Comparison of methods for library construction and short read annotation of shellfish viral metagenomes

Hong‑Ying Wei,Sheng Huang,Jiang‑Yong Wang,Fang Gao,Jing‑Zhe Jiang 한국유전학회 2018 Genes & Genomics Vol.40 No.3

The emergence and widespread use of high-throughput sequencing technologies have promoted metagenomic studies on environmental or animal samples. Library construction for metagenome sequencing and annotation of the produced sequence reads are important steps in such studies and influence the quality of metagenomic data. In this study, we collected some marine mollusk samples, such as Crassostrea hongkongensis, Chlamys farreri, and Ruditapes philippinarum, from coastal areas in South China. These samples were divided into two batches to compare two library construction methods for shellfish viral metagenome. Our analysis showed that reverse-transcribing RNA into cDNA and then amplifying it simultaneously with DNA by whole genome amplification (WGA) yielded a larger amount of DNA compared to using only WGA or WTA (whole transcriptome amplification). Moreover, higher quality libraries were obtained by agarose gel extraction rather than with AMPure bead size selection. However, the latter can also provide good results if combined with the adjustment of the filter parameters. This, together with its simplicity, makes it a viable alternative. Finally, we compared three annotation tools (BLAST, DIAMOND, and Taxonomer) and two reference databases (NCBI’s NR and Uniprot’s Uniref). Considering the limitations of computing resources and data transfer speed, we propose the use of DIAMOND with Uniref for annotating metagenomic short reads as its running speed can guarantee a good annotation rate. This study may serve as a useful reference for selecting methods for Shellfish viral metagenome library construction and read annotation.

Gut Microbiota Community and Its Assembly Associated with Age and Diet in Chinese Centenarians

( Fang Wang ),( Ting Yu ),( Guo Hong Huang ),( Da Cai ),( Xiaolin Liang ),( Hai Yan Su ),( Zhenjun Zhu ),( Danlei Li ),( Yang Yang ),( Pei Hong Shen ),( Rui Feng Mao ),( Lian Yu ),( Mou Ming Zhao ),( 한국미생물 · 생명공학회 2015 Journal of microbiology and biotechnology Vol.25 No.8

Increasing evidence suggests that gut microbiota underpin the development of health and longevity. However, our understanding of what influences the composition of this community of the longevous has not been adequately described. Therefore, illumina sequencing analysis was performed on the gut microbiota of centenarians (aged 100-108 years; RC) and younger elderlies (aged 85-99 years; RE) living in Bama County, Guangxi, China and the elderlies (aged 80-92 years; CE) living in Nanning City, Guangxi, China. In addition, their diet was monitored using a semiquantitative dietary questionary (FFQ 23). The results revealed the abundance of Roseburia and Escherichia was significantly greater, whereas that of Lactobacillus, Faecalibacterium, Parabacteroides, Butyricimonas, Coprococcus, Megamonas, Mitsuokella, Sutterella, and Akkermansia was significantly less in centenarians at the genus level. Both clustering analysis and UniFraq distance analysis showed structural segregation with age and diet among the three populations. Using partial least square discriminate analysis and redundancy analysis, we identified 33 and 34 operational taxonomic units (OTUs) as key OTUs that were significantly associated with age and diet, respectively. Age-related OTUs were characterized as Ruminococcaceae, Clostridiaceae, and Lachnospiraceae, and the former two were increased in the centenarians; diet-related OTUs were classified as Bacteroidales, Lachnospiraceae, and Ruminococcaceae. The former two were deceased, whereas the later one was increased, in the high-fiber diet. The age and high-fiber diet were concomitant with changes in the gut microbiota of centenarians, suggesting that age and high-fiber diet can establish a new structurally balanced architecture of gut microbiota that may benefit the health of centenarians.

Statistical Optimization of Culture Media and Conditions for Production of Mannan by Saccharomyces cerevisiae

Hong-Zhi Liu,Qiang Wang,Yuan-Yuan Liu,Fang Fang 한국생물공학회 2009 Biotechnology and Bioprocess Engineering Vol.14 No.5

