• Journal of Environmental Health Sciences
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• v.34 no.1
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• pp.86-94
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• 2008

Lake Shiwha, an artificial lake located near metropolitan Seoul, offers a unique water environment and has been suspected to have high levels of chemical and microbiological contaminations. Lake Shiwha was originally connected to the sea but currently has four major surface water inputs from agricultural, municipal, industrial areas and in addition an occasional inflow from the sea. The objectives of this study are to investigate the relative contribution of microbial contaminants from each of the inflowing surface waters and to identify appropriate microbial indicator organisms in this unique water environment. We measured the levels of microbial contaminations in the four inflowing surface waters. A number of microbial indicator organisms including total coliform (TC), fecal coliform (FC), E. coli, Enterococci, somatic and male-specific coliphages were analyzed. Bacterial indicator microorganisms were detected and quantified by the $Colilert^{(R)},\;Enterolert^{(R)}$ kit. Surface water (50 l) was sampled by $ViroCap^{TM}\;5"$ cartridge filters and analyzed by the single agar layer method for detecting coliphages. The concentrations of TC, FC, E. coli, and Enterococci were 1543 CFU/100 ml${\sim}1.99{\times}10^6$ CFU/100 ml, 0 CFU/100 ml${\sim}202$ CFU/100ml, 0 CFU/100 ml${\sim}1.80{\sim}10^5$ CFU/100ml, 74 CFU/100 ml${\sim}3408$ CFU/100 ml, respectively. The male-specific and somatic coliphages were detected in three different inflowing surface waters. Isolated E. coli and Enterococci strains were further analyzed by 16s rDNA amplification and subsequent phylogenetic analysis from Jungwang-chun, Ansan-chun, Banwol-chun and penstock of inflowing surface water. Our results indicated that the concentrations of different fecal indicator microorganisms might not be highly correlated with each other. Multiple microbial indicator organisms should be used for monitoring microbial contamination and microbial source tracking methods.

• Asian-Australasian Journal of Animal Sciences
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• v.28 no.1
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• pp.50-57
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• 2015

This study was conducted to investigate the effect of soybean meal (SM) and soluble starch (SS) on biogenic amine production and microbial diversity using in vitro ruminal fermentation. Treatments comprised of incubation of 2 g of mixture (expressed as 10 parts) containing different ratios of SM to SS as: 0:0, 10:0, 7:3, 5:5, 3:7, or 0:10. In vitro ruminal fermentation parameters were determined at 0, 12, 24, and 48 h of incubation while the biogenic amine and microbial diversity were determined at 48 h of incubation. Treatment with highest proportion of SM had higher (p<0.05) gas production than those with higher proportions of SS. Samples with higher proportion of SS resulted in lower pH than those with higher proportion of SM after 48 h of incubation. The largest change in $NH_3$-N concentration from 0 to 48 h was observed on all SM while the smallest was observed on exclusive SS. Similarly, exclusive SS had the lowest $NH_3$-N concentration among all groups after 24 h of incubation. Increasing methane ($CH_4$) concentrations were observed with time, and $CH_4$ concentrations were higher (p<0.05) with greater proportions of SM than SS. Balanced proportion of SM and SS had the highest (p<0.05) total volatile fatty acid (TVFA) while propionate was found highest in higher proportion of SS. Moreover, biogenic amine (BA) was higher (p<0.05) in samples containing greater proportions of SM. Histamines, amine index and total amines were highest in exclusive SM followed in sequence mixtures with increasing proportion of SS (and lowered proportion of SM) at 48 h of incubation. Nine dominant bands were identified by denaturing gradient gel electrophoresis (DGGE) and their identity ranged from 87% to 100% which were mostly isolated from rumen and feces. Bands R2 (uncultured bacterium clone RB-5E1) and R4 (uncultured rumen bacterium clone L7A_C10) bands were found in samples with higher proportions of SM while R3 (uncultured Firmicutes bacterium clone NI_52), R7 (Selenomonas sp. MCB2), R8 (Selenomonas ruminantium gene) and R9 (Selenomonas ruminantium strain LongY6) were found in samples with higher proportions of SS. Different feed ratios affect rumen fermentation in terms of pH, $NH_3$-N, $CH_4$, BA, volatile fatty acid and other metabolite concentrations and microbial diversity. Balanced protein and carbohydrate ratios are needed for rumen fermentation.

