Degradation of organic nitrogenous wastes by a soil streptomycete

A soil streptomycete degraded hair, silk, wool, feather and leather which were collected from solid wastes. The organism was identified taxonomically and designated Streptomyces sp. A956. It degraded leather to the maximum extent and solubilized 35.9% of the total nitrogen, 2.32 mg of glycine equivalent amino nitrogen could be obtained by degradation of 100 mg leather.     

Activity of North Jordan soil streptomycete isolates against Candida albicans

Eighteen percent of 116 different isolates of Streptomyces recovered from soils of northern Jordan showed activity against Candida albicans. The recovered isolates were distributed into three groups according to the diameter of the inhibition zone on the agar plate: group 1 (5–10 mm, slightly active); group 2 (11–15 mm, moderately active); and group 3 (16–35 mm, highly active). Isolates of group 3 were further grouped into four sub-groups and were culturally and morphologically identified. The u.v. spectra of the fermentation broth for the isolates in sub-group 4 were determined, and showed absorbance peaks ranging between 230 and 300 nm.     

Characterization and antimicrobial activity of the bioactive metabolites in streptomycete isolates

Twenty different streptomycete isolates were obtained from soils of southeast Serbia. Five isolates identified as Streptomyces hygroscopicus (SH100, SH101, SH102, SH103, and SH104) showed strong activity against Botrytis cinerea, a parasite found in domestic vines. These isolates were extensively studied for their in vitro antimicrobial activity against Gram-positive bacteria, Gram-negative bacteria, yeasts and fungi, and also antiviral activity against Herpes simplex. The results indicated that the obtained isolates were highly active against Botrytis cinerea, Candida albicans, and Herpes simplex, with an inhibition zone at ≥31 mm. The structure of the bioactive components was determined using elemental analysis, as well as UV/VIS, FTIR, and TLC.     

Characterization and antimicrobial activity of the bioactive metabolites in streptomycete isolates

Twenty different streptomycete isolates were obtained from soils of southeast Serbia. Five isolates identified as Streptomyces hygroscopicus (SH100, SH101, SH102, SH103, and SH104) showed strong activity against Botrytis cinerea, a parasite found in domestic vines. These isolates were extensively studied for their in vitro antimicrobial activity against Gram-positive bacteria, Gram-negative bacteria, yeasts and fungi, and also antiviral activity against Herpes simplex. The results indicated that the obtained isolates were highly active against Botrytis cinerea, Candida albicans, and Herpes simplex, with an inhibition zone of approximately 31 mm. The structure of the bioactive components was determined using elemental analysis, as well as UV/VIS, FTIR, and TLC.     

Degradation of humic acid of a forest soil by some fungal isolates

Summary In a laboratory incubation study the humic acid isolated from a forest soil of Palamau (Bihar) was subjected to biodegradation for a period of six weeks by using nine cultures of fungi. These fungi were tested earlier for their cellulose decomposing ability. The humic acid was used as sole source of carbon, nitrogen and carbon plus nitrogen in Czapek-Dox broth. Of the nine culturesAspergillus awamori (IARI),Penicillium sp. (Ranchi),Humicola insolense (Hissar) were found to be very effective in decomposing humic acid. The humic acid used as sole source of carbon was most efficiently degraded followed by that used as carbon+nitrogen source. When it was used as sole source of nitrogen, it could not be degraded so efficiently. This may be due to unavailability of its nitrogen to these microorganisms.     

A keratin-decomposing streptomycete

A proteolytic actinomycete was isolated from an Indian soil sample. It degraded hair, silk, wool and feather. Protease activity was reported for growth of the organism on these keratin substrates. The organism was taxonomically studied and designated as Streptomyces sp. S₇.     

Proline hydroxylation by cell free extract of a streptomycete

Free L-proline was hydroxylated to free L-hydroxyproline by cell free extract of Streptomyces griseoviridus P8648. The hydroxylation reaction required ferrous ion, 2-ketoglutarate and ascorbate. Zinc ion, ethylenediaminetetraacetic acid and alpha,alpha'-dipyridyl inhibited the reaction. Optimum temperature and pH were 25.0 degrees C and 7.5, respectively.     

