cano-wgMLST_BacCompare: A Bacterial Genome Analysis Platform for Epidemiological Investigation and Comparative Genomic Analysis.

With the decreasing cost of next-generation sequencing, whole-genome sequence-based bacterial genome comparisons are expected to become a mainstream process in the public health domain. Extended multilocus sequence typing (MLST) methods are becoming increasingly popular for use in comparing bacterial genetic relatedness in epidemiological investigations. Several extended MLST schemes based on biological signatures have been reported. Among them, whole-genome MLST (wgMLST) has gradually become one of the most widely used approaches for bacterial strain typing. In addition to using bacterial typing, many researchers aim to identify differences in the genes of compared strains. Because these differences might provide insights into critical bacterial functions, such as virulence and pathogenicity, researchers usually study these genes that differ between strains. Hence, we designed a web service tool based on wgMLST-constructed tree topology coupled with the feature selection method to create the "canonical wgMLST (cano-wgMLST) tree." The genes that differ between strains are shown at each split of the tree, thereby directly providing information for performing a comparative genomic analysis for each strain pair. We demonstrated that this web service tool could be operated efficiently on two datasets consisting of 22 Campylobacter jejuni isolates and 59 S. Heidelberg isolates, respectively. We imposed this tool on a designated web server, cano-wgMLST_BacCompare, to enable users to create a wgMLST tree and canonical wgMLST tree automatically from their uploaded bacterial genomes for not only epidemiological investigation but also comparative genomic analysis. Additionally, detailed information on how to use this service is provided. The cano-wgMLST_BacCompare is available at http://baccompare.imst.nsysu.edu.tw.

The Cano-eMLST Program: An Approach for the Calculation of Canonical Extended Multi-Locus Sequence Typing, Making Comparison of Genetic Differences Among Bunches of Bacterial Strains.

Extended multi-locus sequence typing (eMLST) methods have become popular in the field of genomic epidemiology. Before eMLST methods can be applied in epidemiological investigations, the selection of a suitable scheme is critical. The core genome scheme (cgMLST) has become the most popular eMLST approach for strain typing in the epidemiological domain. In addition to strain typing, many public health researchers and clinical microbiologists wish to investigate which genes cause genetic differences between compared strains. Therefore, a tool that can be used to extract canonical genes with an eMLST scheme would be particularly useful. In this study, we present cano-eMLST, a well-designed program that applies a feature-selection methodology to create a canonical locus combination with discriminatory power by traversing a genetic relatedness tree based on a user-selected scheme. The cano-eMLST program is provided mainly to help infectious disease laboratory researchers identify potential factors related to bacterial pathogenesis. The core program (tree-traversing approach) of cano-eMLST is implemented in Perl and Python. All the necessary dependencies and environmental settings are provided in the encapsulated version (VirtualBox or VMware) and self-installation version (all use source code and libraries).

Acute exacerbation of hepatitis C in hepatocellular carcinoma patients receiving chemotherapy.

Acute hepatitis C exacerbations can occur in cancer patients carrying hepatitis C virus (HCV) when receiving systemic chemotherapy. However, clinical studies evaluating these complications remain rare due to the lack of clinically proven effective and tolerable anti-HCV treatments at late cancer stages. Furthermore, no data were available regarding hepatitis C exacerbation in advanced hepatocellular carcinoma (HCC) patients receiving chemotherapy. To address this issue, 48 patients with HCV-related advanced HCC, who underwent systemic chemotherapy using 5- fluorouracil, cisplatin, and mitoxantrone from 2008 to 2014 were analyzed. Nine patients developed acute hepatitis exacerbations defined by HCV-RNA elevation ≥10-fold and alanine transaminase (ALT) elevation ≥5-fold of the upper normal limit. Six were genotype 1b and 3 were genotype 2. Three patterns of clinical courses were observed including single episode of exacerbation (n = 5), fluctuated flares (n = 3), and delayed exacerbation (n = 1). Hepatic failure developed in five patients. Patients with acute exacerbations were less likely to have pretreatment ascites (11.1% vs. 53.8%; P = 0.028) and displayed a lower baseline ALT (44.1 ± 28.5 U/L vs. 72.6 ± 19.2 U/L; P = 0.007). Paradoxically, despite a high risk of hepatic failure, occurrence of hepatitis C exacerbation was associated with a favorable overall survival (P = 0.027; 22.8 vs. 5.4 months). In conclusion, hepatitis C exacerbation can occur in HCC patients receiving chemotherapy, leading to liver failure. However, the flare was associated with a better overall survival, possibly due to its association with a better baseline liver function. J. Med. Virol. 89:153-160, 2017. © 2016 Wiley Periodicals, Inc.