• Korean Journal of Microbiology
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• v.42 no.1
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• pp.40-46
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• 2006

Galactose joined to glucose by a $\beta(1\rightarrow4)$ glycosidic bond makes lactose and this disaccharide is rich in milk. It is known that lacotse is hydrolyzed to each monomeric sugar by either lactase in human or $\beta-galactosidase$ in bacteria. Ingestion of milk by lactase-deficient persons causes a temporary diarrhea and subsequent chronic diarrhea results in colitis with chronic inflammation. We isolated a $\beta-galactosidase$ producing psycrotolerant strain AS-20 from near cattle shed and investigated the growth at various temperature conditions. Whereas Escherichia coli strains did not grow at $10^{\circ}C$, the AS-20 strain could grow well at this low temperature and showed optimal growth at $30^{\circ}C$. The isolated strain was identified as 97% Hafnia alvei by biochemical properties. This strain could ferment glucose, lacotse, maltose, mannitol, xylose, ONPG, rhamanose and L-arabinose, and decarboxylate lysin and ornithine. To confirm the identity of isolated strain we amplified 16S rDNA by PCR and searched similarity of the 1426 bp DNA sequcence with Genbank database. The strain AS-20 showed 99% similarity with Hafnia alvei. The activity of $\beta-galactosidase$ was 1.5 times higher when the cell was grown at 10 or $20^{\circ}C$ than at $30^{\circ}C$. The highest enzyme activity of AS-20 was also much higher than that of E. coli, which was grown at $30^{\circ}C$.

• Journal of Life Science
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• v.15 no.4 s.71
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• pp.657-663
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• 2005

Collagenases are generally defined as enzymes that are capable of degrading the polypeptide backbone of native collagen under conditions that do not denature the protein. An extracellular collagenase-producing bacterial strain was isolated from kimchi and identified to be Bacillus subtilis JS-17 through morphological, cultural, biochemical characteristics and 16S rDNA sequence analysis. Optimum culture condition of Bacillus subtilis JS-17 for the production of collagenase was $1.5\%$ fructose, $1\%$ yeast extract, $0.5\%\;K_2HPO_4,\;0.4\%\;KH_2PO_4,\;0.01\%\;MgSO_4\cdot7H_2O,\;0.01\%\; MnSO_4\cdot4H_2O,\;,0.1\%$ citrate and $0.1\%\;CaCl_2$. The production of collagenase was optimal at $30^{\circ}C$ for 72 hr. A collagenase was isolated from the culture filtrate of Bacillus subtilis JS-17. The enzyme was purified using Amberlite IRA-900 column chromatography, Sephacryl S-300 HR column chromatography and DEAE-Sephadex A-50 column chromatography The purified collagenase has an specific activity 192.1 units/mg. The molecular weight of the purified enzyme was estimated to be 28 kDa by SDS-PACE. The purified collagenase has $100\%$ activity up to $55^{\circ}C$.

• Environmental Engineering Research
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• v.10 no.5
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• pp.213-226
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• 2005

