptococcal disease detection and pre-emptive treatment should be our focus in Africa as it is cost effective and would reduce mortality. Prevention of the poor outcomes of cryptococcosis through routine screening for serum cryptococcal antigen, with early disease detection and prompt treatment, could be a practical approach in resource-limited settings.
Our study revealed an overall cryptococcal antigenaemia prevalence of 1.6% among HIV infected adult patients in a Lagos hospital, of which 85.7% was severely immunocompromised with CD4+ve Tcell count ≤ 100. Also, we found that WHO clinical stage was not a predicting risk factor for cryptococcal antigenaemia in HIV infection. We recommend the use of point of care test kits which are easy to administer and have excellent sensitivity and specificity and also need no expertise or laboratory infrastructure for the screening of severely immunocompromised patients in resource limited settings such as ours. Targeted screening and subsequent treatment would reduce morbidity and mortality from cryptococcal meningitis among HIV infected patients in our setting.
The authors would like to thank the HIV Clinic Laboratory and Nursing staff for their cooperation during patient recruitment, blood sample collection and rapid testing in this study. We also appreciate IMMY for the donation of CrAg LFA dipsticks which are used for this study.
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