Nitrogen Fixation in Obligate Methanotrophs

Abstract

A number of representative species of obligate methane-oxidizing bacteria were surveyed for their ability to fix N2 by growth experiments and the acetylene reduction test. Although all strains exhibited growth on nitrogen-free plates, only type I1 organisms and the type X methanotroph Methylococcus capsulatus (Bath) grew well in nitrogen-free liquid medium and were capable of active acetylene reduction. N,-fixation in type I1 methanotrophs was less sensitive to 0, than in the type X methanotroph Methylococcus capsulatus (Bath) and batch cultures of type I1 organisms could be established at p 0 2 values of up to 0.2 bar. N,-fixation in Methylococcus capsulatus (Bath) was inhibited at p 0 2 values above 0.1 5 bar and the 'switch-off' of nitrogenase activity by ammonia was also observed in this organism.

NifH and NifD phylogenies: an evolutionary basis for understanding nitrogen fixation capabilities of methanotrophic bacteria

“…Except for Methylococcus spp., gammaproteobacterial MB showed no growth in nitrogen-free medium, even if they were cultivated under microaerobic conditions. The findings of Murrell & Dalton (1983) correspond well with the results of our NifH-and NifD-based trees, in which all alphaproteobacterial MB plus Methylococcus capsulatus form a common cluster, clearly distinct from the gammaproteobacterial MB Methylomonas and Methylobacter. Auman et al (2001) demonstrated acetylene reduction activity in representatives of Methylomonas and Methylobacter.…”

“…Thus, our investigation provides the first evidence that, besides Methylococcus, at least some representatives of type I MB are capable of growth in nitrogen-free media, though under a very limited range of conditions. It should be noted that in the experimental study of Murrell & Dalton (1983), a number of batch cultures of type I MB were also incubated under low pO 2 (0?01-0?02 bar) to achieve growth on nitrogen-free media. However, as pointed out above, the OD increase under these conditions is usually very small because the culture becomes limited by O 2 , and the OD soon declines back to zero.…”

“…Murrell & Dalton (1983) used a combined approach, examining growth under a wide range of pO 2 values and using the acetylene reduction test to assess the dinitrogen-fixation capability of MB. This study revealed that only alphaproteobacterial MB and Methylococcus capsulatus grow well in nitrogen-free liquid medium and actively reduce acetylene.…”

“…Under high partial pressures of oxygen, representatives of Methylocella usually show quite poor growth in nitrogen-free media. This is also typical for other dinitrogen-fixing MB such as Methylococcus, Methylosinus and Methylocystis (Murrell & Dalton, 1983;Dedysh et al, 2000;Auman et al, 2001). Decreasing dissolved oxygen tension can enhance both growth and acetylene reduction activity of these MB.…”

“…Using this approach, we were able to obtain batch cultures of Methylobacter luteus and Methylocystis echinoides IMET 10491 of a relatively high OD 600 , up to 0?35-0?45, on NFMS medium. In addition, we applied the same strategy to establish a batch culture of another type II MB, Methylosinus trichosporium OB3b, on NFMS medium, since this bacterium had already been used as a model organism to study the nitrogen-fixation capability of MB (Murrell & Dalton, 1983;Auman et al, 2001). The cells from these three cultures were used to inoculate a number of flasks with a range of pO 2 values from 0?005 to 0?2 bar.…”