Extremely thermophilic bacteria in New Zealand hot springs

Abstract

Pools in thermal areas were investigated by direct microscopic procedures, by indirect tracer methods and by conventional bacteriological methods of enrichment and isolation, in order to determine the diversity and distribution of extremely thermophilic bacteria. The indirect isotope uptake tracer method was unsuccessful during the earlier part of the project. Although, problems with isotope methodology were largely solved, this did not occur early enough in the investigation to allow use of the method as a means of establishing a biomass activity index for pools. The immunofluorescence method worked well on laboratory grown isolates but was surprisingly unsatisfactory when used in situ. The method was abandoned because of the time required to correct possible problems which may have been due to the choice of the microorganisms used in the study. The implications of these results with regard to ecological studies are discussed in the thesis. Electron microscopy was used to further investigate the morphology and diversity of bacteria in pools. Direct microscopic examination of pool water by phase contrast microscopy showed that over 90% of the pools contained bacteria. Gram staining of slides colonized in situ revealed that Gram negative bacteria were the only types present in most pools. New methods of grid preparation and handling were developed. In situ colonisation of grids showed that practically all the 38 pools over the temperature range of 70°C to 102°C and pH’s from 3.0 to 9.5 contained bacteria. Pools varied as to diversity of species and numbers of cells of the same morphotype in each pool. Some pools contained an apparent monoculture whereas others had a diverse ecosystem. In situ EM technique and subsequent cultural studies revealed that sulphur-respiring archaebacteria were the dominant microorganisms present in springs with temperatures above 90°C with both coccal and rod-shaped forms present. Methanogens, another archaebacterial group, were also isolated but they inhabited pools with lower temperatures (between 65 to 80°C). Isolation and cultivation procedures were also successfully used to obtain cultures of more conventional anaerobes. These were chiefly obtained from neutral springs in the 70 to 85°C range and fermented carbohydrates to a variety of end products but primarily acetate, ethanol or lactate. The diversity of this group of caldoactive glycolytic anaerobes was demonstrated by isolating species which produced quite distinct ratios of the three major end products. Some of these isolates were very similar to glycolytic anaerobes described previously e.g. Thermoanaerobium brockii, Thermoanaerobacter ethanolicus. A number of isolates had properties different from the type strains. Investigation of the aerobic flora of thermal springs were not as intensive as those on anaerobes, largely because these organisms have been well described elsewhere. Thermus and Bacillus species were the most dominant aerobic microflora in many of the New Zealand springs. Their distribution and characteristics conformed to expectations of overseas results. Thermothrix and Sulfolobus species were isolated from a few pools but Thermomicrobium species which have been isolated from the Yellowstone National Park thermal springs, USA, have not been detected in the New Zealand thermal springs.