Degradation of the metal-cyano complex tetracyanonickelate(II) by cyanide-utilizing bacterial isolates

Ten bacterial isolates capable of growth on tetracyanonickelate(II) [K2[Ni(CN)4]] (TCN) as the sole nitrogen source were isolated from soil, freshwater, and sewage sludge enrichments. Seven of the 10 were identified as pseudomonads, while the remaining 3 were classified as Klebsiella species. A detailed investigation of one isolate, Pseudomonas putida BCN3, revealed a rapid growth rate on TCN (generation time, 2 h), with substrate removal and growth occurring in parallel. In addition to TCN, all isolates were able to utilize KCN, although the latter was significantly more toxic; MICs ranged from 0.2 to 0.8 mM for KCN and greater than or equal to 50 mM for TCN. While growth occurred over a wide range of TCN concentrations (0.25 to 16 mM), degradation was most substantial under growth-limiting conditions and did not occur when ammonia was present. In addition, cells grown on TCN were found to accumulate nickel cyanide [Ni(CN)2] as a major biodegradation product. The results show that bacteria capable of growth on TCN can readily be isolated and that degradation (i) appears to parallel the capacity for growth on KCN, (ii) does not occur in the presence of ammonia, and (iii) proceeds via the formation of Ni(CN)2 as a biological metabolite.     

Degradation of mono- and dichlorobenzoic acid isomers by two natural isolates of Alcaligenes denitrificans

Two strains of Alcaligenes denitrificans, designated BRI 3010 and BRI 6011, were isolated from polychlorinated biphenyl (PCB)-contaminated soil using 2,5-dichlorobenzoic acid (2,5-DCBA) and 2,4-DCBA, respectively, as sole carbon and energy sources. Both strains degraded 2-chlorobenzoic acid (2-CBA), 2,3-DCBA, and 2,5-DCBA, and were unable to degrade 2,6-DCBA. BRI 6011 alone degraded 2,4-DCBA. Growth of BRI 6011 in yeast extract and 2,6-DCBA induced pyrocatechase activity, but 2,6-DCBA was not degraded, suggesting the importance of an unsubstituted carbon six of the aromatic ring. Metabolism of the chlorinated substrates resulted in the stoichiometric release of chloride, and degradation proceeded by intradiol cleavage of the aromatic ring. Growth of both strains on 2,5-DCBA induced pyrocatechase activities with catechol and chlorocatechols as substrates. In contrast to dichlorobenzoic acids, growth on 2-CBA, benzoic acid, mono- and dihydroxybenzoic acids induced a pyrocatechase activity against catechol only. Although 2,4-DCBA was a more potent inducer of both pyrocatechase activities, its utilization by BRI 6011 was inhibited by 2,5-DCBA. Specific uptake rates using resting cells were highest with 2-CBA, except when the resting cells had been previously grown on 2,5-DCBA, in which case 2,5-DCBA was the preferred substrate. The higher rates of 2,5-DCBA uptake obtained by growth on that substrate, suggested the existence of a separately induced uptake system for 2,5-DCBA.     

Keratinase of a streptomycete.

Keratinase produced from Streptomyces Sp.A11 decomposed human hair, chicken feather, wool, silk and pure keratin extracted from human epidermis. Purification of the enzyme by DEAE-cellulose column chromatography resulted in 7.5-fold increase in activity relative to the activity of the culture filtrate. The enzyme was inducible, extracellular, homogeneous with a molecular weight of 49,000. The enzyme activity was inhibited by reduced glutathione, phenylmethyl sulphonyl fluoride and 2-mercaptoethanol.     

Purification and characterization of a streptomycete collagenase

A soil streptomycete designated as Streptomyces sp. A₈ produced an extracellular collagen hydrolysing enzyme that appeared to be 'true collagenase'as it degraded native collagen under physiological conditions and cleaved the synthetic hexapeptide 4-phenylazobenzyloxycarbonyl-L-prolyl-L-leucyl-glycyl-L-prolyl-D-arginine into two tripeptides. The enzyme was purified by diethyl aminoethyl cellulose chromatography and Sephadex G-150 gel filtration. The purified enzyme had an apparent molecular weight of about 75000 by SDS-polyacrylamide gel electrophoresis. Treatment with lithium chloride did not dissociate it into subunits. A strong inhibition was observed with chelating agents such as α-α-dipyridyl and 8-hydroxyquinoline. Ethylene diamine tetracetate completely inhibited the enzyme activity. Among the cations tested only Ca^{2 +} and Mg^{2 +} enhanced the collagenase activity. Heavy metal ions like Pb^{2 +}, Ag⁺, Cu^{2 +} and Zn^{2 +} strongly inhibited the enzyme. The EDTA inhibition could be reversed with Ca^{2 +}. Cysteine and reduced glutathione caused significant reduction in enzyme activity. Parachloromercuribenzoate and iodoacetamide had no effect on the collagenase. Amino acid analysis revealed the absence of cysteine and tyrosine. Many of the properties were the same as collagenases of Clostridium histolyticum and Vibrio alginolyticus.     