In view of the increase in Saccharomyces cerevisiae mannan content, the culture medium and condition for S.cerevisiae were optimized in this study. The influence of culture medium ingredients such as carbon and nitrogen sources, inorganic ion, and enzyme activator on mannan production were evaluated using factional design. The mathematical model was established by the quadratic rotary combination design through response surface analysis. The optimized concentrations of culture medium were determined as follows: 4.98 g/100 mL, sucrose; 4.39 g/100 mL, soybean peptone; 3.10 g/100 mL, yeast extract; and 2.21 g/100 mL, glycerol. The optimized culture medium increased mannan production from 82.7 ± 3.4 mg/100 mL to 162.53 ± 3.47 mg/100 mL. The influence of original pH, inoculum size, temperature, and media volume on mannan production was evaluated and confirmed by orthogonale experimental design, with the order of effect as follows: media volume > temperature > initial pH > inoculation size. The optimized culture condition was pH, 5; inoculum size, 5 ml; temperature, 32oC; and media volume, 40 mL. The maximum mannan production increased to 258.5 ± 9.1 mg/100 mL at the optimum culture condition.=It was evident that the mannan production was affected significantly by culture medium and condition optimization (é < 0.01). In view of the increase in Saccharomyces cerevisiae mannan content, the culture medium and condition for S.cerevisiae were optimized in this study. The influence of culture medium ingredients such as carbon and nitrogen sources, inorganic ion, and enzyme activator on mannan production were evaluated using factional design. The mathematical model was established by the quadratic rotary combination design through response surface analysis. The optimized concentrations of culture medium were determined as follows: 4.98 g/100 mL, sucrose; 4.39 g/100 mL, soybean peptone; 3.10 g/100 mL, yeast extract; and 2.21 g/100 mL, glycerol. The optimized culture medium increased mannan production from 82.7 ± 3.4 mg/100 mL to 162.53 ± 3.47 mg/100 mL. The influence of original pH, inoculum size, temperature, and media volume on mannan production was evaluated and confirmed by orthogonale experimental design, with the order of effect as follows: media volume > temperature > initial pH > inoculation size. The optimized culture condition was pH, 5; inoculum size, 5 ml; temperature, 32oC; and media volume, 40 mL. The maximum mannan production increased to 258.5 ± 9.1 mg/100 mL at the optimum culture condition.=It was evident that the mannan production was affected significantly by culture medium and condition optimization (é < 0.01).

miR-638 is a new biomarker for outcome prediction of non-small cell lung cancer patients receiving chemotherapy

Fang Wang,Jian-fang Lou,Yan Cao,Xin-hui Shi,Peng Wang,Jian Xu,Er-fu Xie,Ting Xu,Rui-hong Sun,Jianyu Rao,Pu-wen Huang,Shi-yang Pan,Hong Wang 생화학분자생물학회 2015 Experimental and molecular medicine Vol.47 No.-

MicroRNAs (miRNAs), a class of small non-coding RNAs, mediate gene expression by either cleaving target mRNAs or inhibiting their translation. They have key roles in the tumorigenesis of several cancers, including non-small cell lung cancer (NSCLC). The aim of this study was to investigate the clinical significance of miR-638 in the evaluation of NSCLC patient prognosis in response to chemotherapy. First, we detected miR-638 expression levels in vitro in the culture supernatants of the NSCLC cell line SPC-A1 treated with cisplatin, as well as the apoptosis rates of SPC-A1. Second, serum miR-638 expression levels were detected in vivo by using nude mice xenograft models bearing SPC-A1 with and without cisplatin treatment. In the clinic, the serum miR-638 levels of 200 cases of NSCLC patients before and after chemotherapy were determined by quantitative real-time PCR, and the associations of clinicopathological features with miR-638 expression patterns after chemotherapy were analyzed. Our data helped in demonstrating that cisplatin induced apoptosis of the SPC-A1 cells in a dose- and time-dependent manner accompanied by increased miR-638 expression levels in the culture supernatants. In vivo data further revealed that cisplatin induced miR-638 upregulation in the serum derived from mice xenograft models, and in NSCLC patient sera, miR-638 expression patterns after chemotherapy significantly correlated with lymph node metastasis. Moreover, survival analyses revealed that patients who had increased miR-638 levels after chemotherapy showed significantly longer survival time than those who had decreased miR-638 levels. Our findings suggest that serum miR-638 levels are associated with the survival of NSCLC patients and may be considered a potential independent predictor for NSCLC prognosis.

Efficacy and Survival-associated Factors with Gefitinib Combined with Cisplatin and Gemcitabine for Advanced Non-small Cell Lung Cancer

Fang, Hong,Lin, Rong-Yan,Sun, Ming-Xia,Wang, Qian,Zhao, Yu-Liang,Yu, Jing-Lin,Tian, Yan,Wang, Xiao-Yun Asian Pacific Journal of Cancer Prevention 2014 Asian Pacific journal of cancer prevention Vol.15 No.24

