Genomic and functional insights into the interactions between vaginal Lactobacillus strains and pathogens

The vaginal microbiota of healthy pre-menopausal women is typically dominated by one Lactobacillus species among L. crispatus, L. gasseri, L. jensenii and L. iners. Thanks to a series of antimicrobial activities, strains belonging to these species represent the first barrier against infections and maintain niche homeostasis. On the other hands, the increase abundance in pathogen species is associated with the onset of numerous diseases, leading also to an increase risk of other infections acquisition. The deciphering of factors which influence Lactobacillus survival, as well as the interactions between lactobacilli-pathogens and pathogens-pathogens represent an important topic of study for improving woman health and investigating effective probiotic strategies. Here, we investigated environmental factors and genetic traits that lead to the dominance of either L. crispatus or L. gasseri in the vaginal niche and the possible applications of liposomes loaded with L. gasseri biosurfactants for the treatment and prevention of Staphylococcus aureus biofilm infections. Furthermore, considering the increasing relevance acquired by bacterial extracellular vesicles (EVs) we analysed the role of EVs derived from vaginal lactobacilli and pathogens on both bacterial growth and HIV-1 infections. As a result, we reported for the first time i) common and species-specific genotypic and phenotypic features of L. crispatus and L. gasseri ii) significant antibiofilm activity of liposomes loading vaginal Lactobacillus biosurfactants against multi-drug resistant S. aureus strains iii) absence of growth regulation mediated by EVs derived from lactobacilli on pathogen cultures and vice versa iv) anti-HIV-1 activity of protein derived from L. gasseri EVs and unexpected antiviral effect of pathogen-derived EVs on HIV-1 infections in vitro. In conclusion, this PhD thesis explored characteristics and possible applications of vaginal lactobacilli for the human health, as well as promising antiviral effects of both lactobacilli and pathogen derived EVs.

Exploratory analysis of the role of circulating microRNA in metastatic melanoma patients

MicroRNAs act as oncogene or tumor suppressor gene regulators and are actively released from tumor cells in the circulation. Specific microRNAs can be isolated and quantified in the blood, usually in serum or plasma fractions, where they are uncommonly stable. Cell-free microRNAs serve many, and possibly yet unexplored, functional roles and microRNA levels reflect underlying conditions and have been associated with skin cancer presence, stage and evolution. However, the clinical potential of circulating miRNAs in metastatic melanoma remains largely undefined. From May 2020 to September 2022, we conducted a spontaneous, monocentric, exploratory study on human tissues in vitro, which aimed to evaluate the prognostic and predictive role of circulating miRNAs in metastatic melanoma patients. At the Medical Oncology Unit of Policlinico Sant’Orsola-Malpighi of Bologna, peripheral venous blood samples from patients with metastatic melanoma treated with checkpoint inhibitors (CPI) were collected before the start of CPI (baseline, T0) and longitudinally, approximately every 3 months (T1, T2, etc). Circulating miRNA quantification was performed by droplet digital PCR (Biorad) using an EvaGreen and LNA primer-based assays. QuantaSoft Program (Biorad) calculated the absolute quantifications of each miRNA, indicated as copies/µL. After analysis of the literature, we chose to analyze miR-155-5p, miR-320a and miR-424-5p level. All miRNAs except miR-424-5p show a significantly higher level in plasma of patients who are alive after 1 year of follow-up. High/low levels of baseline miR-155-5p, miR-320a and miR-424-5p are significantly associated with overall survival and progression-free survival. Furthermore, a preliminary analysis on the group of patients who received first-line with anti-PD-1 (N=7), baseline miR-155-5p shows higher levels in responder vs. non responder patients (p 0.06). These data, though promising, are preliminary and need to be further investigated in a larger cohort of patients.