An isolate of Rhizopus nigricans capable of tolerating and removing pentachlorophenol Araceli Tomasini*, VeroÂnica Flores, Diana CorteÂs and Javier Barrios-GonzaÂlez Depto. de BiotecnologõÂa, Universidad Auto Ânoma Metropolitana-Iztapalapa, Ado., Postal 55-535, 09340 MeÂxico, D.F. Mexico *Author for correspondence: Tel.: 58 04 64 63, Fax: 58 04 47 12, E-mail: atc@xanum.uam.mx Received 12 September 2000; accepted 21 February 2001 Keywords: Biodegradation, ®lamentous fungi, pentachlorophenol, Rhizopus nigricans, tolerance Summary Rhizopus nigricans, isolated from an industrial euent (paper mill), was resistant to pentachlorophenol (PCP) in Petri dishes and in submerged cultures (100 and 25 mg l )1 respectively). It was shown that this strain of R. nigricans can remove PCP in submerged culture. When 12.5 mg of PCP l )1 were added at 48 h, this compound had been completely removed by 144 h. Results indicated that the fungus did not produce extracellular lignin peroxidase (LiP) and laccase, but extracellular phenoloxidase production was observed. The synthesis of the latter enzyme was stimulated by the presence of PCP and/or tyrosine. These results indicate that this fungus, and probably other ®lamentous fungi, have an interesting potential to be used in processes for chlorophenol biodegradation. Introduction Pentachlorophenol (PCP) has been used as a wood preservative and pesticide. This compound is xenobiotic and recalcitrant and has been classi®ed as a priority pollutant in the USA. PCP is a common soil and wastewater pollutant (Lamar et al. 1990) and its remov- al is of great importance. Microbial processes have been used for this purpose and one of them studied for PCP degradation utilizes white rot fungi, particularly Phan- erochaete chrysosporium. This fungus has been studied widely (Mileski et al. 1988; Lin et al. 1989; Lamar et al. 1990; Lamar 1992), and it has been demonstrated that P. chrysosporium degraded and mineralized PCP from initial concentrations of 1±500 mg l )1 . Other white rot fungi that degrade PCP are Inonotus dryophilus, Peren- niporia medulla-panis, Trametes versicolor, Phellinus badius, Ganoderma oregonens and Lentinula edodes (Alleman et al. 1992; Okeke et al. 1996). The time for PCP degradation by white rot fungi was 12±60 days depending upon initial PCP concentration, fungal spe- cies, initial biomass, the mycelium age (Lamar et al. 1990; Alleman et al. 1992). The ability of a given microorganism to tolerate high contaminant concentra- tions would be an important advantage in increasing biodegradation eciency (Lamar et al. 1990). The above justi®es the importance of studing other fungi dierent from white rot fungi, which may grow faster and tolerate higher contaminant concentrations and therefore increase degradation eciency. There are few reports about PCP degradation by fungi other than white rot fungi. Cserjesi & Johnson (1971) reported PCP methylation by Trichoderma virgatum, Fahr et al. (1999) reported two brown rot fungi, Gloeophyllum striatum and G. trabeum, which degraded 2,4-dichlorophenol and PCP within 6 weeks. Seigle-Murandi et al. (1993) re- ported a large number of fungal strain belonging to dierent taxonomic groups such as zygomycetes, asco- mycetes, melaconiales, dematiaceae. All these fungi depleted PCP with dierent eciencies, and in some cases a correlation between PCP depletion and pheno- loxidase production was noticed. A wild Rhizopus sp. strain, resistant to PCP, was isolated from a paper mill euent (Tomasini et al. 1996), and was identi®ed as R. nigricans. In this work it is shown that R. nigricans is more resistant to PCP than some white rot fungi, both in plate culture and submerged culture. Moreover it is also shown that R. nigricans removes 62% of PCP, in 168 h in submerged culture (at 125 rev min )1 ), at initial concentrations of 12.5 mg l )1 . Materials and Methods Sensitivity test to PCP in plate culture Sensitivity to PCP by R. nigricans was determined on Petri dishes. The fungus was cultured with the synthetic medium of Melin-Norkrans containing (g l )1 ): glucose 5, malt extract 2, yeast extract 1, KH 2 PO 4 0.5, MgSO 4 á7H 2 0.15, (NH 4 ) 2 HPO 4 , agar 15, pH 5.0 World Journal of Microbiology & Biotechnology 17: 201±205, 2001. 201 Ó 2001 Kluwer Academic Publishers. Printed in the Netherlands.