Resuscitation of Microorganisms after Gamma Irradiation

Abstract

Microbiological analysis of rock exposed to gamma-radiation doses between 0 and 9.34 kGy indicated that some microorganisms became viable but nonculturable (VBNC) and lost metabolic capacity as measured by BIOLOG microtiter plates. To investigate this phenomenon, portions of irradiated rock were placed at 4 degrees C for 2 months in an attempt to resuscitate the microbes to a culturable state. Culturable heterotrophs were enumerated and BIOLOG plates were used to determine the metabolic capability of the microbial community. Culturable bacteria that had previously been nonculturable were found at all doses. The number of colony types decreased from 26 in the nonirradiated control rock to between 9 and 10 in rock irradiated at doses ranging from 2.34 to 9.34 kGy. BIOLOG plates indicated partial recovery of metabolic capacity in all the samples tested. Fatty acid methyl ester analysis of the recovered isolates using the MIDI system (Microbial ID, Inc.) yielded three distinct groups of related bacteria. All resuscitated isolates clustered with the original nonirradiated isolates at the genus level, and 92% of them clustered at the species level. These results indicate that microbes were likely resuscitated from a VBNC state.

The Impact of Gamma Radiation on Sediment Microbial Processes

“…For example, under a chronic dose rate of ϳ2 Gy h Ϫ1 , microorganisms isolated from a spent nuclear fuel pond were capable of surviving total absorbed doses five times greater than tolerated in acute-dose experiments (Ͼ426 Gy h Ϫ1 ) (20,21). Furthermore, microbes from the indigenous endolithic community of a proposed repository were capable of surviving low gamma doses in a viable-but-nonculturable state (22), such that resuscitation may be possible when environmental conditions become more favorable (23). This highlights the importance of gathering low dose rate data, particularly since lower dose rates may allow species to respond via upregulating repair mechanisms (24) or even adapting over geological timescales, relevant to radwaste disposal scenarios.…”

“…For instance, studies of survival of microorganisms from clay buffer material have suggested that typically only 10% of the population survives after doses of ϳ1.6 kGy (9) and that the dose rate may not have a significant impact on the viability of microbial populations (19). On the other hand, indigenous members of an endolithic microbial community from a proposed high-level radioactive waste repository may have been able to survive in a nonculturable state after irradiation (9.34 kGy at 1.63 Gy min Ϫ1 ), to be rejuvenated when conditions become favorable (22,23).…”

Boundaries for Biofilm Formation: Humidity and Temperature

“…These conditions will favor biofilm formation by surviving, indigenous microorganisms if nutrients are present for growth. Some indigenous bacteria within Yucca Mountain have been shown to be responsible for MIC (5), and others have been shown to survive, or resuscitate, after irradiation (22,23). Therefore, the bounding limits of tem- perature and RH for biofilm formation will be important for predictive models of the stability of the high-level nuclear waste packages and support structures in the proposed repository.…”

Review of Microbial Responses to Abiotic Environmental Factors in the Context of the Proposed Yucca Mountain Repository

“…However, this work showed that microbes in YM samples can survive γ-radiation in VBNC state. In a subsequent study, Pitonzo et al (1999b) stored the irradiated rock samples for 2 months at 4 o C, in an attempt to resuscitate the VBNC microbes to a culturable state. Culturable bacteria that had previously been nonculturable were found at all radiation doses in the samples after cold storage, but in numbers that were orders of magnitude lower than the culturable organisms in the unirradiated samples.…”

“…Radiation-resistant microorganisms in the rock samples became viable but not culturable (VBNC) after a cumulative dose of 2.33 kGy and their metabolic capability was reduced considerably. In a subsequent study, Pitonzo et al (1999b) stored the irradiated rock samples for 2 months at 4 o C, in an attempt to resuscitate the VBNC microbes to a culturable state. Results showed that microbes can survive considerable radiation doses in a VBNC state, and recovered some of their previous culturable and metabolic capabilities, once the radiation stress was removed.…”