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ABSTRACT: The complete mitochondrial genome of Pseudochauhanea macrorchis was determined and compared with other monogenean mitochondrial genomes from GenBank. The circular genome was 15,031 bp in length and encoded 36 genes (12 protein-coding genes, two ribosomal RNAs, and 22 transfer RNAs) typically found in flatworms. Structures of the mitochondrial genome were mostly concordant with that known for Microcotyle sebastis and Polylabris halichoeres, but also contained two noted features-a gene rearrangement hot spot and the highly repetitive region (HRR) in major non-coding region (NCR). The gene rearrangement hot spot located between the cox3 and nad5 genes, including a cluster of tRNA genes, nad6 gene and one major NCR. The HRR seemed to be a unique feature of the polyopisthocotylean mitochondrial genomes. In conclusion, the present study provided new molecular data for future studies of the comparative mitochondrial genomics and also served as a resource of markers for the studies of species populations and monogenean phylogenetics.
Molecular Biology Reports 04/2012; 39(8):8115-25. · 2.93 Impact Factor
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ABSTRACT: This study presents the complete mitochondrial (mt) genome of Polylabris halichoeres, which is the largest mt genome sequenced of monogeneans so far and the second complete sequence after Microcotyle sebastis from the Microcotylidae. It is basically similar to that of M. sebastis, with the exception of a high level of gene rearrangement located between trnC and trnL((UUR)), a translocation of trnM and trnH, as well as a highly repetitive region (HRR) in the large non-coding region (NCR). We also find a series of trnI pseudogenes (ΨI) and one unknown short open reading frame (ORF) in the large NCR. Although the ORF cannot be unambiguously regarded as an atp8 gene, we cannot rule out the possibility that it has other functional importance, but it need further study in the future.
Mitochondrial DNA 06/2011; 22(1-2):3-5. · 1.49 Impact Factor
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ABSTRACT: The serum of rabbitfish (Siganus oramin) has been confirmed previously to have killing effect to Cryptocaryon irritans, an important marine ciliate protozoan that causes a disease referred to as "marine white spot disease". Herein, we find the serum of the rabbitfish also shows antibacterial activity against both gram-positive and gram-negative bacteria and has killing effect on two other parasites: Trypanosoma brucei brucei, Ichthyophthirius multifiliis. Results of scanning electron microscopy indicated that after treating with rabbitfish serum, the surface of the Staphylococcus aureus was wrinkled and pores were formed on the surface of Escherichia coli. Serum of the rabbitfish possesses a strong killing effect to Ichthyophthirius multifiliis in vitro, causing a similar effect as to C. irritans. The serum of rabbitfish also showed strong killing effect to T. b. brucei in vitro, with the minimus trypanocidal titre (MTT) only to be 1.5% in 1 h. Results of laser confocal fluorescence microscopy indicated that rabbitfish serum could also induce cell rupture of T. b. brucei. A novel antimicrobial protein (SR-LAAO) was isolated from the serum of rabbitfish by using ultrafiltration, reversed phase high performance liquid chromatography (RP-HPLC) and Native polyacrylamide gel electrophoresis (Native-PAGE). Results of gel overlay assay showed that the protein could act alone to inhibit the growth of S. aureus and E. coli. Results of western blot and automated Edman degradation showed that it was the same as the antiparasitic protein (APP) reported before to have killing effect on C. irritans. Full length cDNA sequence of the SR-LAAO was cloned. BLAST research suggested that the cDNA of SR-LAAO has a close similarity with a number of L-amino acid oxidases (LAAOs) and possesses two conserved motifs that exist in LAAOs. Combined, these results demonstrate that this protein which has antimicrobial activity to some pathogenic organisms was a novel LAAO found in the serum of rabbitfish. Immunohistochemical analysis demonstrated tissue specific expression and localization of SR-LAAO in the spleen, kidney, gill and blood of the rabbitfish, but was not found in other tissues. These results suggest that this protein may contribute considerably to the host non-specific immune defense mechanism to combat microbes of the rabbitfish and has the potency for using in future drug development.
