ABSTRACT Nutrient loss from the weathering process is a major challenge for tropical agriculture. Biochar with nutrient retention capacity has been proposed as an amendment to retain plant-available nutrients. Meanwhile, information on diazotrophic population responses to biochar application in the humid tropics is still poorly explored. A field study was carried out over three cropping cycles of maize in a typic paleudults of Peninsular Malaysia. During the first cropping cycle, the soil was amended with palm kernel shell biochar (PK), rice husk biochar (RH), palm kernel biochar with fertilizer (FPK), rice husk biochar with fertilizer (FRH), fertilizer (F), and control soil (C). Soil samples were taken at each harvesting stage and analyzed for pH, cation exchange capacity (CEC), total N, ammonium (NH4+–N), nitrate (NO3––N), microbial biomass N, and urease activity. Total and active diazotrophs were quantified from soil DNA and RNA, respectively, employing quantitative polymerase chain reaction (qPCR) amplification of the nifH gene. Palm kernel shell and rice husk biochars maintained a significant NH4+–N and NO3––N, respectively, during the second cropping cycle. Both biochars promote the total and active diazotrophic population. A detrimental impact on the nifH transcript was detected from fertilization even when biochar was co-amended. Two possible land management options for tropical soil were proposed from the overall data. First, the application of fertilizer with biochar can reduce N loss against weathering. Second, the application of biochar alone may improve biological N2 fixation in tropical soils.