Biological activity of infusion and decoction extracts of Hibiscus sabdariffa L. calyces

To compare the phytochemical and biological profiles, infusion and decoction extracts of dried Hibiscus sabdariffa calyces were prepared. Infusion extract had the highest phenolic content (292.42 ± 0.62 mg GAE/g extract) and antioxidant activity (TEAC = 3.09 ± 0.28). It also potently inhibited the activities of α-amylase (58.57 ± 1.4%), α-glucosidase (66.92 ± 2.5%) and 5-lipoxygenase (58.97 ± 1.17 μg/ml). All extracts showed weak cytotoxicity as tested on Vere monkey kidney cells.

Characterization of a Novel Simian Sapelovirus Isolated from a Cynomolgus Monkey using PLC/PRF/5 Cells

We isolated a novel simian sapelovirus (SSV), Cam13, from fecal specimen of a cynomolgus monkey by using PLC/PRF/5 cells. The SSV infection of the cells induced an extensive cytopathic effect. Two types of virus particles with identical diameter (~32 nm) but different densities (1.348 g/cm3 and 1.295 g/cm3) were observed in the cell culture supernatants. The RNA genome of Cam13 possesses 8,155 nucleotides and a poly(A) tail, and it has a typical sapelovirus genome organization consisting of a 5’ terminal untranslated region, a large open reading frame (ORF), and a 3’ terminal untranslated region. The ORF encodes a single polyprotein that is subsequently processed into a leader protein (L), four structural proteins (VP1, VP2, VP3, and VP4) and seven functional proteins (2A, 2B, 2C, 3A, 3B, 3C, and 3D). We confirmed that 293 T, HepG2/C3A, Hep2C, Huh7 and primary cynomolgus monkey kidney cells were susceptible to SSV infection. In contrast, PK-15, Vero, Vero E6, RD-A, A549, and primary green monkey kidney cells were not susceptible to SSV infection. We established an ELISA for the detection of IgG antibodies against SSV by using the virus particles as the antigen. A total of 327 serum samples from cynomolgus monkeys and 61 serum samples from Japanese monkeys were examined, and the positive rates were 88.4% and 18%, respectively. These results demonstrated that SSV infection occurred frequently in the monkeys. Since Cam13 shared 76.54%–79.52% nucleotide sequence identities with other known SSVs, and constellated in a separate lineage in the phylogeny based on the entire genome sequence, we propose that Cam13 is a new genotype of the simian sapelovirus species.

Cytotoxic and Genotoxic Effects of Fluconazole on African Green Monkey Kidney (Vero) Cell Line

Fluconazole is a broad-spectrum triazole antifungal that is well-established as the first-line treatment for Candida albicans infections. Despite its extensive use, reports on its genotoxic/mutagenic effects are controversial; therefore, further studies are needed to better clarify such effects. African green monkey kidney (Vero) cells were exposed in vitro to different concentrations of fluconazole and were then evaluated for different parameters, such as cytotoxicity (MTT/cell death by fluorescent dyes), genotoxicity/mutagenicity (comet assay/micronucleus test), and induction of oxidative stress (DCFH-DA assay). Fluconazole was used at concentrations of 81.6, 163.2, 326.5, 653, 1306, and 2612.1μM for the MTT assay and 81.6, 326.5, and 1306μM for the remaining assays. MTT results showed that cell viability reduced upon exposure to fluconazole concentration of 1306μM (85.93%), being statistically significant (P<0.05) at fluconazole concentration of 2612.1μM (35.25%), as compared with the control (100%). Fluconazole also induced necrosis (P<0.05) in Vero cell line when cells were exposed to all concentrations (81.6, 326.5, and 1306μM) for both tested harvest times (24 and 48 h) as compared with the negative control. Regarding genotoxicity/mutagenicity, results showed fluconazole to increase significantly (P<0.05) DNA damage index, as assessed by comet assay, at 1306μM versus the negative control (DI=1.17 vs DI=0.28, respectively). Micronucleus frequency also increased until reaching statistical significance (P<0.05) at 1306μM fluconazole (with 42MN/1000 binucleated cells) as compared to the negative control (13MN/1000 binucleated cells). Finally, significant formation of reactive oxygen species (P<0.05) was observed at 1306μM fluconazole vs the negative control (OD=40.9 vs OD=32.3, respectively). Our experiments showed that fluconazole is cytotoxic and genotoxic in the assessed conditions. It is likely that such effects may be due to the oxidative properties of fluconazole and/or the presence of FMO (flavin-containing monooxygenase) in Vero cells.