A Surge in the Seroprevalence of Toxoplasmosis among the Residents of Islands in Gangwha-gun, Incheon, Korea

Seroepidemiological changes of Toxoplasma gondii infection among the residents of the islands of Gangwha-gun, Incheon for 2 years were surveyed and evaluated by ELISA using a crude extract antigen. In 2010, sera of 919 adult residents in Gyodong-myeon and 313 adults in Samsan-myeon were collected and checked for IgG antibody titers, which showed 14.5% (133 sera) and 19.8% (62 sera) positive rates, respectively. In 2011, sera of 955 adults in Gyodong-myeon and 341 adults in Samsan-myeon were examined, which showed an increase of positive rates to 23.8% (227 sera) and 31.7% (108 sera), respectively. Totally, the seroprevalence of the first year was 15.8% and it increased rapidly to 25.8% in the second year. The positive rates of both sexes increased simultaneously with the significant ratio of males to females by 1.7-2.2 fold (P<0.05). In both myeons, 661 sera were collected every year and showed changes in optical density (OD) in 177 sera; newly found as positives in 73 persons (11.0%), negative conversion in 10 persons (1.5%), and maintained or increased in 94 persons (14.2%). This rapid increase in the prevalence of toxoplasmosis in Gangwha islands may be due to in part peculiar changes in the toxoplasmic environment of the islands and presumably the consumption of the pork bred domestically within the islands or imported from high endemic nations. It is necessary to find out symptomatic toxoplasmic patients and confirm the risk factors for further infection in the islands of Gangwha-gun.     

Free radicals enzymatically triggered by Clonorchis sinensis excretory-secretory products cause NF-κB-mediated inflammation in human cholangiocarcinoma cells

Chronic clonorchiasis, caused by direct and continuous contact with Clonorchis sinensis worms and their excretory-secretory products, is associated with hepatobiliary damage, inflammation, periductal fibrosis and even development of cholangiocarcinoma. Our previous report revealed that intracellular reactive oxygen species were generated in C. sinensis excretory-secretory product-treated human cholangiocarcinoma cells; however, their endogenous sources and pathophysiological roles in host cells were not determined. In the present study, we found that treatment of human cholangiocarcinoma cells with excretory-secretory products triggered increases in free radicals via a time-dependent activation of NADPH oxidase, xanthine oxidase and inducible nitric oxide synthase. This increase in free radicals substantially promoted the degradation of cytosolic IκB-α, nuclear translocation of nuclear factor-κB subunits (RelA and p50), and increased κB consensus DNA-binding activity. Excretory-secretory product-induced nuclear factor-κB activation was markedly attenuated by preincubation with specific inhibitors of each free radical-producing enzyme or the antioxidant, N-acetylcysteine. Moreover, excretory-secretory products induced an increase in the mRNA and protein expression of the proinflammatory cytokines, IL-1β and IL-6, in an nuclear factor-κB-dependent manner, indicating that enzymatic production of free radicals in ESP-treated cells participates in nuclear factor-κB-mediated inflammation. These findings provide new insights into the pathophysiological role of C. sinensis excretory-secretory products in host chronic inflammatory processes, which are initial events in hepatobiliary diseases.     

Genomic sequence analysis of a potential QTL region for fat trait on pig chromosome 6

On pig chromosome 6, the SW71 microsatellite is located in the region corresponding to several quantitative trait loci (QTL), such as those for intramuscular fat content and for body weight at 4 weeks of age. The genomic sequence of approximately 909 kb was obtained from seven BAC clones encompassing the SW71 region corresponding to human 18q11.21-q11.22. By searching the NCBI GenBank using BLASTX and BLASTN, this 909-kb segment was found to contain eight genes, RAB31, TXNDC2, VAPA, APCDD1, NAPG, FAM38B, C18orf30, and C18orf58, and one putative gene (DN119777). The average G + C content in the sequence of this contig was 45.75% and 33 CpG islands were detected. CpG islands were scattered throughout the region in which most of the putative genes were located. Dense CpG islands of approximately 840 bp were observed, including within the 5' UTR and exon 1 of the orthologs of the RAB31, VAPA, APCDD1, and NAPG genes. Comparative analysis of conserved segments of six species showed that K(a)/K(s) ratios of the TXNDC2 gene in collinear and rearranged segments were significantly different at 4.1 and 1.3, respectively. In conclusion, we demonstrated the genomic organization of pig chromosome 6, including the gene order surrounding SW71, which provides important information for comparative mapping. Moreover, the genes revealed in this study may be positional candidate genes associated with QTL on chromosome 6 that affect fat deposition in pigs.     

