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Most current researches focus solely on reducing soil chromium availability. It is difficult to reduce soil Cr(VI) concentration below 5.0 mg kg-1 using single remediation technology. This study introduced a sustainable soil Cr(VI) reduction and stabilization system, Penicillium oxalicum SL2–nanoscale zero-valent iron (nZVI), and investigated its effect on Cr(VI) reduction efficiency and microbial ecology. Results showed that P. oxalicum SL2–nZVI effectively reduced soil total Cr(VI) concentration from 187.1 to 3.4 mg kg–1 within 180 d, and remained relatively stable at 360 d. The growth curve of P. oxalicum SL2 and microbial community results indicated that γ-ray irradiation shortened the adaptation time of P. oxalicum SL2 and facilitated its colonization in soil. P. oxalicum SL2 colonization activated nZVI and its derivatives, and increased soil iron bioavailability. After restoration, the negative effect of Cr(VI) on soil microorganisms was markedly alleviated. Cr(VI), Fe(II), bioavailable Cr/Fe, Eh, EC and urease (SUE) were the key environmental factors of soil microbiota. Notably, Penicillium significantly stimulated the growth of urease-positive bacteria, Arthrobacter , Pseudarthrobacter , and Microvirga , synergistically reducing soil chromium availability. The combination of P. oxalicum SL2 and nZVI is expected to form a green, economical and long-lasting Cr(VI) reduction stabilization strategy. [Display omitted] • P. oxalicum SL2–nZVI effectively reduced soil total Cr(VI) and available chromium. • Gamma-ray irradiation pretreatment facilitated P. oxalicum SL2 colonization in soil. • The negative effect of Cr(VI) on soil microorganisms was mitigated after remediation. • Cr- and Fe- related indicators, Eh, EC and SUE were the key environmental factors. • Penicillium colonization significantly stimulated the growth of urease-positive bacteria. [ABSTRACT FROM AUTHOR]