Carotegenic Virgibacillus halodenitrificans from Wadi El-Natrun Salt Lakes: Isolation, Optimization, Characterization and Biological Activities of Carotenoids

SIMPLE SUMMARY: As one of the two cases of soda lakes in the world, Wadi El-Natrun salt lakes are the intended isolation source in this study. They encompass a characteristic microbial niche of symbionts producing unique bioactive agents, among them carotenoids. Herein, halophilic carotenoid-producing microbes were isolated and purified. Based on the carotenoids content, the potent isolate, namely Virgibacillus halodenitrificans, was molecularly identified, and characterized physiologically, biochemically and morphologically as well. Its carotenoids content was optimized using statistical methods. Under optimized conditions, the maximum productivity was obtained and the extracted carotenoids were identified by UV-Vis analysis, FTIR, Raman spectroscopy, TLC and LC–MS, which confirmed such halophilic carotenoids composed of β-carotene, lutein and β-Apo-8′-carotenal mixture. Meanwhile, the carotenoids were utilized as a powerful antibacterial and antifungal agent. Notably, they exerted higher antibiofilm potentiality at a concentration of 20 μg/mL. Finally, halophilic carotenoids are competing as new biocidal agents, in particular with their biocompatibility and efficiency. ABSTRACT: Carotenoids, as phytonutrient pigments, are signified by their unique beneficial features that serve human health and the surrounding ecosystem. Haloalkaliphiles from soda lakes produce different natural bioactive molecules; however, their ability to produce carotenoids has been limited. Therefore, this study focused on the screening and isolation of carotenoid-producing haloalkaliphilic microbes. Out of 10 isolates, a powerful carotigenic bacterium was isolated, characterized phenotypically and identified on the molecular level as Virgibacillus halodenitrificans. By employing statistical approaches like Plackett–Burman design and central composite design, the influence of significant nutritional variables on carotenoids production was screened and optimized. Predictive modeling manifested that carotenoid yield was 36.42 mg/mL, a 2.12-fold enhancement compared to the basal conditions through inoculating 1.8% of bacterial biomass on optimized medium containing yeast extract (2 g/mL), peptone (10 g/mL) and NaCl (233.6 g/mL). The carotenoids content was confirmed by UV-Vis spectrum which detected a characteristic unique peak with left and right shoulders at 461 nm, 490 and 522 nm. However, FTIR and Raman spectroscopy showed the presence of several functional groups. Meanwhile, LC–MS confirmed that the examined carotenoids were composed of β-carotene, lutein and β-Apo-8′-carotenal mixture. As a bioactive agent, the carotenoids of V. halodenitrificans DASH showed characteristic antagonistic potency against a wide spectrum of Gram-positive and Gram-negative pathogens. Interestingly, a potent antifungal capacity was observed against Candida albicans, reflecting promising mycocidal efficacy against COVID-19 white fungal post-infections. Furthermore, carotenoids (20 μg/mL) inhibited the biofilm formation of P. aeruginosa and S. aureus by 54.01 ± 3.97% and 80.082 ± 0.895%, respectively. Our results proposed that haloalkaliphiles of Wadi El-Natrun lakes are promising sources of carotenoids that exhibited efficiency as safe, biocompatible and natural bioactive agents for environmental, medical and industrial applications.