Bioaugmentation of Activated Sludge by an Indigenous 3-Chloroaniline-Degrading <i>Comamonas testosteroni</i> Strain, I2 <i>gfp</i>

TLDR

Comamonas testosteroni I2, isolated from activated sludge, can mineralize 3‑chloroaniline via a meta‑cleavage pathway that transiently accumulates a yellow chlorocatechol intermediate. The study aimed to evaluate whether inoculating this strain into activated sludge could remediate 3‑chloroaniline‑contaminated wastewater. The strain was chromosomally tagged with gfp (I2‑gfp) and introduced into a lab‑scale semicontinuous activated‑sludge reactor, where its persistence and the reactor microbial community were monitored by DGGE of 16S rRNA genes. I2‑gfp persisted for at least 45 days and achieved complete 3‑chloroaniline degradation within two weeks after a six‑day adaptation, but subsequent operation only removed 50 % of the contaminant, and DGGE revealed a distinct community shift, indicating that bioaugmentation requires regular resupplementation.

Abstract

ABSTRACT A strain identified as Comamonas testosteroni I2 was isolated from activated sludge and found to be able to mineralize 3-chloroaniline (3-CA). During the mineralization, a yellow intermediate accumulated temporarily, due to the distal meta -cleavage of chlorocatechol. This strain was tested for its ability to clean wastewater containing 3-CA upon inoculation into activated sludge. To monitor its survival, the strain was chromosomally marked with the gfp gene and designated I2 gfp . After inoculation into a lab-scale semicontinuous activated-sludge (SCAS) system, the inoculated strain maintained itself in the sludge for at least 45 days and was present in the sludge flocs. After an initial adaptation period of 6 days, complete degradation of 3-CA was obtained during 2 weeks, while no degradation at all occurred in the noninoculated control reactor. Upon further operation of the SCAS system, only 50% 3-CA removal was observed. Denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes revealed a dynamic change in the microbial community structure of the activated sludge. The DGGE patterns of the noninoculated and the inoculated reactors evolved after 7 days to different clusters, which suggests an effect of strain inoculation on the microbial community structure. The results indicate that bioaugmentation, even with a strain originating from that ecosystem and able to effectively grow on a selective substrate, is not permanent and will probably require regular resupplementation.