• Journal of Microbiology
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• v.40 no.4
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• pp.274-281
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• 2002

The competitive quantitative PCR method targeting pcbC gene was developed for monitoring 4-chlorobiphenyl(4CB)-degrading bacteria, Pseudomonas sp. strain DJ-12, in soil microcosms. The method involves extraction of DNA from soil contaminated with 4CB, PCR amplification of a pcbC gene fragment from the introduced strain with a set of strain-specific primers, and quantification of the elec-trophoresed PCR product by densitometry. To test the adequacy of the method, Pseudomonas sp. strain DJ-12 was introduced into both contaminated and non-contaminated soil microcosms amended with 4CB. Pseudomonas sp. strain DJ-12 was monitored and quantified by a competitive quantitative PCR in comparison with 4CB degradation and the result was compared to those obtained by using the conventional cultivation method. We successfully detected and monitored 4CB-degrading bacteria in each microcosm and found a significant linear relationship between the number of 4CB-degrading bacteria and the capacity for 4CB biodegradation. The results of DNA spiking and cell-spreading experiments suggest that this competitive quantitative PCR method targeting the pcbC gene for monitoring 4CB- degrading bacteria appears to be rapid, sensitive and more suitable than the microbiological approach in estimating the capacity of 4CB biodegradation in environmental samples.

• Korean Journal of Microbiology
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• v.32 no.2
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• pp.126-131
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• 1994

The pchABCD genes in Pseudomonas sp. DJ-12 speciyin degradation o 4-chlorobiphenyl(4CB) were cloned in Eschericia coli. The cloned cells of E. coli CU1 and CU101 showed to produce 2,3-dihydroxybiphenyl (2,3-DHBP) from 4-chlorobiphenyl by dechlorination, as Pseudomonas so. DJ-12 produced 2,3-DHBP from both biphenyl and 4CB. In particular, E. coli CU101 transformed with the recombinant plasmid of pCU101 revealed dechlorination activity to produce 2,3-DHBP from 4CB without production of 4-chlorobenzoic acid. Therefore, the pcbAB genes (2.2 kb in size) cloned from the chromosome of Pseudomonas sp. DJ-12 were found to have dechlorination activity on 4CB to produce 2,3-DHNP.

• Korean Journal of Microbiology
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• v.31 no.2
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• pp.129-134
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• 1993

The pchAB genes of Pseudomonas sp. DJ-12 produce the enzymes of 4-chlorobipheny] (4CB) dioxygenase and dihydrodiol dehydrogenase which act on the first and second steps in degradation of 4CB and biphenyl. The genes were cloned in E coli XLI-Blue. The pcbAB genes of about 2.2 kb in size were contained in the pCUlO1 hybrid plasmid in the cloned cell of CUIOI. The genes were found to have their own promoter and three restriction sites for HindlII. 2,3-dihydroxybiphenyl was detected by the resting cell assay, as the metabolite transformed from biphenyl by the cloned cell of CUIOI. This means that the pcbAB genes are well expressed in E. coli. But dechlorination was unlikely involved in the pchAB gene expression but was believed to occur by functioning on 4CBA produced after ring-cleavage of 4CB.

• Korean Journal of Microbiology
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• v.30 no.1
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• pp.53-59
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• 1992

Four bacterial strains capable of catabolizing 4-chlorobiphen!;l (4CB) were isolated from the industrial waste waters. The bacterial isolates designated as PO$. P20, P27, and P1242. respectively, were examined for their catabolic activities. And in order to examine molecular homology of the 4CB catabolizing genes of these bacterial isolates. Southern hybridization was conducted with bphABC genes of P. p.srudoalculigrnrs KF707 as a DNA probe. The metabolites of 2-hydroxy-6-0x0-6-(4'-chlorophenyl)hexa-2 .4-dienoic acid and Cchlorobenzoate were detected to be produced by the isolatc:~ in the MM2 liquid cultures. But Cchlorobenzoate was further catabolized to produce 4.-hydroxybenzoate by DJ-12, P08. and P27. but not by P20 and P1242. As results of hybridization, homologous regions were commonly observed in Xhol fragments of 2.2 and 1.8 kb and in EcoRl fragment of 11 kb in the DJ- 12. P08, and P27 isolates. But in any restriction enzyme digests ot the P20 and PI242 isolates. homologous region was not detected. The cbp genes of the bactcria capable of catabolizing 4CB in nature could be divided into two groups by divergence<

