We found that anemia was common in this cohort on second line ART. The prevalence was highest in women who also had lower hemoglobin values and more severe degrees of anemia. Being female and a CD4 count <200 cell/ul were risk factors for anemia while BMI ≥30 kg/m2 and being on ART for more than 10 years were associated with reduced risk of anemia.
Although ART use is associated with a decrease in the prevalence of anemia, the prevalence in our study population was high [1, 8, 18]. Among the general adult population in Malawi, the prevalence of anemia ranges from 17% in men to 28% in non-pregnant women [19, 20]. Compared to this, we report a higher prevalence of 33.2% in people on second line ART− 15.7% in men and 43.6% in women. The prevalence of anemia in this study is similar to the prevalence observed in people on first line ART regimens where it was reported at 38.2% in a study of people on first line ART regimens . However this prevalence is lower than the prevalence in ART naïve individuals which was reported at 77.4% in one study . This is consistent with studies that show that ART reduces the prevalence of anemia in people living with HIV [21, 22].
The high prevalence of anemia in our study population is still worrying and points to a larger problem— that anemia remains untreated in large numbers of people on ART. This is a concern because anemia lowers the quality of life, an important treatment goal while on ART, and increases the risk of progression to AIDS and death [4,5,6]. The high prevalence of anemia highlights the need to address causes of anemia in people on ART.
The mechanism of anemia in this study population as in other populations living with HIV is multifactorial and includes both HIV-related and non-HIV related causes. The HIV-related causes are HIV itself which causes anemia of chronic disease due to chronic inflammation; opportunistic infections such as mycobacteria and fungi and neoplasms such as lymphoma which infiltrate bone marrow and inhibit maturation of progenitor cells as well as medications used during treatment of HIV and associated conditions such as zidovudine and cotrimoxazole [3, 7]. Treating HIV-related anemia should be with effective ART; appropriate treatment for the opportunistic conditions and removal of any suspected medications [1, 7, 23]. In our study, a few anemic participants (13/377) had VL >1000 cps/ml and were suspected to have treatment failure while the rest of the participants had viral suppression. We did not have data on current and past medical history and drug history (other than the ART history). However since this was a very ART experienced cohort, it is possible that participants had a significant history of opportunistic conditions and drugs that can cause anemia.
Endemic causes of anemia play a role in both the general population and in people living with HIV. In sub-Saharan Africa, these are chronic malnutrition; nutritional deficiencies such as iron deficiency, vitamin B12 deficiency and folate deficiency; infections and parasite infestations such as malaria, schistosomiasis and hookworms; pregnancy and hemoglobin disorders such as thalassemia, other hemoglobinopathies and other rare congenital hematologic disorders [24,25,26]. In settings such as ours where the anemia prevalence is already high from endemic causes, HIV infection worsens pre-existing anemia through its effects of chronic inflammation and immunosuppression .
Despite iron deficiency being a common cause of anemia within African populations, there was no preponderance of microcytosis nor hypochromia in our study population to suggest that iron deficiency is an important cause of anemia in the study participants . Other studies have reported high rates of iron deficiency in people living with HIV and in those on ART . A possible reason for the low prevalence of iron deficiency anemia in our study population is that this was an urban population which is associatied with lower prevalence of iron deficiency that rural populations . In addition, the study participants were continuously in care at the HIV clinics with access to clinical assessments and hemoglobin level checks. The participants had many opportunities to be diagnosed with anemia and to have iron supplementation. The WHO recommends presumptive iron supplementation, anti-helminthic and anti-malarial therapy to treat anemia in resource limited settings where large workloads for health workers and inadequate laboratory capabilities make diagnosing specific causes of anemia difficult . This approach is recommended in the Malawi Standard Treatment Guidelines and in other African countries . However, a study in Mozambique showed that there is need for additional efforts to find and treat specific causes of anemia in PLHIV including in those on ART .
Nearly half of the female participants in this study had anemia, consistent with studies that show high prevalence of anemia in women living with HIV. Women living HIV are at a higher risk of severe anemia than men [4, 30]. The high prevalence in the women also reflects an overall higher prevalence of anemia in women in the general population . In addition, six out of the seven women that had severe anemia in this study had microcytosis that indicates iron deficiency anemia − all were given iron supplementation. In contrast, almost all men had mild anemia except for 4 cases with moderate anemia. Whether additional factors such as differential ART failure in women, contribution of anemia in failure of first line ART regimen or use of NNRTI for prevention of mother to child transmission (PMTCT) may have contributed to the high prevalence of anemia in women this study is not known. Women could benefit from routine screening for anemia before starting ART and routine hemoglobin monitoring while they are on ART.
There are several reasons for the high proportion of women in our study population. In Malawi, more women are infected with HIV than men; women tend to present for health services more frequently than men  and with the Option B+ program that has been implemented since 2011, pregnant women are systematically targeted for HIV testing and those who have HIV start life-long ART regardless of the CD4 cell count . As a result, more women are on ART than men. Our study population has similar gender demographics with the overall clinic population so that women are not overrepresented in the study population. From our findings, there was no difference in time spent on first line ART, time on second line ART and in virologic suppression between men and women.
We did not see an association between AZT in the current second line ART regimen and anemia in our analysis. This is in contrast with the widely recognized risk of anemia reported in association with AZT use [4, 31, 32]. We suspect that the lack of an association was because participants who would have been anemic at initiation of ART were not started on an AZT-based ART regimen and those that were suspected of having AZT associated anemia would have already been switched from AZT before these data were collected. Huffam et al. also reported that prior ART experience could be protective against subsequent development of AZT associated anemia in later regimens . This is relevant to this study population that had significant prior ART experience. Participants in this study spent an average of 7.4 years on ART. In addition, some studies have reported that AZT is not a significant risk factor for anemia when it is used as part of a combinational ART regimen as is the case in our study population [21, 29].
In this study, we described anemia in among an ART-experienced cohort on PI-based second line ART. A strength of this analysis is that it was conducted on a well-characterized cohort that is generalizable because it used data collected during routine clinic visits.
An important limitation of the study is the lack of data on other known risk factors for anemia such as the presence of fever, opportunistic infections (e.g. TB, oral candidiasis) and concurrent use of other medication which were not measured and were not included in the analysis . When analyzing the data on AZT, we were not able to look at substitutions that were made prior to the date of sample collection. This could have the effect of confounding by indication the significance of AZT in the ART regimens. Because Hb monitoring is not routinely done for patients on ART outside of the two facilities included in this study, the prevalence reported may be different than that in the wider HIV-infected population on second line ART.