ALBERT

Description: Background: Streptococcus equi ssp. zooepidemicus (S. zooepidemicus) is an important pathogen causing swine streptococcosis in China. Pathogenicity islands (PAIs) of S. zooepidemicus have been transferred among bacteria through horizontal gene transfer (HGT) and play important roles in the adaptation and increased virulence of S. zooepidemicus. The present study used comparative genomics to examine the different pathogenicities of S. zooepidemicus. Results: Genome of S. zooepidemicus ATCC35246 (Sz35246) comprises 2,167,264-bp of a single circular chromosome, with a GC content of 41.65%. Comparative genome analysis of Sz35246, S. zooepidemicus MGCS10565 (Sz10565), Streptococcus equi. ssp. equi. 4047 (Se4047) and S. zooepidemicus H70 (Sz70) identified 320 Sz35246-specific genes, clustered into three toxin-antitoxin (TA) systems PAIs and one restriction modification system (RM system) PAI. These four acquired PAIs encode proteins that may contribute to the overall pathogenic capacity and fitness of this bacterium to adapt to different hosts. Analysis of the in vivo and in vitro transcriptomes of this bacterium revealed differentially expressed PAI genes and non-PAI genes, suggesting that Sz35246 possess mechanisms for infecting animals and adapting to a wide range of host environments. Analysis of the genome identified potential Sz35246 virulence genes. Genes of the Fim III operon were presumed to be involved in breaking the host-restriction of Sz35246. Conclusion: Genome wide comparisons of Sz35246 with three other strains and transcriptome analysis revealed novel genes related to bacterial virulence and breaking the host-restriction. Four specific PAIs, which were judged to have been transferred into Sz35246 genome through HGT, were identified for the first time. Further analysis of the TA and RM systems in the PAIs will improve our understanding of the pathogenicity of this bacterium and could lead to the development of diagnostics and vaccines.

Description: Background: Penicillium digitatum is one of the most destructive postharvest pathogen of citrus fruits, causing fruit decay and economic loss. The emergence of fungicide-resistant strains made the control of P. digitatum more difficult. While the genome of P. digitatum is available, there has been few reports about its resistant mechanism from the transcriptome perspective and there has been no large-scale functional annotation of the genome using expressed genes derived from transcriptomes. Methods: Total RNA of P. digitatum strain HS-F6 (prochloraz-resistant strain) and HS-E3 (prochloraz-susceptible strain) before and after prochloraz-treatment were extracted and sequenced on an Illumina Hiseq 2000 platform. The transcriptome data of four samples were compared and analyzed using differential expression analysis, novel transcripts prediction and alternative splicing analysis, SNP analysis and quantitative real-time PCR. Results: We present a large scale analysis about the transcriptome data of P. digitatum. The whole RNA was extracted from a prochloraz-resistant strain (HS-F6) and a prochloraz-susceptible strain (HS-E3) before and after prochloraz-treatment and sequenced by Illumina technology. A total of more than 100 million reads were generated and de novo assembled into 9760 transcripts that contained annotated genes after quality control and sequence assembling. 6625 single nucleotide variations (SNVs) were identified from the sequences aligned against the reference genome. Gene expression profiling analysis was performed upon prochloraz treatment in HS-F6 and HS-E3, and differential expression analysis was used to identify genes related to prochloraz-response and drug-resistance: there are 224 differentially expressed genes in HS-E3 and 1100 differentially expressed genes in HS-F6 after prochloraz-treatment. Moreover, gene expression profile in prochloraz-resistant strain HS-F6 is quite different from that in HS-E3 before prochloraz-treatment, 1520 differential expression genes were identified between the two strains. Gene ontology (GO) term enrichment and KEGG enrichment were then performed to classify the differential expression genes. Among these genes, there are a lot of transporter encoding genes including 14 MFS (Major Facilitator Superfamily) transporters, 8 ABC (ATP-binding cassette transporter) and 3 MATE (multidrug and toxic compound extrusion family) transporters. Meanwhile, the roles of typical MFS, ABC and MATE proteins in prochloraz resistance were investigated using real-time quantitative PCR. Conclusions: The sequencing-based transcriptome data of P. digitatum demonstrate differences between prochloraz-resistant and prochloraz-susceptible strains with prochloraz-treatment. The differences existed in expressed transcripts, splice isoforms and GO categories, which would contribute to our knowledge on the molecular mechanisms involved in drug resistance of P. digitatum.

