Characterization of methyl parathion degradation by a Burkholderia zhejiangensis strain, CEIB S4-3, isolated from agricultural soils.

Through the use of an enrichment technique, we isolated from the agricultural soils of Morelos in central México a strain of Burkholderia zhejiangensis identified as CEIB S4-3, it's could use the pesticide methyl parathion (MP) as the only source of carbon and degrade completely p-nitrophenol (PNP). For more efficient MP and PNP degradation by the CEIB S4-3 strain, the absence of an extra carbon source, a large inoculum and an MP concentration up to 50 mg/l are required. Sequence and annotation analysis of the draft genome, showed presence of mpd functional gene, which was expressed and its activity on the MP was confirmed. Additionally, the genes coding for enzymes in the benzoquinone pathway (conducted by Gram-negative bacteria) and the benzenotriol pathway (conducted by Gram-positive bacteria) were found, which was corroborated by identification of intermediary metabolites by HPLC. Thus, we propose that B. zhejiangensis CEIB S4-3 uses both degradation pathways.

Isolation of the opdE gene that encodes for a new hydrolase of Enterobacter sp. capable of degrading organophosphorus pesticides.

Microbial enzymes that can hydrolyze organophosphorus compounds have been isolated, identified and characterized from different microbial species in order to use them in biodegradation of organophosphorus compounds. We isolated a bacterial strain Cons002 from an agricultural soil bacterial consortium, which can hydrolyze methyl-parathion (MP) and other organophosphate pesticides. HPLC analysis showed that strain Cons002 is capable of degrading pesticides MP, parathion and phorate. Pulsed-field gel electrophoresis and 16S rRNA amplification were performed for strain characterization and identification, respectively, showing that the strain Cons002 is related to the genus Enterobacter sp. which has a single chromosome of 4.6 Mb and has no plasmids. Genomic library was constructed from DNA of Enterobacter sp. Cons002. A gene called opdE (Organophosphate Degradation from Enterobacter) consists of 753 bp and encodes a protein of 25 kDa, which was isolated using activity methods. This gene opdE had no similarity to any genes reported to degrade organophosphates. When kanamycin-resistance cassette was placed in the gene opdE, hydrolase activity was suppressed and Enterobacter sp. Cons002 had no growth with MP as a nutrients source.

Characterization of cellulolytic activities of Bjerkandera adusta and Pycnoporus sanguineus on solid wheat straw medium / Caracterización de las actividades celulolíticas de Bjerkandera adusta y Pycnoporus sanguineus en un medio sólido de paja de trigo

Cellulolytic properties of two white rot fungi, Bjerkandera adusta and Pycnoporus sanguineus, cultivated on wheat straw agar medium, were characterized and compared. Optimal growing parameters for maximum enzyme production for both fungi were wheat straw medium pH 5 and 28ºC. B. adusta showed, on the 6th day of culture, carboxymethylcellulose (CMC)ase activity levels 1.6 times higher than maximal P. sanguineus activity, achieved on the 8th day. B. adusta supernatants also displayed higher activity levels towards xylan (3.6-fold) compared to those of P. sanguineus. However, enzymes from P. sanguineus were more robust resisting one hour incubation at high temperatures (up to 80ºC), and exhibiting activity and stability in pH range from 2 to 8. Cellulolytic activities, with molecular masses ranging from 25 to 90 kDa, from the two species were detected in zymograms.

Production of two novel laccase isoforms by a thermotolerant strain of Pycnoporus sanguineus isolated from an oil-polluted tropical habitat.

A thermotolerant and halotolerant strain of Pycnoporus sanguineus was isolated from an oil-polluted site in a tropical area located in Veracruz, Mexico. This strain was able to grow at 47 degrees C and in culture medium containing 500 mM NaCl. The strain was also tolerant to the presence of 30,000 ppm of crude Maya oil. A 68-kDa protein purified from submerged cultures exhibited laccase activity towards 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS), guaiacol, syringaldazine, and o-dianisidine, for which it presented the highest affinity (Km = 43 microM). Two-dimensional gel electrophoresis analysis showed that, unusual for laccases, the enzyme has two active isoforms, with isoelectric points of 7.00 and 7.08. The purified enzyme showed high thermostability, retaining 40% of its original activity after 3 h at 60 degrees C. This property seems to correlate with a long "shelf-life," given that at 40 degrees C enzyme activity was only gradually lost over a 5-day period incubation. Both the fungus and its laccase are likely to have high potential for biotechnological applications.