Fish & Shellfish Immunology 02/2011; 30(4-5):1095-108. · 3.32 Impact Factor
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ABSTRACT: Streptococcus iniae is a major bacterium that causes invasive disease in cultured fish worldwide. The protection relies mainly on anti-microbial compounds and vaccines, and there is much interest in developing S. iniae vaccine based on conserved protein immunogens. Subcellular localization of protein has important influence on its immunogenicity. The surface and extracellular proteins of pathogenic bacteria can be easily recognized by the infected host compare to intracellular proteins, which are the feasible vaccine development targets. However, a putative hydrophobic membrane protein (designated MtsB) of the ATP-binding cassette (ABC) transporter system was found to be protective against S. iniae HD-1 infection when used as an injection vaccine administered intraperitoneally into tilapia. The MtsB protein is present on the cytoplasmic membrane and is expressed in vivo during Kunming mice infection by S. iniae HD-1. This is believed to be the first report on the use of a hydrophobic membrane protein of the ABC system as an S. iniae subunit vaccine.
Vaccine 01/2011; 29(3):391-4. · 3.77 Impact Factor
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ABSTRACT: Streptococcus iniae (S. iniae) is a major pathogen that causes considerable morbidity and mortality in cultured fish worldwide. The pathogen's ability to adapt to the host affects the extent of infection, hence understanding the mechanisms by which S. iniae overcomes physiological stresses during infection will help to identify potential virulence determinants of streptococcal infection. Grow S. iniae under iron-restricted conditions is one approach for identifying host-specific protein expression. Iron plays an important role in many biological processes but it has low solubility under physiological condition. Many microorganisms have been shown to be able to circumvent this nutritional limitation by forming direct contacts with iron-containing proteins through ATP-binding cassette (ABC) transporters. The ABC transporter superfamilies constitute many different systems that are widespread among living organisms with different functions, such as ligands translocation, mRNA translation, and DNA repair.
An ABC transporter system, named as mtsABC (metal transport system) was cloned from S. iniae HD-1, and was found to be involved in heme utilization. mtsABC is cotranscribed by three downstream genes, i.e., mtsA, mtsB, and mtsC. In this study, we cloned the first gene of the mtsABC transporter system (mtsA), and purified the corresponding recombinant protein MtsA. The analysis indicated that MtsA is a putative lipoprotein which binds to heme that can serve as an iron source for the microorganism, and is expressed in vivo during Kunming mice infection by S. iniae HD-1.
This is believed to be the first report on the cloning the ABC transporter lipoprotein from S. iniae genomic DNA. Together, our data suggested that MtsA is associated with heme, and is expressed in vivo during Kunming mice infection by S. iniae HD-1 which indicated that it can be a potential candidate for S. iniae subunit vaccine.
BMC Microbiology 01/2010; 10:309. · 3.04 Impact Factor
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ABSTRACT: SsHSP14.1, a novel sHSP from the hyper-thermophilic archaeon Sulfolobus solfataricus (S. solfataricus), is reported herein to function to protect EcoR I from heat-induced inactivation. A predicted salt bridge and hydrophobic interactions were found to be important for this function.
Protein and Peptide Letters 12/2009; 17(6):751-8. · 1.94 Impact Factor
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ABSTRACT: T-A cloning is the most commonly used molecular cloning technique in which T-vector is the core tool. T-vectors commonly used now are blue-white positive-selection vectors based on mechanism of alpha-complement of beta-galactose of Escherichia coli. Disadvantages of these traditional T-vectors in application include insufficient positive rate and the existence of false-positive clones, which, especially the latter, makes screening of transformant clones via colony PCR necessary. In the present study, a novel T-vector, based on the strategy of inhibiting the growth of negative transformants by means of a lethal gene and innovative design of insertion site, was developed. The innovative design of the insertion site for PCR fragments not only guarantees complete blocking of the expression of the lethal gene when PCR fragments are inserted, but also eliminates the formation of false-positive clones. All (100%) of 20 randomly chosen grown colonies were shown to be positive via colony PCR in four separate experimental T-A clonings of PCR fragments with lengths between 219 and 2100 bp. The novel T-vector developed in the present study makes colony PCR screening unnecessary, because of its extremely high efficiency of negative transformant inhibition and the mechanism for elimination of false-positive clones. Temperature (42 degrees C)-dependent positive selection greatly simplifies the procedure and lowers the cost of T-A cloning. Furthermore, it is an ideal T-vector for constructing libraries of PCR-amplified DNA fragments such as SSH (suppressive subtraction hybridization) libraries because of its zero negative background performance.
Biotechnology and Applied Biochemistry 04/2009; 53(Pt 4):247-51. · 1.53 Impact Factor