Combining radiofrequency ablation and ethanol injection may achieve comparable long-term outcomes in larger hepatocellular carcinoma (3.1-4 cm) and in high-risk locations.

Radiofrequency ablation (RFA) is more effective for hepatocellular carcinoma (HCC) < 3 cm. Combining percutaneous ethanol injection and RFA for HCC can increase ablation; however, the long-term outcome remains unknown. The aim of this study was to compare long-term outcomes between patients with HCC of 2-3 cm versus 3.1-4 cm and in high-risk versus non-high-risk locations after combination therapy. The primary endpoint was overall survival and the secondary endpoint was local tumor progression (LTP). Fifty-four consecutive patients with 72 tumors were enrolled. Twenty-two (30.6%) tumors and 60 (83.3%) tumors were of 3.1-4 cm and in high-risk locations, respectively. Primary technique effectiveness was comparable between HCC of 2-3 cm versus 3.1-4 cm (98% vs. 95.5%, p = 0.521), and HCC in non-high risk and high-risk locations (100% vs. 96.7%, p = 1.000). The cumulative survival rates at 1 year, 3 years, and 5 years were 90.3%, 78.9%, and 60.3%, respectively, in patients with HCC of 2-3 cm; 95.0%, 84.4%, and 69.3% in HCC of 3.1-4.0 cm (p = 0.397); 90.0%, 71.1%, and 71.1% in patients with HCC in non-high-risk locations; and 92.7%, 81.6%, and 65.4% in high-risk locations (p = 0.979). The cumulative LTP rates at 1 year, 3 years, and 5 years were 10.2%, 32.6%, and 32.6%, respectively, in all HCCs; 12.6%, 33.9%, and 33.9% in HCC of 2-3 cm; 4.8%, 29.5%, and 29.5% in HCC of 3.1-4 cm (p = 0.616); 16.7%, 50.0%, and 50.0% in patients with HCC in non-high-risk locations; and 8.8%, 29.9%, and 29.9% in patients with HCC in high-risk locations (p = 0.283). The cumulative survival and LTP rates were not significantly different among the various subgroups. Combining RFA and percutaneous ethanol injection achieved comparable long-term outcomes in HCCs of 2-3 cm versus 3.1-4.0 cm and in high-risk versus non-high-risk locations. A randomized controlled or cohort studies with larger sample size are warranted.

[Isothermal unidirectional elongation method of gene synthesis].

Gene synthesis is gaining popularity as a versatile means of cloning genes. The prevailing synthesis methods are PCR-based and ligase-based gene synthesis. Both of them are not independent of the sequence content of the target gene. In the present study, a new gene synthesis method-isothermal unidirectional elongation method (IUEM) is proposed, which involves the cooperation of three enzymes, and is independent or partially independent of the target gene sequence by the specially designed oligonucleotides that form hairpins. The synthesis process is isothermal and serial in IUEM. Several factors that affect the elongation were studied, and two DNA segments of length 254 bp and 300 bp were synthesized with IUEM. The synthesized segments could work as building blocks for the synthesis of longer DNA sequences.