• Journal of Microbiology and Biotechnology
• /
• v.23 no.11
• /
• pp.1617-1626
• /
• 2013

The Bacterial community structure and its complexity of the enrichment culture during the isolation and screening of imidacloprid-degrading strain were studied using denaturating gradient gel electrophoresis analysis. The dominant bacteria in the original tea rhizosphere soil were uncultured bacteria, Rhizobium sp., Sinorhizobium, Ochrobactrum sp., Alcaligenes, Bacillus sp., Bacterium, Klebsiella sp., and Ensifer adhaerens. The bacterial community structure was altered extensively and its complexity reduced during the enrichment process, and four culturable bacteria, Ochrobactrum sp., Rhizobium sp., Geobacillus stearothermophilus, and Alcaligenes faecalis, remained in the final enrichment. Only one indigenous strain, BCL-1, with imidacloprid-degrading potential, was isolated from the sixth enrichment culture. This isolate was a gram-negative rod-shaped bacterium and identified as the genus Ochrobactrum based on its morphological, physiological, and biochemical properties and its 16S rRNA gene sequence. The degradation test showed that approximately 67.67% of the imidacloprid (50 mg/l) was degraded within 48 h by strain BCL-1. The optimum conditions for degradation were a pH of 8 and $30^{\circ}C$. The simulation of imidacloprid bioremediation by strain BCL-1 in soil demonstrated that the best performance in situ (tea soil) resulted in the degradation of 92.44% of the imidacloprid (100 mg/g) within 20 days, which was better than those observed in the ex situ simulations that were 64.66% (cabbage soil), 41.15% (potato soil), and 54.15% (tomato soil).

• Mycobiology
• /
• v.44 no.3
• /
• pp.155-161
• /
• 2016

The most economically important species used in a wide range of fermentation industries throughout Asia belong to Aspergillus section Flavi, which are morphologically and phylogenetically indistinguishable, with a few being toxigenic and therefore a major concern. They are frequently isolated from Korean fermentation starters, such as nuruk and meju. The growing popularity of traditional Korean alcoholic beverages has led to a demand for their quality enhancement, therefore requiring selection of efficient non-toxigenic strains to assist effective fermentation. This study was performed to classify the most efficient strains of Aspergillus section Flavi isolated from various types of traditional wheat nuruk, based on a polyphasic approach involving molecular and biochemical evaluation. A total of 69 strains were isolated based on colony morphology and identified as Aspergillus oryzae/flavus based on internal transcribed spacer and calmodulin gene sequencing. Interestingly, none were toxigenic based on PCR amplification of intergenic regions of the aflatoxin cluster genes norB-cypA and the absence of aflatoxin in the culture supernatants by thin-layer chromatography analysis. Saccharification capability of the isolates, assessed through ${\alpha}-amylase$ and glucoamylase activities, revealed that two isolates, TNA24 and TNA15, showed the highest levels of activity. Although the degrees of variation in ${\alpha}-amylase$ and glucoamylase activities among the isolates were higher, there were only slight differences in acid protease activity among the isolates with two, TNA28 and TNA36, showing the highest activities. Furthermore, statistical analyses showed that ${\alpha}-amylase$ activity was positively correlated with glucoamylase activity (p < 0.001), and therefore screening for either was sufficient to predict the saccharifying capacity of the Aspergillus strain.

