LS-6-P-3244 Ultrastructural changes and death in cultured aquatic bacteria induced by ultraviolet radiation
Aquatic bacteria play crucial roles in the cycling of organic matter and energy flow in aquatic ecosystem [1]. Ultraviolet radiation (UV) is a non-ionizing radiation, which corresponds to 9% of solar radiation that falls on the aquatic ecosystems [2]. It shifts bacterial communities’ composition and may induce cellular changes. Because of the increasing incidence of UV on aquatic ecosystems, studies focused on the effects of this radiation on bacteria at both cellular and community levels are very important [3]. Here,we investigated the occurrence of death processes and ultrastructure of cultured aquatic bacteria exposed to UV. For bacterial cultures, samples were collected from Funil Reservoir (Brazil, RJ), serially diluted and seed in two non-selective solid media (R2A and TSA). Bacterial colonies were transferred to non-selective liquid media (TSB) and in stationary phase were exposed to ultraviolet radiation (UVR) during 3 h. For this experiment was used UVR in wavelength range corresponding to UV-A+ UV-B (emission peaks: 365/312nm). Hourly samples were collected and the following parameters were analyzed: (i) growth curves by optical density through spectrophotometry and cell density through fluorescence microscopy using DAPI; (ii) cellular viability using fluorescent markers for membrane integrity (Live/dead Baclight kit); and (iii) ultrastructural changes by transmission electron microscopy (TEM) [4]. Our density analyses show that bacteria exposed to UVR showed a decreased bacterial density (p<0.05). Cell viability evaluation showed that UVR induced a significant increase of the aquatic bacteria mortality in culture (p<0.05). On the third hour of exposure to UVR, we detected a higher number of non-viable cells in comparison with those found after the first hour of radiation exposure and controls (p=0.037). 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 that bacteria exposed to UVR had higher proportion of cellular damages (51.38%) compared to control cells (9.54%). Damaged bacteria showed loss of cellular envelope integrity, shape changes and cellular elongation. Moreover, 11.9% of bacteria in death process presented emptying of cell components. Altogether, the results of the present work demonstrate that the UV affects fresh water bacterial communities in culture by inducing ultrastructural changes and cell death.
References
[1] L.R. Pomeroy et al, Oceanography, 20 (2007) 28-33
[2] J.M. Anderson et al, Edward Arnold, 108 (1981) 13-35
[3] G.J. Herndl et al, Nature, 361(1993) 717-719
[4] T.P. Silva et al, Antonie van Leeuwenhoek, 105 (2014) 1-14
This work was supported by CAPES, CNPq and Fapemig.
Fig. 1: Bacterial death curve obtained by cell and optical density analyses during ultraviolet exposure. In (A and B), compare cell death curve in UVR-treated group and control. Bacterial cultures were submitted to ultraviolet radiation. A sharp decline of cell density (A) and optical density (B) is detected from the first hour in UVR-exposed bacteria. |
Fig. 2: Visualization of live/dead aquatic bacteria in culture by fluorescence microscopy. In (A), Live bacteria are stained in green while dead cells are seen in red. (B) Bacterial viability in UVR-treated group and control. After 3 h, of UVR exposition, the frequency of dead bacteria is greater compared with 2 and 1 h (*p<0.05). |
Fig. 3: Ultrastructure of aquatic bacteria. (A) A intact bacteria show nucleoid areas (N) and typical cellular, as highlighted in (Ai). (B-D) Damaged bacteria are observed after exposition to UVR. Observe bacterial changes, such as cell elongation and condensation (B), disintegration of the cell envelope (C, arrow) and emptying of cell components (D). |
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