Description: Grain development in maize is an essential process in the plant’s life cycle and is vital for use of the plant as a crop for animals and humans. However, little is known regarding the protein regulatory networ...

Description: Background: Cranberries (Vaccinium macrocarpon Ait.), renowned for their excellent health benefits, are an important berry crop. Here, we performed transcriptome sequencing of one cranberry cultivar, from fruits at two different developmental stages, on the Illumina HiSeq 2000 platform. Our main goals were to identify putative genes for major metabolic pathways of bioactive compounds and compare the expression patterns between white fruit (W) and red fruit (R) in cranberry. Results: In this study, two cDNA libraries of W and R were constructed. Approximately 119 million raw sequencing reads were generated and assembled de novo, yielding 57,331 high quality unigenes with an average length of 739 bp. Using BLASTx, 38,460 unigenes were identified as putative homologs of annotated sequences in public protein databases, including NCBI NR, NT, Swiss-Prot, KEGG, COG and GO. Of these, 21,898 unigenes mapped to 128 KEGG pathways, with the metabolic pathways, secondary metabolites, glycerophospholipid metabolism, ether lipid metabolism, starch and sucrose metabolism, purine metabolism, and pyrimidine metabolism being well represented. Among them, many candidate genes were involved in flavonoid biosynthesis, transport and regulation. Furthermore, digital gene expression (DEG) analysis identified 3,257 unigenes that were differentially expressed between the two fruit developmental stages. In addition, 14,473 simple sequence repeats (SSRs) were detected. Conclusions: Our results present comprehensive gene expression information about the cranberry fruit transcriptome that could facilitate our understanding of the molecular mechanisms of fruit development in cranberries. Although it will be necessary to validate the functions carried out by these genes, these results could be used to improve the quality of breeding programs for the cranberry and related species.

Description: Patients over age 60 maked up more than 50% of newly diagnosed patients with acute myeloid leukemia (AML). Futhermore, with an aging population, more and more older AML patients were diagnosed in China. But the treatment approaches of this disease were variable, with many uncertainties and controversies. Treatment options for older patients with adverse prognostic features, such as poor performance status, unfavorable cytogenetics or an antecedent hematologic disorder were limited, and outcomes were poor. Aggres­sive induction chemotherapy had a high mortality and relatively low efficacy in this population. There were several new therapeutic schemes for older patients with AML. CAG regimens consisting of low-dose cytarabine, aclarubicin and granulocyte-colony stimulating factor for the treatment of older patients with AML showed higher rates of CR (42-68%). Decitabine, a DNA-hypomethylating agent induces differentiation and apoptosis of leukemic cells. The current National Comprehensive Cancer Care guidelines suggested decitabine as alterative options for older patients with AML. Previous studies have shown that decitabine demonstrated efficacy in a phase II multicenter study of older patients with AML, with a CR rate of 25%, 30-day mortality of 7%, median overall survival 7.7 months and little extramedullary toxicity. In our study, decitabine(15 mg/m2/d, d1-5) combined with CAG regimens (aclarubicin 20 mg/d, d3-6, Ara-C 10 mg/m2, q12h, d3-9, G-CSF 300ug, qd, d1-9) treated 27 older patients with AML, repeated every 4 weeks. Effectiveness and safety were assessed. 27 older patients with newly diagnosed AML who were in Tianjin First Central Hospital of China from January 2011 to December 2013 were enrolled in our study. They were all treated with decitabine combined with CAG regimens. The characteristics of the 27 patients were described in Table I. The study population included 15 males and 12 females, with a median age of 68 years (range 60-79 years). All patients had Eastern Cooperative Oncology Group (ECOG) performance status of