• Korean Journal of Microbiology
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• v.28 no.1
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• pp.41-46
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• 1990

A bacterial isolate of DJ-12 capable of degrading 4-chlorobenzoic acid (4CBA) as well as 4-chlorobiphenyl (4CB) was used in this study. Its biodegradability of 4CBA was tested and the location of the genes coding for degradation of 4CBA was investigated by the nethod of in vivo cloning. The genes were found to be existed in the plasmid of pDJ121 which is about 65kb in size and which has 9, 11, 10, and 19 restriction sites for EcoRI, HindIII, SalI, and PstI, respectively. The hybrid plasmid of pDK450 was constructed by ligation of the EcoRI fragments of pDJ121 with pKT230 as a vector. In the recombinant cells selected through transformation of the hybrid vector into Pseudomonas putida KT2440, the 4CBA-degrading genes of DJ-12 were proved to be cloned and expressed in the Pseudomonas sp.

• Journal of Microbiology and Biotechnology
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• v.18 no.5
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• pp.952-957
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• 2008

A Gram-negative bacterium, named LY402, was isolated from contaminated soil. 16S rDNA sequencing and measurement of the physiological and biochemical characteristics identified it as belonging to the genus Enterohacter. Degradation experiments showed that LY402 had the ability to aerobically transform 79 of the 91 major congeners of Aroclor 1242, 1254, and 1260. However, more interestingly, the strain readily degraded certain highly chlorinated and recalcitrant polychlorinated biphenyls (PCBs). Almost all the tri- and tetra-chlorobiphenyls (CBs), except for 3,4,3',4'-CB, were degraded in 3 days, whereas 73% of 3,4,3',4'-, 92% of the penta-, 76% of the hexa-, and 37% of the hepta-CBs were transformed after 6 days. In addition, among 12 octa-CBs, 2,2',3,3',5,5',6,6-CB was obviously degraded, and 2,2',3,3',4,5,6,6'- and 2,2',3,3',4,5,5',6'-CB were slightly transformed. In a metabolite analysis, mono- and dichlorobenzoic acids (CBAs) were identified, and parts of them were also transformed by strain LY402. Analysis of PCB degradation indicated that strain LY402 could effectively degrade PCB congeners with chlorine substitutions in both ortho- and para-positions. Consequently, this is the first report of an Enterobacteria that can efficiently degrade both low and highly chlorinated PCBs under aerobic conditions.

• Journal of Microbiology and Biotechnology
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• v.5 no.2
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• pp.68-73
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• 1995

Pseudomonas sp. P20 was shown to be capable of degrading biphenyl and 4-chlorobiphenyl (4CB) to produce the corresponding benzoic acids wnich were not further degraded. But the potential of the strain for biodegradation of 4CB was shown to be excellent. The pcbA, B, C and D genes responsible for the aromatic ring-cleavage of biphenyl and 4CB degradation were cloned from the chromosomal DNA of the strain. In this study, the pebC and D genes specifying degradation of 2, 3-dihydroxybiphenyl (2, 3-DHBP) produced from biphenyl by the pebAB-encoded enzymes were cloned by using pBluescript SK(＋) as a vector. From the pCK102 (9.3 kb) containing pebC and D genes, pCK1022 inserted with a EcoRI-HindIII DNA fragment (4.1 kb) carrying pebC and D and a pCK1092 inserted with EcoRI-XbaI fragment (1.95 kb) carrying pebC were constructed. The expression of pcbC and D' in E. coli CK102 and pebC in E. coli CK1092 was examined by gas chromatography and UV-vis spectrophotometry. 2.3-dihydroxybiphenyl was readily degraded to produce meta-cleavage product (MCP) by E. coli CK102 after incubation for 10 min, and then only benzoic acid(BA) was detected in the 24-h old culture. The MCP was detected in E. coli CK1022 containing pebC and 0 genes (by the resting cells assay) for up to 3 h after incubation and then diminished completely in 8 h, whereas the MCP accumulated in the E. coli CK1092 culture even after 6 h of incubation. The 2, 3-DHBP dioxygenases (product of pebC gene) produced by E. coli CK1, CK102, CK1023, and CK1092 strains were measured by native PAGE analysis to be about 250 kDa in molecular weight, which were about same as those of Pseudomonas sp. DJ-12, P. pseudoa1caligenes KF707, and P. putida OU83.

