A novel strain of Micrococcus sp. DUT_AHX, which was isolated from the sludge of a nitrobenzene (NB)-manufacturing plant and could utilize NB as the sole carbon source, was identified on the basis of physiological and biochemical tests and 16S ribosomal DNA (rDNA) sequence analysis. It can grow at the temperature up to 40 °C or in the presence of NaCl concentration up to 12 g/L in Luria-Bertani (LB) medium. The optimal degradation conditions are as follows: temperature 37 °C, pH 7.0, and shaking speed 150 r/min. The strain involves a partial reductive pathway due to the release of ammonia and can also utilize 2-aminophenol as the sole carbon source. Furthermore, the enzyme activity tests show that crude extracts of NB-grown strain DUT_AHX mainly contain 2-aminophenol 1, 6-dioxygenase activity. The exploitation of salt-tolerant bacteria will be a remarkable improvement in NB bioremediation and wastewater treatment at high salinity and high temperature.

The coupled effects of nitrogen source and methane monooxygenase (MMO) on the growth and poly-β-hydroxybutyrate (PHB) accumulation capacity of methanotrophs were explored. The ammonia-supplied methanotrophs expressing soluble MMO (sMMO) grew at the highest rate, while N2-fixing bacteria expressing particulate MMO (pMMO) grew at the lowest rate. Further study showed that more hydroxylamine and nitrite was formed by ammonia-supplied bacteria containing pMMO, which might cause their slightly lower growth rate. The highest PHB content (51.0%) was obtained under nitrogen-limiting conditions with the inoculation of nitrate-supplied bacteria containing pMMO. Ammonia-supplied bacteria also accumulated a higher content of PHB (45.2%) with the expression of pMMO, while N2-fixing bacteria containing pMMO only showed low PHB production capacity (32.1%). The maximal PHB contents of bacteria expressing sMMO were low, with no significant change under different nitrogen source conditions. The low MMO activity, low cell growth rate and low PHB production capacity of methanotrophs continuously cultivated with N2 with the expression of pMMO were greatly improved in the cyclic NO3− N2 cultivation regime, indicating that long-term deficiency of nitrogen sources was detrimental to the activity of methanotrophs expressing pMMO.Download high-res image (207KB)Download full-size image

A novel strain of Streptomyces sp. DUT_AHX was isolated from sludge contaminated with nitrobenzene and identified on the basis of physiological and biochemical tests and 16S ribosomal DNA (rDNA) sequence analysis. The optimal degradation conditions were as follows: temperature 30°C, pH 7.0–8.0, shaking speed 150–180 r/min, and inocula 10% (V/V). The strain, which possessed a partial reductive pathway with the release of ammonia, was also able to grow on mineral salts basal (MSB) medium plates with 2-aminophenol, phenol, or toluene as the sole carbon source. Furthermore, the enzyme activity tests showed crude extracts of nitrobenzene-grown DUT_AHX contained 2-aminophenol 1,6-dioxygenase activity. The 17-kb plasmid was isolated by the modified alkaline lysis method and was further cured by sodium dodecyl sulphate (SDS) together with 37°C. As a result, the cured derivative strain DUT_AHX-4 lost the 2-aminophenol 1,6-dioxygenase activity. The results suggested that the catabolic genes encoding the nitrobenzene-degrading enzymes were plasmid-associated. Moreover, the plasmid DNA was amplified with degenerate primers by touchdown PCR and an expected size fragment (471 bp) was generated. The Blast results revealed that the gene encoding a 157 amino acid polypeptide was 39%–76% identical to YHS domain protein. The further examination of the plasmid would demonstrate the molecular basis of nitrobenzene catabolism in Streptomyces, such as regulation and genetic organization of the catabolic genes.