• Asian Pacific Journal of Cancer Prevention
• /
• v.16 no.2
• /
• pp.579-583
• /
• 2015

This study was conducted on female patients with different gynecological problems attending the gynecology out-patient departments of two tertiary care hospitals in Peshawar city of Khyber Pakhtunkhwa, Pakistan between August 2012 and October 2013. The 200 patients had an age range of 21-65 years. Smears were taken with cervical brushes and preserved in preservative medium and processed for manual liquid based cytology (MLBC) for Pap staining. Out of 200 collected samples, 30 samples were found inadequate on cytology. Of the remaining 170 samples, 164 (96.47%) were normal, 5 (2.94%) were of atypical squamous cells of unknown significance (ASCUS) and 1 (0.6%) was of high grade squamous intraepithelial lesion (HSIL). On PCR all the samples were positive for beta globin gene fragment including those reported inadequate on cytology. Out of the 5 ASCUS samples, 2 samples were positive for HPV, one each for HPV 16 and HPV 18, and the rest of the 3 samples were negative for HPV DNA. The 1 sample of HSIL was positive for HPV 16 on PCR. Out of 164 normal samples on cytology, only 1 sample was HPV 16 positive. So overall, 4 (2%) out of 200 samples were positive for HPV DNA, where 3 were HPV 16 (1.5%), and 1 was HPV 18 (0.5%) positive, and thus the ratio of infection with of HPV 16 to HPV 18 was 3:1 in the general population. In conclusion, PCR based HPV detection is a more sensitive method for screening of HPV infection than cytology as sample inadequacy does not affect the results. However, it can be combined with cytology methods in a HPV positive female to achieve the maximum results.

• Proceedings of the Korean Society for Bioinformatics Conference
• /
• 2004.11a
• /
• pp.272-278
• /
• 2004

Recently, Pepe et al. (2003) employed the receiver operating characteristic (ROC) approach to rank candidate genes from a microarray experiment that can be used for the biomarker development with the ultimate purpose of the population screening of a cancer, In the cancer microarray experiment based on n patients the researcher often wants to compare the tumor tissue with the normal tissue within the same individual using a common reference RNA. This design is referred to as a reference design or an indirect design. Ideally, this experiment produces n pairs of microarray data, where each pair consists of two sets of microarray data resulting from reference versus normal tissue and reference versus tumor tissue hybridizations. However, for certain individuals either normal tissue or tumor tissue is not large enough for the experimenter to extract enough RNA for conducting the microarray experiment, hence there are missing values either in the normal or tumor tissue data. Practically, we have $n_1$ pairs of complete observations, $n_2$ 'normal only' and $n_3$ 'tumor only' data for the microarray experiment with n patients, where n=$n_1$+$n_2$+$n_3$. We refer to this data set as a mixed data set, as it contains a mix of fully observed and partially observed pair data. This mixed data set was actually observed in the microarray experiment based on human tissues, where human tissues were obtained during the surgical operations of cancer patients. Pepe et al. (2003) provide the rationale of using ROC approach based on two independent samples for ranking candidate gene instead of using t or Mann -Whitney statistics. We first modify ROC approach of ranking genes to a paired data set and further extend it to a mixed data set by taking a weighted average of two ROC values obtained by the paired data set and two independent data sets.

• Asian Pacific Journal of Cancer Prevention
• /
• v.17 no.3
• /
• pp.1053-1055
• /
• 2016

Background: Polycythemia rubra vera (PV), being a primary polycythemia, is caused by neoplastic proliferation of erythroid, megakaryocytic and granulocytic lineages which result in panmyelosis. PV patients have a somatic acquired mutation in the Janus kinase (JAK2) pathway, rendering cell proliferation independent of the normal regulatory mechanisms that regulate erythropoiesis. The rational of this study was to determine the prevalence of the JAK-2 V617F mutation in Pakistani patients with PV. Materials and Methods: In this cross sectional study, 26 patients with PV were enrolled from January 2010 to December 2014. Patients were diagnosed based on WHO criteria for PV. All were screened for G-T point mutation (V617F) in the JAK2 gene on chromosome 9 by an allele specific PCR. Results: The mean age was $53.4{\pm}9.31years$ (range 36-72) and the male to female ratio was 2:1. The frequency of JAK2 V617F positivity in our PV patients was found to be 92.3%. Overall 30.7% of patients were asymptomatic and remaining 69.3% presented with symptomatic disease. The mean hemoglobin was $18.1{\pm}1.9g/dl$ with the mean hematocrit of $55.6{\pm}8.3%$. The mean total leukocyte count was $12.8{\pm}7.1{\times}10^9/l$ and the platelet count was $511{\pm}341.9{\times}10^9/l$. A positive correlation of JAK2 V617F mutation was established with high TLC count (P=0.01). No correlation of JAK2 V617F could be established with age or gender (P>0.05). Conclusions: The JAK2 V617F mutation frequency in our PV patients was similar to those reported internationally. Screening for the mutation in all suspected PV cases could be beneficial in differentiating patients with reactive and clonal erythrocytosis.

