High growth rate and substrate exhaustion results in rapid cell death and lysis in the thermophilic bacterium Geobacillus thermoleovorans

Abstract

Batch cultures of the thermophilic bacterium Geobacillus thermoleovorans T80 attained extremely high-specific glucose utilization rates leading to high specific growth rates, followed by extensive cell death and lysis with the onset of substrate exhaustion. The dramatic decrease in live cell numbers, as determined by flow cytometry, was accompanied by the release of soluble protein. Once the growth phase reached the point of commitment to lysis created by the impending exhaustion of substrate, the addition of extra carbon substrate did not halt the rapid death rate and lysis, although, towards the end of the exponential growth phase, the substrate was utilized producing only a small additional biomass concentration as a result of the net effect of cell growth and death. This lytic phenomenon was observed when a range of different carbon substrates (glucose, pyruvate, acetate, n-hexadecane, nutrient broth), as well as ammonium (the nitrogen source) in the presence of excess carbon source, reached near exhaustion. The rate and extent of cell death and the ensuing lysis depend on the culture growth rate. Cultures batch grown with a lower initial substrate concentration, or at a lower temperature, or at lower dilution rates for continuous-flow cultures, exhibited a lower rate and extent of cell death and lysis. Batch re-culture of the persister cells resulted in a behavior identical to that of the original culture indicating that these cells were not genetically modified. The glucose utilization, cell growth and death rates were mathematically described based on Monod kinetics and estimated values of pertinent biokinetic constants are reported.

Primers for overlookednirK, qnorB,andnosZgenes of thermophilic Gram-positive denitrifiers

“…Geobacillus , are soil inhabitants that have been reported to be the most abundant and/or dominant active group of bacteria in certain ecosystems (Felske et al ., ; Rahman et al ., ; Janssen, ), studies on functional – physiological and ecological – roles, in particular on nitrogen transformations, of these and other thermophilic organisms in both hot as cold environments are virtually nonexistent. Apart from a number of earlier reports on phenotypic denitrification and denitrification enzymes in Geobacillus (Garcia, ; Ho et al ., ; Manachini et al ., ; Nazina et al ., ), in‐depth research on thermophilic denitrifying bacteria has only been started recently (Pavlostathis et al ., ; Mishima et al ., ; Nara et al ., ; Fukuda et al ., ; Matsumoto et al ., ; Salomonsson et al ., ; Jung et al ., ).…”

Application ofrecAandrpoBsequence analysis on phylogeny and molecular identification ofGeobacillusspecies

“…These metabolic capabilities of geobacilli suggest that they may have important biotechnological applications, both in the industrial and environmental fields (Hasan et al. 2006; Pavlostathis et al. 2006).…”

“…Recent studies by Feitkenhauer et al (2001Feitkenhauer et al ( , 2003 had revealed the ability of Geobacillus thermoleovorans on phenol degradation. These metabolic capabilities of geobacilli suggest that they may have important biotechnological applications, both in the industrial and environmental fields (Hasan et al 2006;Pavlostathis et al 2006).…”

The complete denitrification pathway of Geobacillus thermoleovorans

“…It is an alternative process for respiration used by bacteria under low oxygen conditions, through which they can reduce nitrate and/or nitrite to dinitrogen gas through the production of the intermediates nitric oxide and nitrous oxide [1]. In recent studies, the thermophilic bacterium Geobacillus thermoleovorans was shown to reduce nitrate to nitrogen gas, indicating the presence of a denitrification pathway [2]. The first catalytic step of this pathway can be achieved by two alternative enzymes: a membrane-bound or periplasmic-bound reductase (NAR and NAP, respectively).…”