There is a growing concern about antibiotic and metal resistance in natural and clinical settings. This study examined antibiotic and metal resistance for ancient Stenotrophomonas maltophilia isolated from permafrost. Whole-genome sequencing was applied to examine antibiotic resisrance genes and metal resistance genes. The permafrost S. maltophilia exhibited resistance to chloramphenicol, trimethoprim-sulfamethoxazole, and erythromycin, which is consistent with the presence of multidrug e ux pump smeDEF. The absence of qnrB, qnrR, oqxB, and smrA is consistent with its sensitivity for tetracycline, and cipro oxacin. Also, the absence of bcrABC is consistent with the strain's sensitivity to bacitracin. The strain also showed metal resistance to Zn 2+ , Cu 2+ , Cr 6+ , Hg 2+ , which is consistent with the presence of czcABCD, cueR and copA, chrR, merT, and merD. The strain showed worse resistance (lower MICs) to cipro oxacin, tetracycline, chloramphenicol, trimethoprim/sulfamethoxazole, and 4 metals Zn 2+ , Ni 2+ , Cu 2+ , and Cr 6+ , compared with clinical S. maltophilia; the former strain belongs to the lower ARG and MRG gene clusters compared to latter one. Among 12 genomic islands (GIs) in the permafrost strain, 5 contained resistance genes, with only one kind of ARG or MRG, while S. maltophilia from other environments exhibited a minimum of 2 kinds of resistance gene clustering on a single GI. This study demonstrates that continuous exposure to the clinical environment can create a selective pressure on bacteria inducing resistance to antibiotics and metals, involving more ARGs and MRGs via horizontal gene transfer route.