• Asian Pacific Journal of Cancer Prevention
• /
• v.16 no.17
• /
• pp.7889-7892
• /
• 2015

Background: Primary idiopathic myelofibrosis (PMF) is a clonal Ph-chromosome negative myeloproliferative neoplasm characterized by dysregulated kinase signaling and release of abnormal cytokines. In the recent past, following JAK2 V617F mutation invention, important revolution has been made in the molecular diagnostic biology of this disease. The rational of this study was to determine the mutational status of JAK2 V617F in Pakistan patients with PMF. Materials and Methods: In this cross sectional study, 20 patients with PMF were enrolled from January 2011 to December 2014. Diagnosis was based on WHO criteria for PMF. All patients were screened for G-T point mutation (V617F) in the JAK2 gene on chromosome 9 by allele specific PCR. Results: The mean age was $57.9{\pm}16.5years$. The male to female ratio was 3:1. The frequency of JAK2 V617F positivity in our PMF patients was found to be 55%. Positive correlations of JAK2 V617F mutation were established with high TLC count, raised LDH and marked splenomegaly (P<0.05). No correlation of JAK2 V617F could be established with age and gender (P>0.05). Conclusions: The JAK2 V617F mutation frequency in our PMF patients was similar to those reported previously. In our hands JAK2 V617F mutated patients expressed an aggressive disease phenotype. Screening for the mutation in all suspected PMF cases could be beneficial in differentiating patients with reactive and clonal marrow fibrosis.

• YAKHAK HOEJI
• /
• v.38 no.3
• /
• pp.293-299
• /
• 1994

Forty nine clinical isolates of S. aureus showing resistance to erythromycin(EM) were selected from 83 strains isolated recently in Korea. Fourteen strains of S. aureus showing inducible resistance to MLS antibiotics were selected by disc agar diffusion method. Colony hydridization was executed using two MLS inducible resistance genes, ermA and ermC, identified previously from S. aureus as probes. S. aureus 375 and S. aureus 507 whose genes were not homologous to those probes were finally selected. It was confirmed that the resistance genes of S. aureus 375 and S. aureus 507 had no homology with those probes in southern hybridization test using ermA, ermC and ermAM as probes. It was determined that S. aureus 375 had a plasmid whose size was about 35 kb. To know if the plasmid may have the genes related to inducible resistance to MLS antibiotics, it was attempted to transform Bacillus subtillis BR151 and S. aureus RN4220 with the plasmid isolated from S. aureus 375. It was shown that the gene related to inducible resistance to MLS antibiotics did not exist in this plasmid. These results indicate that two clinical isolates of S. aureus showing inducible resistance to MLS antibiotics have novel genes that have no homology with MLS resistance genes identified so far. It is assumed that these genes may exist in chromosomal DNA.