To treat wastewater efficiently by a one-step process of nitrogen removal, a new bacterial strain producing $N_2$ gas from ${NH_4}^+$ under an aerobic condition was isolated and identified. The cell was motile and a Gram-negative rod, and usually occurred in pairs. By 16S-rDNA analysis, the isolated strain was identified as Enterobacter asburiae with 96% similarity. The isolate showed that the capacity of $N_2$ production under an oxic condition was approximately three times higher than that under an anoxic condition. Thus, the consumption of ${NH_4}^+$ by the isolate was significantly different in the metabolism of $N_2$ production under the two different environmental conditions. The optimal conditions of the immobilized isolate for $N_2$ production were found to be pH 7.0, $30^{\circ}C$ and C/N ratio 5, respectively. Under all the optimum reaction conditions, $N_2$ production by the immobilized isolate resulted in reduction of ORP with both the consumption of DO and the drop of pH. The removal efficiencies of $COD_{Cr}$, and TN were 56.1 and 60.9%, respectively. The removal rates of $COD_{Cr}$, and TN were the highest for the first 2.5 hrs with the removal $COD_{Cr}/TN$ ratios of 32.1, and afterwards the rates decreased as reaction proceeded. For application of the immobilized isolate to a practical process of ammonium removal, a continuous operation was executed with a synthetic medium of a low C/N ratio. The continuous bioreactor system exhibited a satisfactory performance at 12.1 hrs of HRT, in which the effluent concentrations of ${NH_4}^+$-N was measured to be 15.4 mg/L with its removal efficiency of 56.0%. The maximum removal rate of ${NH_4}^+$-N reached 1.6 mg ${NH_4}^+$-N/L/hr at 12.1 hrs of HRT(with N loading rate of $0.08\;Kg-N/m^3$-carrier/d). As a result, the application of the immobilized isolate appears a viable alternative to the nitrification-denitrification processes.

• Journal of Embryo Transfer
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• v.20 no.2
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• pp.169-176
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• 2005

Loop-mediated isothermal amplification (LAMP) is novel DNA amplification methods that amplifies a target sequence specifically under isothemal condition. The present study was to assess the in vitro viability afier biopsy and sexing rate of different types of embryo biopsied. In vivo compact morulae and blastocyst embryos were obtained from Korean Native Cow (KNC) superovulated with FSH (Antorin, R-10) on 7 Day after artificial insemination. in vitro compact morulae and blastocyst embryos were obtained with KNC or Holsteins that were gained on 6, 7 or 8 day after in vitro fertilization(IVF) with frozen semen. Biopsy of bovine embryo was carried out in a $80{\mu}l$ drop with $Ca^{2+}-Mg^{2+}$ free D-PBS and the viability of biopsied embryos were evaluated in IVMD (IFP, Japan) medium at 12 hrs culture time. The sex ratio of biopsied Hanwoo embryos were male vs. female of $43.5\%\;vs.\;56.5\%$ in vivo and $33.9\%\;vs.\;49.2\%$ in vitro respectively, and male rate of biopsied Holstein embryos were significantly higher than female $(70.8\;vs.\;29.2\%)$. and indefinite rate of in vitro embryos was $16.9\%$ and in vivo was not. The degeneration rate of biopsied embryo, in vitro embryos were significantly higher than in vivo $(13.2\%\;vs,\;0.0\%,\;p<0.05)$. The survivability of in vivo embryo were between biopsied following punching method was significantly (P<0.05) higher than bisection method produced embryos $(100\%\;vs.\;83.3\%)$ and in vitro had no difference. However, the degeneration rate of biopsied embryo by bisection method was significantly higher than punching methods between in vivo and in vitro $(16.7\;vs.\;22.6\%,\;respectively,\;p<0.05)$. In conclusion, these results indicate that punching method was optimal and survivability after embryo biopsy was useful for reducing the damage caused by the embryo biopsy procedure for LAMP-based embryo sexing.

• Asian-Australasian Journal of Animal Sciences
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• v.29 no.11
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• pp.1555-1561
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• 2016

Shank skin color of Korean native chicken (KNC) shows large color variations. It varies from white, yellow, green, bluish or grey to black, whilst in the majority of European breeds the shanks are typically yellow-colored. Three shank skin color-related traits (i.e., lightness [$L^*$], redness [$a^*$], and yellowness [$b^*$]) were measured by a spectrophotometer in 585 progeny from 68 nuclear families in the KNC resource population. We performed genome scan linkage analysis to identify loci that affect quantitatively measured shank skin color traits in KNC. All these birds were genotyped with 167 DNA markers located throughout the 26 autosomes. The SOLAR program was used to conduct multipoint variance-component quantitative trait locus (QTL) analyses. We detected a major QTL that affects $b^*$ value (logarithm of odds [LOD] = 47.5, $p=1.60{\times}10^{-49}$) on GGA24 (GGA for Gallus gallus). At the same location, we also detected a QTL that influences $a^*$ value (LOD = 14.2, $p=6.14{\times}10^{-16}$). Additionally, beta-carotene dioxygenase 2 (BCDO2), the obvious positional candidate gene under the linkage peaks on GGA24, was investigated by the two association tests: i.e., measured genotype association (MGA) and quantitative transmission disequilibrium test (QTDT). Significant associations were detected between BCDO2 g.9367 A>C and $a^*$ ($P_{MGA}=1.69{\times}10^{-28}$; $P_{QTDT}=2.40{\times}10^{-25}$). The strongest associations were between BCDO2 g.9367 A>C and $b^*$ ($P_{MGA}=3.56{\times}10^{-66}$; $P_{QTDT}=1.68{\times}10^{-65}$). However, linkage analyses conditional on the single nucleotide polymorphism indicated that other functional variants should exist. Taken together, we demonstrate for the first time the linkage and association between the BCDO2 locus on GGA24 and quantitatively measured shank skin color traits in KNC.