Objective: To analyze the efficacy and survival associated factors of gefitinib combined with cisplatin and gemcitabine for advanced non-small cell lung cancer. Materials and Methods: A total of 57 patients with advanced non-small cell lung cancer (NSCLC), who received platinum-based chemotherapy regimens for more than 1 cycle, were treated with gefitinib combined with cisplatin and gemcitabine until disease progression. Efficacy, survival time and adverse reactions were observed. The Kaplan-Meier method was adopted for analysis of survival and Cox regression for associated influencing factors. Results: The patients were followed up until October 31, 2013, and the median follow-up time was 19 months. Of 57 patients, there were 4 (7.0%) with complete remission (CR), 8 (14.0%) with partial remission, 31 (54.4%) with stable disease, and 14 (24.6%) with disease progression. The remission rate was 21.1% and the disease control rate was 75.4%. The median progression-free survival (PFS) time and the median overall survival time were 10 months and 15.2 months. The one-year, two-year and three-year survival rates were 47.4%, 23.3% and 10.0%. Gender and pathological types were the independent risk factors influencing PFS time (P=0.028, P=0.009). Tumor pathological type and early efficacy were independent factors for the prognosis (P=0.018, P=0.000). Adverse reactions were mostly rashes of I~II degree and diarrhea and slightly increasing level of aminopherase. The skin adverse event incidence of III degree or above was 1.8% (1/57) and brain metastasis was foudn in 31.6% (18/57). Conclusions: Gefitinib combined with cisplatin andgemcitabine, is effective for patients with IIIb~IV NSCLC who received multiple cycles of chemotherapy.

Complete mitochondrial genome of the meadow moth, Loxostege sticticalis (Lepidoptera: Pyraloidea: Crambidae), compared to other Pyraloidea moths

Hong-Fang Ma,Xi-Xi Zheng,Ming-Hui Peng,Hai-Xu Bian,Miao-Miao Chen,Yan-Qun Liu,Xing-Fu Jiang,Li Qin 한국응용곤충학회 2016 Journal of Asia-Pacific Entomology Vol.19 No.3

The mitochondrial genome (mitogenome) is an important topic for comparative and evolutionary genomics, as well as phylogenetic and population genetics. However, there are limited data regarding the mitochondrial genome available of Pyraloidea, one of the largest superfamilies in Lepidoptera. In this report, we present the complete mitogenome of the meadow moth, Loxostege sticticalis L. (Lepidoptera: Pyraloidea: Crambidae), which is a serious economic pest of both crops and weedsworldwide, thereby enhancing the available genomic information for Pyraloidea. This circular genome is 15,218 bp in length, containing 13 protein-coding genes (PCGs), two rRNA genes (rRNAs), and 22 tRNA genes (tRNAs), with a typical gene orientation and order comparable to other sequenced Pyraloidea insects. The genome composition of the major strand exhibits highly AT bias (80.82%), with a slightly positive AT skew indicating the occurrence of more As than Ts. The L. sticticalis mitogenome has a total of 130 bp of intergenic spacer sequences spread over 15 regions, ranging in size from 1 to 48 bp, of which only two are common among the 23 total Pyraloidea moths that have data collected on the mitogenome (one is located between tRNAGln and ND2 with variation change in length and a limited sequence conservation, and the other is located between tRNASer(UCN) and ND1 with a conserved 6 bp motif ‘ATACTA’). The A + Trich region of 331 bp in the genome is comprised of non-repetitive sequences but contains an ATAGN motif followed by a poly-T stretch of 17 bp, a microsatellite-like (TA)11 element preceded by an ATTTA motif, and a poly-A stretch upstream tRNAMet. These conserved structures identified in the A + T-rich region are presented in all of the sequenced Pyraloidea species. We provide a mitogenome-based phylogeny of Pyraloidea species, in which L. sticticalis shares close ancestry to Ostrinia species with substantial evidence. Our phylogenetic analyses strongly divide Crambidae into two sister lineages, one consisting of Pyraustinae and Spilomelinae, while the other contains Crambinae, Acentropinae, Scopariinae, Schoenobiinae and Glaphyriinae. The mitogenome dataset also supports the basal split between Pyraustinae and Spilomelinae.

Morphology of the immature stages of Adoretus tenuimaculatus Waterhouse, 1875 (Coleoptera: Scarabaeidae: Rutelinae: Adoretini)

Hong-Fang Ma,Ce Li,Lu Jiang 한국응용곤충학회 2018 Journal of Asia-Pacific Entomology Vol.21 No.4

The immature stages of the brown chafers Adoretus tenuimaculatus Waterhouse, 1875 were investigated morphologically using scanning electron microscopy. The scarabaeiform larvae of A. tenuimaculatus are peculiar for bearing a pair of eye spots on the head capsule, devoid of palidium but numerous hamate setae on the raster. The larvae possess a crown-like apical sensory area on each antenna, maxillary and labial palp. The pupa possesses dorsally six pairs of dioneiform organs on the intersegment regions between the first seven abdominal segments. This study provides the first ultra morphological study of the larvae in Adoretini. Both advantages and disadvantages of the scanning electron microscopy in the larval taxonomy of Scarabaeidae were briefly discussed.