• The Plant Pathology Journal
• /
• v.30 no.1
• /
• pp.58-67
• /
• 2014

The multifunctional triple gene block protein 1 (TGB1) of the Potexvirus Alternanthera mosaic virus (AltMV) has been reported to have silencing suppressor, cell-to-cell movement, and helicase functions. Yeast two hybrid screening using an Arabidopsis thaliana cDNA library with TGB1 as bait, and co-purification with TGB1 inclusion bodies identified several host proteins which interact with AltMV TGB1. Host protein interactions with TGB1 were confirmed by biomolecular fluorescence complementation, which showed positive TGB1 interaction with mitochondrial ATP synthase delta' chain subunit (ATP synthase delta'), light harvesting chlorophyll-protein complex I subunit A4 (LHCA4), chlorophyll a/b binding protein 1 (LHB1B2), chloroplast-localized IscA-like protein (ATCPISCA), and chloroplast ${\beta}$-ATPase. However, chloroplast ${\beta}$-ATPase interacts only with $TGB1_{L88}$, and not with weak silencing suppressor $TGB1_{L88}$. This selective interaction indicates that chloroplast ${\beta}$-ATPase is not required for AltMV movement and replication; however, TRV silencing of chloroplast ${\beta}$-ATPase in Nicotiana benthamiana induced severe tissue necrosis when plants were infected by AltMV $TGB1_{L88}$ but not AltMV $TGB1_{L88}$, suggesting that ${\beta}$-ATPase selectively responded to $TGB1_{L88}$ to induce defense responses.

• Proceedings of the Korean Environmental Sciences Society Conference
• /
• 2006.11a
• /
• pp.285-289
• /
• 2006

In screening of algicidal bacteria, we isolated a marine bacterium which had potent algicidal effects on harmful algal bloom (HAB) species. This organism was identified as a strain very close to Bacillus subtilisby 16S rRNA gene sequencing. This bacterium, Bacillus sp. SY-1, produces very active algicidal compounds against the harmful dinoflagellate Cochlodinium polykrikoides. We isolated three algicidal compounds (MS 1056, 1070, 1084) and identified them by amino acid analyses, fast atom bombardment mass spectrometry (FAB-MS), infrared spectroscopy (IR), $^1H$, $^{13}C$, and extensive two-dimensional nuclear magnetic resonance (2D NMR) techniques including $^1H-^{15}N$ HMBC analysis. One of them, MS 1056, contains a b-amino acid residue with an alkyl side chain of $C_{15}$. MS 1056, 1070, and 1084 showed algicidal activities against C. polykrikoides with an $LC_{50}$ (6 hrs) of 2.3, 0.8, $0.6\;{\mu}g/ml$, respectively. These compounds also showed significant algicidal activities against other harmful dinoflagellates and raphidophytes. In contrast, MS 1084 showed no significant growth inhibition against various organisms coexisting with HAB species in natural environments, including bacteria, eukaryotic microalgae, and cyanobacteria, although it inhibited growth of some fungi and yeasts. These observations imply that algicidal bacterium Bacillus sp. SY-1 and its algicidal compounds could play an important role in regulating the onset and development of HABs in the natural environments.

• Asian-Australasian Journal of Animal Sciences
• /
• v.23 no.2
• /
• pp.272-278
• /
• 2010

In this study, we cloned, sequenced and characterized porcine y+L Amino Acid Transporter-1 (y+LAT1). By screening a translated EST database with the protein sequence of the human $y^{+}$LAT1 and by using rapid amplification of cDNA ends (RACE), the full-length cDNA encoding porcine $y^{+}$LAT1 was isolated from porcine intestine RNA. It was 2,111 bp long, encoding a 511 amino acid trans-membrane glycoprotein composed of 12 transmembrane domains. The predicted amino acid sequence was found to be 91%, 90%, 87% and 87% identical to those of cattle, human, mouse and rat $y^{+}$LAT1 respectively. Real-time RT-PCR results indicated that the small intestine had the highest $y^{+}$LAT1 mRNA abundance and the lung had the lowest $y^{+}$LAT1 mRNA abundance. Baby hamster kidney (BHK) cells transfected with green fluorescent protein (GFP) tagged porcine $y^{+}$LAT1 cDNA indicated that the cellular localization of the gene product in BHK was on the plasma membrane.