Hematological complications are documented as the second most common cause of morbidity and mortality in HIV/AIDS patients. Drugs for HIV/AIDS showed side effects on different hematological parameters. This study was aimed at assessing the changes in hematological profile of HIV infected pregnant women after initiation of highly active antiretroviral treatment. Retrospective cohort study was conducted to assess the hematological changes in HIV infected pregnant women after initiation of highly active antiretroviral (HAART) at ART clinic of Debremarkos hospital in Ethiopia from May to August 2017. Data available between May 2008 and August 2017 was collected from 380 patient cards of pregnant women. The data was entered into Excel spread sheet and was transported into STATA software for analysis. All data were presented as mean ± Standard error of the mean. Comparisons were made using the Paired T-test. A p-value of <0.05 was considered as a statistically significant difference. The result from the present study shows that the HAART initiation in pregnant women with HIV increases the CD4 cell count, the mean corpuscular hemoglobin concentration, white blood cell count, lymphocyte count and total mid cell count at ninety-five percent confidence interval. From the total of 380 patient cards with complete CD4 count, 200 patient cards were recorded with TDF/3TC/EFV ART regimen. The remaining 180 patient cards were documented with AZT/3TC/NVP regimen. Comparing the effect of the two Regimens on CD4 count, TDF/3TC/EFV increases the CD4 count significantly in pregnant women with HIV (p<0.0001). The other regimen (AZT/3TC/NVP) also increases the CD4 count from initial value but this increase in CD4 count after AZT/3TC/NVP initiation in pregnant women with HIV is not statistically significant (P>0.05). HAART initiation in pregnant women with HIV increases lymphocyte count, total mid cell count, red blood cell count, white blood cell count, total hemoglobin, hematocrit, mean cell volume and total platelet. On the contrary, the initial value of granulocyte count, mean cell hemoglobin, mean corpuscular hemoglobin concentration, red cell distribution width and mean platelet volume decreases after antiretroviral treatment in pregnant women with HIV.