• Korean Journal of Soil Science and Fertilizer
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• v.33 no.1
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• pp.47-51
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• 2000

2,4,5-trichlorophenoxyacetic acid (2,4,5-T)-degrading bacterial strains were isolated from rice field and field in suburbs of Taejon. Of the total 100 isolates, 19 strains were selected by fast growth on solid minimal media containing 2,4,5-T as a sole of carbon and energy, and they were identified to genus level. 11 strains were identified as Pseudomonas, 4 strains as Acinetobacter, 1 strains were as Alcaliagenes and 3 strains were not identified. Strains MU19 and MU92 which were identified as Pseudomonas were capable of degradation for 4 kinds of chlorinated aromatic hydrocarbons, 2,4-D, 2,4,5-T, MCPA and 3CB. Acinetobacter sp. MU38 showed the highest degradability in liquid minimal media at 48 hours after inoculation, and Pseudomonas spp. MU19. MU57, MU73, and MU92 were able to degrade carbon source at higher rates. As the results Acinetobacter sp. MU38 and Pseudomonas spp. MU19 and MU92 were capable of biodegradation for broad range of halogenated aromatic hydrocarbons, and had higher rates of degradation for 2,4,5-T.

• Korean Journal of Soil Science and Fertilizer
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• v.33 no.4
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• pp.261-265
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• 2000

To develope the enhanced bacterial strains capable of biodegradation for various chlorinated aromatic compounds, 100 bacterial strains were isolated from soil samples of suburbs of Taejon, Cheongju, and Jeonju by the enrichment culture. These strains can degrade pentachlorophenol (PCP) which is a kind of wood preservatives. Nineteen strains of the isolates were selected by fast colony-forming rate on solid minimal media containing PCP as an only source of carbon and energy. These strains were identified to genus level. Fifteen strains were identified as Pseudomonas, 1 strain as Acinetobacter and 3 strains were not. Genus Alcaligenes strains were not found among them. Pseudomonas sp. MU135. MU139, MU163 and MU 184 were able to degrade for 4 kinds of chlorinated compounds, PCP, 2,4-D, MCPA and 3CB. Pseudomonas sp. If was observed that MU139 exhibits the highest degradability in liquid minimal media at 72 hours after inoculation. Pseudomoans sp. MU147, MU177, MU184 and MU192 also degraded the compounds at higher rates. As the results, Pseudomonas sp. MU139 and unidentified strain MU184 had biodegrability for broad range of chlorinated compounds and higher rates of degradation for PCP.

• Asian-Australasian Journal of Animal Sciences
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• v.33 no.1
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• pp.24-34
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• 2020

Objective: The effects of Pleurotus ostreatus on the feed utilization of broad bean stalks (BBS), rape straw (RS), paddy straw (PS), and corn stalk (CS) was examined. Methods: The four roughages were co-cultured with Pleurotus ostreatus. The chemical composition; enzyme activities of laccase, carboxymethylcellulase (CMCase) and xylanase; carbohydrate and protein fractions (based on The Cornell Net Carbohydrate and Protein System [CNCPS]) were assessed at different days after inoculation (7, 14, 21, 28 d) and un-inoculated roughages (control, 0 d). The digestibility of nutrient components and the gas production of roughage with various incubation times were monitored at 0, 2, 4, 6, 9, 12, 24, 36, 48, 60, and 72 h using an in vitro ruminal fermentation method. Results: A higher CMCase activity (0.1039 U/mL) and earlier time to peak (14 d) were detected in Pleurotus ostreatus cultured with CS (p<0.05). Significantly, the incubation length-dependent responses of cumulative gas production were observed from 24 to 72 hours post fermentation (p<0.05), and these incubation length-dependent effects on cumulative gas production of PS and CS appeared earlier (24 h) for PS and CS than those (48 h) for BBS and RS (p<0.05). The fast-degradable carbohydrate (CA) content for all four roughages significantly increased over time (p<0.05). Nonetheless, increased degradation efficiency for CA treated with Pleurotus ostreatus was detected at both 21 and 28 days of incubation (p<0.05). With the exception of PS (p<0.05), there were no significant difference among the roughages (p>0.05) in slowly-degradable carbohydrate (CB2) at different incubation times (p<0.05). Conclusion: Assessment of the alterations in chemical composition, CNCPS system fractions, and the fermentation kinetics after biological pretreatment may yield a valuable database for evaluating the biological pretreatment of Pleurotus ostreatus in ruminant feed.