• Korean Journal of Microbiology
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• v.52 no.2
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• pp.166-174
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• 2016

In order to find an efficient bacterial strain that can carry out nitrification and denitrification simultaneously, we isolated many heterotrophic nitrifying bacteria from wastewater treatment plant. One of isolates NS13 showed high removal rate of ammonium and was identified as Alcaligenes faecalis by analysis of its 16S rDNA sequence, carbon source utilization and fatty acids composition. This bacterium could remove over 99% of ammonium in a heterotrophic medium containing 140 mg/L of ammonium at pH 6-9, $25-37^{\circ}C$ and 0-4% of salt concentrations within 2 days. It showed even higher ammonium removal at higher initial ammonium concentration in the medium. A. faecalis NS13 could also reduce nitrate and nitrous oxide by nitrate reductase and nitrous oxide reductase, respectively, which was confirmed by detection of nitrate reductase gene, napA, and nitrous oxide reducase gene, nosZ, by PCR. One of metabolic intermediate of denitrification, $N_2O$ was detected from headspace of bacterial culture. Based on analysis of all nitrogen compounds in the bacterial culture, 42.8% of initial nitrogen seemed to be lost as nitrogen gas, and 46.4% of nitrogen was assimilated into bacterial biomass which can be removed as sludge in treatment processes. This bacterium was speculated to perform heterotrophic nitrification and aerobic denitrification simultaneously, and may be utilized for N removal in wastewater treatment processes.

• Microbiology and Biotechnology Letters
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• v.26 no.2
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• pp.122-129
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• 1998

It were reported that antifungal mechanism of Enterobacter cloacae is a volatile ammonia that produced by the strain in soil, and the production of ammonia is related to the bacterial urease activity. A powerful bacterium SH14 against soil-borne pathogen Fusarium solani, which cause root rot of many important crops, was selected from a ginseng pathogen suppressive soil. The strain SH14 was identified as Bacillus subtilis by cultural, biochemical, morphological method, and $API^{circledR}$ test. From several in vitro tests, the antifungal substance that is produced from B. subtilis SH14 was revealed as heat-stable and low-molecular weight antibiotic substance. In order to construct the multifunctional biocontrol agent, the urease gene of Bacillus pasteurii which can produce pathogenes-suppressive ammonia transferred into antifungal bacterium. First, a partial BamH I digestion fragment of plasmid pBU11 containing the alkalophilic B. pasteurii l1859 urease gene was inserted into the BamH I site of pEB203 and expressed in Escherichia coli JM109. The recombinant plasmid was designated as pGU366. The plasmid pGU366 containing urease gene was introduced into the B. subtilis SH14 with PEG-induced protoplast transformation (PIP) method. The urease gene was very stably expressed in the transformant of B. subtilis SH14. Also, the optimal conditions for transformation were established and the highest transformation frequency was obtained by treatment of lysozyme for 90 min, and then addition of 1.5 ${mu}g$/ml DNA and 40% PEG4000. From the in vitro antifungal test against F. solani, antifungal activity of B. subtilis SH14(pGu366) containing urease gene was much higher than that of the host strain. Genetical development of B. subtilis SH14 by transfer of urease gene can be responsible for enhanced biocontrol efficacy with its antibiotic action.