Ip, C. | Search bioRxiv

bioRxiv2023

Pan-genotypic probe-based enrichment to improve efficiency of Hepatitis B virus sequencing

Hepatitis B Virus (HBV) genome sequencing can be used to provide more complete genetic information at the population and individual level to shed light on the limitations of current interventions, and inform new strategies for elimination. HBV sequencing is challenging due to the partially dsDNA genome, high diversity, low viral loads and presence of large amounts of host genetic material in clinical samples. Here we describe the design and use of a pan-genotypic panel of 74 HBV specific capture-probes and nuclease treatment in improving sequencing efficiency. We processed 20 plasma samples (viral loads 1.98 to 4.07 log10, genotypes A-E) and three positive controls (human total brain RNA and bacteriophage lambda DNA) in triplicate to compare DNAse vs. RNAse vs. no nuclease treatment. We prepared libraries using the Takara Bio SMARTer Stranded Total RNA-Seq Kit v3, split the library in two, enriching half with the custom-designed probe panel and xGen Hybridization and Wash Kit (IDT), the other half was not enriched. Both libraries were sequenced on the NovaSeq6000 platform with 2x150nt paired-end reads. Capture resulted in a 47,970 fold increase in the number of reads mapped to the HBV genome in the "no nuclease" arm (243 HBV reads per million reads sequenced in the capture pool vs. 5x10-3 reads per million in the no-capture pool). Out of 20 samples, only 1 without capture generated HBV reads (viral load 3.89 log10 IU/ml) vs. 19 samples with capture. HBV sequence yield was increased in the capture arm and resulted in 2.30 log10 (95% confidence interval 1.99 - 2.48 log10) increase in HBV reads (per million reads sequenced) per log10 increase in viral load. The proportion of HBV reads increased a median of 12 fold with RNAse treatment. We developed a targeted pan-genotypic sequencing method using a custom panel of biotinylated oligos that increases the sequencing efficacy of HBV. This method will allow us to gain a better insight into HBV diversity.

genomics↗

bioRxiv2021

Safety and efficacy of a new vaginal gel, Feminilove, for the treatment of symptoms associated with vaginal dryness and vulvovaginal atrophy in women: an in vitro and in vivo study

Vaginal dryness is a common symptom associated with vulvovaginal atrophy of menopause. The impact of vaginal dryness is very significant as it negatively affects quality of life, daily activities, sexual satisfaction as well as on interpersonal relationships. Symptoms of vaginal dryness is often underreported and undertreated. Recently, vaginal lubricants and moisturizers have been applied as one of the alternative and safe approaches to relieve vaginal dryness for women with mild to moderate vaginal dryness. We evaluated the safety and beneficial effects of a new type of estrogen-free vaginal gel, Feminilove BIO-FRESH moisturizing vaginal gel, using in vitro and in vivo experimental tools. Our results suggest that; 1) Feminilove vaginal gel exhibits minimal cell cytotoxicity on various human vaginal cells; 2) Feminilove vaginal gel exhibits minimal side-effects on the structure of vaginal mucosa stratum of experimental animals; 3) Feminiove vaginal gel inhibits the growth of pathogenic vaginal bacteria (E. coli) while promotes the growth of beneficial vaginal bacteria (Lactobacillus spp); 4) Feminilove vaginal gel elicits an anti-inflammatory response on vaginal epithelial cells; and 5) Feminilove vaginal gel promotes the production of tropoelastin and collagen on cultural vaginal smooth muscle and may restore loose vaginal wall (i.e., tightening effects). In summary, our results indicate that Feminilove BIO-FRESH moisturizing vaginal gel is a safe and effective remedy for vaginal dryness and vulvovaginal atrophy in women.

pharmacology and toxicology↗