Effects of cashew nut testa levels as an alternative to wheat bran in gestating sow diets

Fang, Lin Hu,Hong, Young Gi,Hong, Jin Su,Jeong, Jae Hark,Han, Young Geol,Kwon, In Hyuk,Kim, Yoo Yong Asian Australasian Association of Animal Productio 2018 Animal Bioscience Vol.31 No.6

Objective: This study was conducted to evaluate the effects of dietary cashew nut testa (CNT) as an alternative feed ingredient to wheat bran on reproductive performance, litter performance, milk composition, and blood profiles of gestating sows. Methods: Forth multiparous sows ($Yorkshire{\times}Landrace$) were fed experimental diets starting at 35 days of pregnancy and an initial average body weight (BW) of $211.53{\pm}8.86kg$. Each sow was assigned to a treatment based on BW, backfat thickness (BF) and parity with 10 sows per treatment. Treatments were as follows: i) corn-soybean meal based diet with 6% of wheat bran (C0); ii) basal diet with 2% of CNT and 4% of wheat bran (C2); iii) basal diet with 4% of CNT and 2% of wheat bran (C4); and iv) basal diet with 6% of CNT (C6). Results: There were no statistically significant differences in BW and BF of gestating sows throughout the experimental period. However, changes in BF (p = 0.09) and the daily feed intake of sows (p = 0.09) tended to linearly increase during the lactation period. The weaning to estrus interval (WEI) showed a quadratic response to CNT treatment (p = 0.02), and the C2 diet showed the shortest WEI. Litter birth weight (p = 0.04) and piglet birth weight (p = 0.06) were linearly decreased with increase in CNT. Furthermore, there had no significant differences in piglet weight and litter weight in 21 day. Insulin concentration at day 70 of gestation was linearly reduced with increasing CNT level in diets (p = 0.03). Conclusion: When 6% CNT replaced wheat bran in gestating sow diets, there were no negative effects on sow performance, but litter birth weight and piglet birth weight were decreased when CNT level increased in gestating sow diets.

CD-200 induces apoptosis and inhibits Bcr-Abl signaling in imatinib-resistant chronic myeloid leukemia with T315I mutation

FANG, ZHENGHUAN,JUNG, KYUNG HEE,YAN, HONG HUA,KIM, SOO JUNG,SON, MI KWON,RUMMAN, MARUFA,LEE, HYUNSEUNG,KIM, KI WOON,YOO, HYE-DONG,HONG, SOON-SUN Spandidos Publications 2015 International journal of oncology Vol.47 No.1

<P>Chronic myeloid leukemia (CML) is characterized by a constitutively active Bcr-Abl tyrosine kinase. Although Imatinib has been proven to be an effective drug against CML, its resistance has been observed with disease relapse due to T315I predominant point mutation. Liriodendron tulipifera L., one of the fastest growing hardwood tree species, exerts antioxidant activity and anti-inflammatory effects. However, its anticancer effect has been minimally reported. In this study, we extracted CD-200 from Liriodendron tulipifera L. and investigated its effect on cell survival or apoptosis in CML cells with Bcr-Abl/T315I (BaF3/T315I) as well as wild-type Bcr-Abl (BaF3/WT). CD-200 inhibited cell proliferation in the BaF3/WT cells, and also in the BaF3/T315I cells with Imatinib resistance. Moreover, it strongly inhibited Bcr-Abl signaling pathways in a dose-dependent manner. Also, it significantly increased the sub-G1 phase and the expression of cleaved PARP and caspase-3, as well as the TUNEL-positive apoptotic cells. In addition, we observed that CD-200 induced apoptosis with a loss of mitochondrial membrane potential by decreasing the expression of Mcl-1 and survivin. Furthermore, CD-200 showed a significant inhibition in tumor growth, compared to Imatinib in BaF3/T315I mouse xenograft models. Taken together, our study demonstrates that CD-200 exhibits apoptosis induction and anti-proliferative effect by blocking the Bcr-Abl signaling pathways in the Bcr-Abl/T315I with resistance to Imatinib. We suggest that CD-200 may be a natural product to target Bcr-Abl and overcome Imatinib resistance in CML patients.</P>

The Effectiveness of Two Hospitals Implementing Magnet Hospital Concept to Improve Nursing Staff Retention

Fang Chih Yeh,Li Fang Liu,Nai Hong Wang 한국간호과학회 2009 한국간호과학회 학술대회 Vol.2009 No.10