Purification and characterization of a streptomycete collagenase

A soil streptomycete designated as Streptomyces sp. A8 produced an extracellular collagen hydrolysing enzyme that appeared to be 'true collagenase' as it degraded native collagen under physiological conditions and cleaved the synthetic hexapeptide 4-phenylazobenzyloxycarbonyl-L-prolyl-L-leucyl-glycyl-L-prolyl-D-a rginine into two tripeptides. The enzyme was purified by diethyl aminoethyl cellulose chromatography and Sephadex G-150 gel filtration. The purified enzyme had an apparent molecular weight of about 75,000 by SDS-polyacrylamide gel electrophoresis. Treatment with lithium chloride did not dissociate it into subunits. A strong inhibition was observed with chelating agents such as alpha-alpha-dipyridyl and 8-hydroxyquinoline. Ethylene diamine tetraacetate completely inhibited the enzyme activity. Among the cations tested only Ca2+ and Mg2+ enhanced the collagenase activity. Heavy metal ions like Pb2+, Ag+, Cu2+ and Zn2+ strongly inhibited the enzyme. The EDTA inhibition could be reversed with Ca2+. Cysteine and reduced glutathione caused significant reduction in enzyme activity. Parachloromercuribenzoate and iodoacetamide had no effect on the collagenase. Amino acid analysis revealed the absence of cysteine and tyrosine. Many of the properties were the same as collagenases of Clostridium histolyticum and Vibrio alginolyticus.     

Expression of streptomycete cholesterol oxidase inEscherichia coli

Summary A streptomycete gene coding for extracellular cholesterol oxidase (choA) was subcloned and expressed inEscherichia coli. The pUCO series recombinants were obtained by inserting thechoA gene into the uniqueKpnI site of pUC19 vector. Expression was observed with pUCO192A and pUCO193 constructs in which the cloned gene(s) were aligned with the upstreamlacZ promoter. Isopropyl -d-thioglucopyranoside (IPTG) enhanced this expression up to 2.5-fold. Specific Cho activity in the cell extracts of the stable pUCO193 transformant were 0.004 U and 0.007 U per mg protein without and with IPTG induction, respectively. Cho activity was detected in the spent medium of this culture, suggesting possible secretion of the enzyme.     

Isolation and physical characterization of streptomycete plasmids

Summary Covalently closed circular DNA was isolated from a strain of Streptomyces coelicolor ATCC 10147 and from a strain of Streptomyces coelicolor subspecies flavus ATCC 19894, using two different methods. The two plasmids were of uniform monomer size: 8.9 kb for pS 10147, the plasmid from S. coelicolor ATCC 10147, and around 125 kb for the plasmid from S. coelicolor ATCC 19894.A restriction enzyme map was constructed for pS 10147, using seven enzymes. Four of the enzymes, (BamHI, Bgl,II, PvuII, and XhoI) cut pS 10147 once while PstI made two cuts. The GC content of this plasmid was calculated to be 72%. The possible utilisation of pS 10147 as a cloning vector in Streptomyces is discussed.     

Degradation of estrogens by Rhodococcus zopfii and Rhodococcus equi isolates from activated sludge in wastewater treatment plants

We have isolated four strains of Rhodococcus which specifically degrade estrogens by using enrichment culture of activated sludge from wastewater treatment plants. Strain Y 50158, identified as Rhodococcus zopfii, completely and rapidly degraded 100 mg of 17beta-estradiol, estrone, estriol, and ethinyl estradiol/liter, as demonstrated by thin-layer chromatography and gas chromatography-mass spectrometry analyses. Strains Y 50155, Y 50156, and Y 50157, identified as Rhodococcus equi, showed degradation activities comparable with that of Y 50158. Using the random amplified polymorphism DNA fingerprinting test, these three strains were confirmed to have been derived from different sources. R. zopfii Y 50158, which showed the highest activity among these four strains, revealed that the strain selectively degraded 17beta-estradiol during jar fermentation, even when glucose was used as a readily utilizable carbon source in the culture medium. Measurement of estrogenic activities with human breast cancer-derived MVLN cells showed that these four strains each degraded 100 mg of 17beta-estradiol/liter to 1/100 of the specific activity level after 24 h. It is thus suggested that these strains degrade 17beta-estradiol into substances without estrogenic activity.