Description: Abstract 4247 Iron overload is caused by multiple blood transfusion and excess gastrointestinal absorption, leading to most of the mortality and morbidity associated with anemia diseases (i.e. aplastic anemia, myelodysplastic syndromes, thalassemia and myelofibrosis). It can cause tissue damage and ultimately dysfunction of visceral organs (mainly in the heart, liver, and endocrine glands). Nevertheless, it is unknown whether iron overload affects the hematopoiesis of bone marrow (BM). In recent years, a number of papers reported that iron chelation therapy could enhance the erythropoiesis and even reduce the cytopenia in iron overload anemia (i.e. myelodysplastic syndromes and myelofibrosis). As it is well known that iron overload could increase the production of reactive oxygen species (ROS) in the tissue, and also ROS could affect the hematopoiesis of BM, we hypothesized that iron overload could increase the ROS of BM, and then result in the deficient hematopoiesis. To confirm this hypothesis, we studied the relationship between ROS and the hematopoiesis of iron overload BM. The intracellular concentration of ROS in cells were analyzed by a flow cytometer after incubated with 50 μM (final concentration) 2′–7′-dichlorofluorescin diacetate (DCF; Sigma) for 10 min at 37°C in a humidified atmosphere of 5% CO2 in air. We found that the ROS in the hematopoietic cells of BM from 26 patients with iron overload (16 cases with myelodysplastic syndromes and 10 cases with myelofibrosis) was much higher than that of normal control (p

Description: Interleukin 21(IL-21) is a new member of interleukin 2 cytokine families which was discovered in 2000. IL-21 is produced by activated CD4 positive cells, and is known to influence T, B, NK cells and DC, and has potent anti-tumor effects. For example, IL-21 can improve the proliferation of B lymphocytes, enhance the production of IgG1; improve the proliferation and enhance the anti-tumor activity of both NK and T cells. The cytokine-induced killer (CIK) cells, which are characterized with the phenotype of CD3+CD56+, are the effective cells on adoptive cellular immunotherapy against tumors. We hypothesize that IL-21 could also affect the proliferation and function of CIK cells, thus play a certain role in the anti-tumor immunotherapy by CIK cells. The peripheral blood mononuclear cells (PBMC) and cord blood mononuclear cells (CBMC) from healthy donors were stimulated with anti-CD3 (OKT3) monoclonal antibody and IFNgamma and then expanded with IL-2 and with/without IL-21(200ng/ml). CD3+CD56+ CIK cells were counted by flow cytometry. Net lactate dehydrogenase release from target cells incubated with CIK cells was used as an index of CIK cells cytotoxicity against chronic myeloid leukemia cell line K562 and a variety of tumor target cells from patients. The concentration of the IFNgamma in the culture supernatant was measured by enzyme-linked-immunoassay, the quantity of IFNgamma RNA was measured by RT-PCR assay, and the cytotoxic activity against K562 cells by the culture supernatant was also detected. Cultured with IL-21, at day 14, the quantity of CIK cells was increased from a median of 17.5% to 26.5% (PBMC original) and from 33.8% to 55.9% (CBMC original); The cytotoxic activity rates against K562 cells by CIK cells were increased from 24.0% to 52.2% (PBMC original) and from 35.1% to 79.7% (CBMC original); The concentration of IFNgamma in the culture supernatant was increased for 1.9-fold (PBMC original), and for 3.2-fold (CBMC original); The cytotoxic activity against K562 cells by the culture supernatant was increased for 1.8-fold (PBMC original) and for 2.7-fold (CBMC original); The expression of IFNgamma RNA in CIK cells was also markedly increased derived from both PBMC and CBMC when cultured with IL-21. Moreover, the cytotoxic activity against leukemia cells from 11 patients (6 with acute lymphoblastic leukemia, 5 with acute myeloid leukemia) by CIK cells derived from CBMC were also detected. The cytotoxic activity rates were at a median of 68.3% (range, 34.7%–86.4%) when CIK cells were cultured with IL-21, rates that contrasted drastically to the cytotoxic activity rates when CIK cells were cultured without IL-21, which were only at a median of 37.4% (range, 16.1%–60.0%). In conclusion, our data indicated that IL-21 could enhance the expansion of CIK cells and their anti-tumor activity derived from both PBMC and CBMC in vitro, IFNgamma was evolved in this course although the mechanism still need to be explored. These observations open up the possibility of imagining a future clinical application of IL-21 in the anti-tumor immunotherapy by CIK cells.