BackgroundThe mosquito transmitted Dengue virus (DENV) remains a significant public health problem in many tropical and subtropical countries. Increasing evidence has suggested that during the infection process cellular lipids play important roles at several stages of the replication cycle. This study sought to characterize the changes in lipid metabolism gene expression and investigated the role of one enzyme, fatty acid synthase, in DENV infection.MethodsTranscriptional profiles of genes associated with lipid metabolism were evaluated by real-time PCR after infection of different cell lines (HepG2 and HEK293T/17) and with different DENVs (laboratory adapted and low passage). Expression profiles of genes were evaluated by western blotting. A critical lipid metabolism protein, fatty acid synthase was down-regulated through siRNA and inhibited with orlistat and the effect on DENV infection determined by flow cytometry, plaque assay, western blotting and confocal microscopy.ResultsThe results showed alterations of gene transcription and expression were seen in genes variously associated with lipogenesis, lipolysis and fatty acid β-oxidation during DENV infection. Interference of fatty acid synthase with either siRNA or orlistat had marked effects on virus production, with orlistat having an EC50 value of 10.07 μM at 24 h post infection. However, non-structural protein expression was largely unaffected.ConclusionsWhile drug treatment reduced virus titer by up to 3Log10, no significant effect on DENV non-structural protein expression was observed, suggesting that fatty acid synthase acts through an effect on virion formation.Electronic supplementary materialThe online version of this article (doi:10.1186/s12985-017-0685-9) contains supplementary material, which is available to authorized users.

Lactococcus garvieae BCC 43578 produces a novel class II bacteriocin, garvieacin Q (GarQ), 70 amino acids in length and containing a 20-amino-acid N-terminal leader peptide. It is cleaved at the Gly-Gly site to generate the mature GarQ (5,339 Da), which is especially inhibitory against Listeria monocytogenes ATCC 19115 and other L. garvieae strains. Ribosomally synthesized antimicrobial polypeptides of bacterial origin, designated peptide bacteriocins, play an important role in bacterial competition, allowing bacteriocinproducing bacteria considerable survival advantages. Most bacteriocins from lactic acid bacteria (LAB) are small, heatstable, cationic, amphiphilic, membrane-permeabilizing polypeptides (11,12,20). LAB bacteriocins are categorized into two main classes (5), class I (lanthionine-containing lantibiotics) and class II (non-lanthionine-containing bacteriocins), which has 4 subclasses, namely, class IIa (pediocin-like like), IIb (two peptide), IIc (circular), and IId (nonpediocin linear one peptide).Lactococcus is one of the most important LAB genera, which is widely found in fermentation food and in the environment (3,4). A large number of strains of Lactococcus spp. have been found to produce bacteriocins. Lactococcal bacteriocins are represented in all the classes and subclasses (28). To date, only two bacteriocins from Lactococcus garvieae have been reported: garviecin L1-5 (2.5 kDa) from L. garvieae L1-5, isolated from bovine milk (26), and garvicin ML (6,022 Da) from L. garvieae DCC43, isolated from the intestinal content of mallard duck (Anas platyrhynchos) (2, 23).Here, we describe the isolation and purification of a novel class II bacteriocin, garvieacin Q (GarQ), obtained from culture supernatant of L. garvieae BCC 43578. A genomic DNA fragment containing garQ was cloned and sequenced, allowing the identification of garQ's open reading frame, as well as some of the adjacent DNA sequences.Purification and molecular characterization of GarQ from L. garvieae BCC 43578. Of 30 strains of L. garvieae isolated from nham (local fermented pork sausage), only L. garvieae BCC 43578 exhibited antilisterial activity using the spot-on-lawn method as described previously (18), with Listeria monocytogenes ATCC 19115 as the indicator strain. Thus, its bacteriocin (GarQ) was purified and characterized. Culture supernatant obtained from the 18-h culture grown at 30°C in MRS broth produced 1.6 ϫ 10 6 activity units (AU)/liter with a specific activity of 57 AU/mg protein. Purification of GarQ was performed using a sequential series of chromatographies (Amberlite XAD-16, SP-Sepharose, and reverse-phase high-performance liquid chromatography [HPLC]), resulting in a final yield and fold purification of GarQ of 0.13% and 12,754-fold (731,429 AU/mg protein), respectively (Table 1).

Canine oral tumors are relatively common neoplasms in dogs. For disease monitoring and early diagnosis, salivary biomarkers are appropriate because saliva collection is non-invasive and requires no professional skills. In the era of omics, matrix-assisted laser desorption/ionization with time-of-flight mass spectrometry (MALDI-TOF MS) coupled with liquid chromatography-tandem MS (LC-MS/MS) are suitable to identify potential disease-associated peptides and proteins. The present study aimed to use MALDI-TOF MS and LC-MS/MS to search for particular peptide mass fingerprints (PMFs) and conceivable biomarkers in saliva of dogs with early- and late-stage oral melanoma (EOM and LOM, respectively), oral squamous cell carcinoma (OSCC), benign oral tumors (BN), and periodontitis and healthy controls (CP). Pooled saliva samples in each group were used to be representative of population change. Unique PMFs were obtained and specific peptide fragments were sequenced by LC-MS/MS and BLAST-searched with mammalian protein databases. Seven peptide fragments appeared in the tumor groups (EOM, LOM, OSCC and BN) at 1096, 1208, 1322, 1794, 1864, 2354 and 2483 Da, two peptide fragments appeared in the LOM and OSCC groups at 2450 and 3492 Da, and in the CP controls at 2544 and 3026 Da. Also, protein–chemotherapy drug interaction networks were exhibited. Using western blot analysis, the expression of sentrin-specific protease 7 (SENP7), a peptide fragment at 1096 Da, in OSCC was significantly increased, as was the expression of TLR4, a peptide fragment at 3492 Da, in LOM and OSCC, compared with the CP group. The expression of nuclear factor kappa B (NF-κB), a TLR4 partner, was notably increased in OSCC compared with CP, BN and EOM. The expression was also enhanced in LOM compared with EOM. Expressed protein sequences from western blots were verified by LC-MS/MS. Western blots were then performed with individual samples in each group. The results showed the elevated expression of TLR4 in LOM and OSCC, compared with that in CP and BN, the increased expression of NF-κB in LOM and OSCC, compared with CP and in LOM compared with BN, and the enhanced expression of SENP7 in LOM and OSCC, compared with that in CP and BN. In conclusion, discrete clusters of EOM, LOM, OSCC, BN and CP groups and potential protein candidates associated with the diseases were demonstrated by salivary proteomics. Western blot analysis verified SENP7, TLR4 and NF-κB as potential salivary biomarkers of canine oral tumors.