Confirming single nucleotide polymorphisms from expressed sequence tag datasets derived from three cattle cDNA libraries

Using the Phred/Phrap/Polyphred/Consed pipeline established in the National Livestock Research Institute of Korea, we predicted candidate coding single nucleotide polymorphisms (cSNPs) from 7,600 expressed sequence tags (ESTs) derived from three cDNA libraries (liver, M. longissimus dorsi, and intermuscular fat) of Hanwoo (Korean native cattle) steers. From the 7,600 ESTs, 829 contigs comprising more than two EST reads were assembled using the Phrap assembler. Based on the contig analysis, 201 candidate cSNPs were identified in 129 contigs, in which transitions (69%) outnumbered transversions (31%). To verify whether the predicted cSNPs are real, 17 SNPs involved in lipid and energy metabolism were selected from the ESTs. Twelve of these were confirmed to be real while five were identified as artifacts, possibly due to expressed sequence tag sequence error. Further analysis of the 12 verified cSNPs was performed using the program BLASTX. Five were identified as nonsynonymous cSNPs, five were synonymous cSNPs, and two SNPs were located in 3'-UTRs. Our data indicated that a relatively high SNP prediction rate (71%) from a large EST database could produce abundant cSNPs rapidly, which can be used as valuable genetic markers in cattle.     

Cloning and characterization of an exoinulinase from Bacillus polymyxa

A gene encoding an exoinulinase (inu) from Bacillus polymyxa MGL21 was cloned and sequenced. It is composed of 1455 nucleotides, encoding a protein (485 amino acids) with a molecular mass of 55522 Da. Inu was expressed in Escherichia coli and the His-tagged exoinulinase was purified. The purified enzyme hydrolyzed sucrose, levan and raffinose, in addition to inulin, with a sucrose/inulin ratio of 2. Inulinase activity was optimal at 35°C and pH 7, was completely inactivated by 1 mM Ag⁺ or Hg²⁺. The K _m and V _max values for inulin hydrolysis were 0.7 mM and 2500 M min^–1 mg^–1 protein. The enzyme acted on inulin via an exo-attack to produce fructose mainly.     

Molecular cloning and characterization of vitelline precursor protein B1 from Clonorchis sinensis

In trematodes, vitelline precursor proteins are required for eggshell formation. A cDNA clone of Clonorchis sinensis (CsVpB1) was selected from an EST pool, encoding a polypeptide of 245 amino acids. The CsVpB1 polypeptide demonstrated homology with vitelline precursor proteins from trematodes with high sequential identities. In a phylogenic tree, CsVpB1 clustered with trematode VpB proteins. The CsVpB1 polypeptide was found to be rich in tyrosine residues, including putative predihydroxyphenyl alanine (DOPA) residues, involved in cross-linking of the precursor proteins. Mouse immune sera were raised against a recombinant CsVpB1 protein. In adult C. sinensis, CsVpB1 protein was exclusively localized in vitelline follicles. Based on these results, the CsVpB1 cDNA is believed to encode a VpB of C. sinensis.     

A human infection of Echinostoma hortense in duodenal bulb diagnosed by endoscopy

As gastroduodenoscopy performed more frequently, case reports of human echinostomiasis are increasing in Korea. A Korean woman presented at a local clinic with complaints of abdominal pain and discomfort that had persisted for 2 weeks. Under gastroduodenoscopy, two motile flukes were found attached on the duodenal bulb, and retrieved with endoscopic forceps. She had history of eating raw frog meat. The two flukes were identified as Echinostoma hortense by egg morphology, 27 collar spines with 4 end-group spines, and surface ultrastructural characters. This report may prove frogs to be a source of human echinostome infections.     

Crystal structures of 26 kDa Clonorchis sinensis glutathione S-transferase reveal zinc binding and putative metal binding

The crystal structures of CsGST in two different space groups revealed that Asp26 and His79 coordinate a zinc ion. In one space group, His46 of an adjacent molecule participates in the coordination within 2.0 Å. In the other space group, Asp26, His79 and a water molecule coordinate a zinc ion. The CsGST–D26H structure showed that four histidine residues – His26 and His79 from one molecule and the same residues from a symmetry-related neighboring molecule – coordinate a zinc ion. The coordinated zinc ions are located between two molecules and mediate molecular contacts within the crystal.