Bactérias aquáticas cultiváveis sob efeito da radiação ultravioleta: avaliação da curva de crescimento, viabilidade celular e ultraestrutura

Abstract

Aquatic bacteria is a key component of microbial loop in the aquatic food web. These organisms have important roles in the cycling of organic matter and energy flow in aquatic ecosystems. Experimental models using aquatic bacteria are essential for understanding how isolated factors affect bacterial communities dynamics. Ultraviolet radiation is a non-ionizing radiation which corresponds to 9% of solar radiation that falls on the aquatic ecosystems. It shifts bacterial communities composition and induces cellular changes by damaged caused in DNA and proteins structures. Because the incidence of ultraviolet radiation on aquatic ecosystems has been increasing, studies focused on the effects of this radiation on bacteria at both cellular and community level are very important. The present work aimed to establish a culture model of freshwater aquatic bacteria and investigate the action of ultraviolet radiation on the cultured bacterial community. For bacterial cultures, samples were collected from Funil Reservoir (RJ), serially diluted and spread-plate in two non-selective solid culture media: R2A and TSA. Colonies were characterized through morphological aspects and transferred to non-selective liquid media (TSB). The grown curve was established by optical density through spectrophotometry and by cell density using a fluorescent marker (DAPI) through fluorescence microscopy. Liquid cultures were exposed to ultraviolet radiation (UVA+UVB) during three hours. Hourly samples were collected and the following parameters were analyzed: (i) growth curves by optical density and cell density; (ii) cellular viability using the Baclight kit; and (iii) ultraestructural changes by transmission electron microscopy (TEM). The use of different culture media (R2A and TSA) enabled the establishment of a freshwater aquatic bacteria model. A higher number of morphotype colonies was observed when R2A medium was used and, therefore, this culture medium was considered more suitable for culturing aquatic bacteria.Bacteria exposed to ultraviolet radiation showed a decreased bacterial density, as measured by optical and bacterial density. Ultraviolet radiation induced a significant increase of the aquatic bacteria mortality in culture, as demonstrated by the cell viability evaluations. On the third hour of exposure to ultraviolet radiation, we detected a higher number of non-viable cells in comparison with those found after the first hour of radiation exposure and controls. Moreover, our data also demonstrated that the ultraviolet exposure time negatively interferes with the aquatic bacterial growth and positively with aquatic bacterial mortality. TEM revealed great ultraestructural diversity of aquatic bacteria in the culture model and demonstrated that ultraviolet radiation induces ultraestructural changes and death in aquatic bacteria. In the ultraviolet radiation-treated-group, it was found a higher frequency of gram-positive bacteria with S-layer and/or capsule than in control group. Most gram-positive bacteria in UV group presented intact ultraestructure. In contrast, gram-negative bacteria were more affected by the ultraviolet radiation. These findings indicate that the S-layer have a protective role against ultraviolet radiation and that repair mechanisms were likely triggered by the bacterial community in response to this radiation.. Altogether, the results of the present work demonstrate that the ultraviolet radiation affects fresh water bacterial communities in culture by inducing ultrastructural changes and cell death. Moreover, it was established a culture model of fresh water bacteria that may contribute to the understanding of different aspects of the bacterial community and other studies focused on environmental stress