Trichloroethylene (TCE) is a prevalent groundwater contaminant detected worldwide, and microbes are sensitive indicators and initial responders to these chemical contaminants causing disturbances to their ecosystem. In this study, microbes isolated from San Marcos River sediment were screened for their TCE degradation potential. Among the twelve isolates (SAN1-12), five isolates demonstrated TCE degradation within 5 days at 25 °C and 40 mg/L of TCE concentration in the following order: SAN8 (87.56%), SAN1 (77.31%), SAN2 (76.58%), SAN3 (49.20%), and SAN7 (3.36%). On increasing the TCE concentration to 80 mg/L, the degradation efficiency of these isolates declined, although SAN8 remained the prominent TCE degrader with 75.67% degradation. The prominent TCE-degrading isolates were identified as Aeromonas sp. SAN1, Bacillus sp. SAN2, Gordonia sp. SAN3, and Bacillus proteolyticus SAN8 using 16S rRNA sequencing. The TCE degradation and cell biomass of Bacillus proteolyticus SAN8 were significantly improved when the incubation temperature was increased from 25 °C to 30 °C. However, both slightly acidic and alkaline pH levels, as well as higher TCE concentrations, lowered the efficacy of TCE degradation. Nevertheless, these conditions led to an increase in bacterial cell biomass.