Anthropogenic habitat fragmentation reduces gene flow, often resulting in small, isolated populations with decreased genetic diversity. These impacts can make populations more susceptible to extinction through environmental and genetic stochasticity. The San Joaquin kit fox (Vulpes macrotis mutica; SJKF) is endemic to California and is currently listed as federally endangered. The desert kit fox (Vulpes macrotis ssp.; DKF) occurs throughout most arid regions of the western United States and contains multiple recognized subspecies. Little is known about the status of DKF but SJKF numbers are known to have declined from historical levels. Gene flow between SJKF and DKF has not been investigated. Using 18 nuclear microsatellite loci, we conducted a preliminary investigation of these questions. We identified lower heterozygosity and allelic richness in SJKF (n = 34; He = 0.54 ± SE = 0.05) than DKF (n = 31; He = 0.64 ± 0.06). We observed high genetic distance between the SJKF and DKF (FST = 0.15). Admixture analyses provided no significant evidence of contemporary gene flow between SJKF and DKF. Future research using Mojave samples from closer to the San Joaquin Valley and higher-resolution genomic markers will enable more definitive conclusions.