Abstract. Reemerged Plasmodium vivax malaria in South Korea has not yet been eradicated despite continuous governmental efforts. It has rather become an endemic disease. Our study aimed to determine the genetic diversity in P. vivax merozoite surface protein-1 (PvMSP-1) and circumsporozoite protein (PvCSP) genes over an extended period after its reemergence to its current status. Sequence analysis of PvMSP-1 gene sequences from the 632 P. vivax isolates during 1996-2007 indicates that most isolates recently obtained were different from isolates obtained in the initial reemergence period. There was initially only one subtype (recombinant) present but its subtypes have varied since 2000; six MSP-1 subtypes were recently found. A similar variation was observed by CSP gene analysis; a new CSP subtype was found. Understanding genetic variation patterns of the parasite may help to analyze trends and assess extent of endemic malaria in South Korea.

A cystine-dependent anti-oxidative stress response is characterized in Deinococcus geothermalis for the first time. Nevertheless, the same transcriptional directed Δdgeo_1985F mutant strain was revealed to have an identical phenotype to the wild-type strain, while the reverse transcriptional directed Δdgeo_1985R mutant strain was more resistant to oxidative stress at a certain concentration of HO than the wild-type strain. The wild-type and mutant strains expressed equal levels of superoxide dismutase and catalase under HO-induced stress. Although the expression levels of the general DNA-damage response-related genes recA, pprA, ddrA, and ddrB were up-regulated by more than five-fold in the wild-type strain relative to the Δdgeo_1985R mutant strain, the mutant strain had a higher survival rate than the wild-type under HO stress. The Δdgeo_1985R mutant strain highly expressed a cystine-transporter gene (dgeo_1986), at levels 150-fold higher than the wild-type strain, leading to the conclusion that this cystine transporter might be involved in the defensive response to HO stress. In this study, the cystine transporter was identified and characterized through membrane protein expression analysis, a cystine-binding assay, and assays of intracellular HO, cysteine, and thiol levels. The genedisrupted mutant strain of the cystine importer revealed high sensitivity to HO and less absorbed cystine, resulting in low concentrations of total thiol. Thus, the absorbed cystine via this cystine-specific importer may be converted into cysteine, which acts as a primitive defense substrate that non-enzymatically scavenges oxidative stress agents in D. geothermalis.

BackgroundTo develop a plant-based vaccine against Plasmodium vivax, two P. vivax candidate proteins were chosen. First, the merozoite surface protein-1 (MSP-1), a major asexual blood stage antigen that is currently considered a strong vaccine candidate. Second, the circumsporozoite protein (CSP), a component of sporozoites that contains a B-cell epitope.MethodsA synthetic chimeric recombinant 516 bp gene encoding containing PvMSP-1, a Pro-Gly linker motif, and PvCSP was synthesized; the gene, named MLC, encoded a total of 172 amino acids. The recombinant gene was modified with regard to codon usage to optimize gene expression in Brassica napus. The Ti plasmid inducible gene transfer system was used for MLC chimeric recombinant gene expression in B. napus. Gene expression was confirmed by polymerase chain reaction (PCR), beta-glucuronidase reporter gene (GUS) assay, and Western blot.ResultsThe MLC chimeric recombinant protein expressed in B. napus had a molecular weight of approximately 25 kDa. It exhibited a clinical sensitivity of 84.21% (n = 38) and a clinical specificity of 100% (n = 24) as assessed by enzyme-linked immunosorbent assay (ELISA). Oral immunization of BALB/c mice with MLC chimeric recombinant protein successfully induced antigen-specific IgG1 production. Additionally, the Th1-related cytokines IL-12 (p40), TNF, and IFN-γ were significantly increased in the spleens of the BALB/c mice.ConclusionsThe chimeric MLC recombinant protein produced in B. napus has potential as both as an antigen for diagnosis and as a valuable vaccine candidate for oral immunization against vivax malaria.

A BLASTP search has shown the presence of a gene homologous to an alternative thymidylate synthase (TS), thyX, in Corynebacterium glutamicum ATCC 13032. To determine if thyX is functionally analogous to thyA, thyX was cloned in a plasmid and the resulting construct was transferred by transformation into a thyA mutant of Escherichia coli. The ThyX from C. glutamicum compensated for the defect in TS-deficient E. coli. A functional knockout of the thyX gene was constructed by allelic replacement using a sucrose counter-selectable suicide plasmid and confirmed by PCR and reverse transcriptase-PCR analyses. This mutant was viable without thymidine supplementation, suggesting that thyX is not an essential gene in C. glutamicum. Growth of the thyX mutant was dependent upon coupling activity of dihydrofolate reductase (DHFR) with ThyA for the synthesis of thymidine, and thus showed sensitivity to the inhibition of DHFR by the experimental inhibitor, WR99210. This indicates that thymidine synthesis was at least partially dependent on thyX expression. As it approached stationary phase, the thyX mutant lost viability much more rapidly than the parental wild type and the mutant complemented the thyX gene, suggesting that the activity of the ThyX enzyme is important in that phase of the growth cycle.

The csgA gene encodes an extracellular protein that plays an essential role in the regulation of fruiting-body formation and sporulation of Myxococcus xanthus. The csgA suppressor allele soc-500 (formerly referred to as csp-500) was selected based on its ability to restore sporulation to csgA cells under developmental conditions at 32 degrees C. The soc-500 allele was subsequently found to induce sporulation of csgA+ or csgA cells simply by shifting the temperature of vegetatively growing cells to 15 degrees C. Low-temperature-induced sporulation of soc-500 strains occurred in the absence of two requirements for fruiting-body sporulation: low nutrient levels and a high temperature. Low temperature alone caused the expression of many developmentally regulated genes but did not support the development of wild-type cells. The soc-500 allele appears to activate genes involved with sensing nutritional stress. At low temperature on a nutritionally rich medium, soc-500 induced expression of the tps gene which is normally expressed following nutritional shiftdown. The soc-500 allele was cloned and integrated into the wild-type chromosome by site-specific recombination. It was dominant over the wild-type allele in merodiploids and is contained on a 3-kbp DraI-ClaI restriction fragment. The soc-500 transcriptional unit spans a 300-bp PstI-PstI restriction fragment, since deletion of the PstI restriction fragment inhibits both csgA suppression and low-temperature induction. These results suggest that the soc-500 mutation lies in a gene that is involved in nutrient sensing.