Microbes and Metabolism - Environmental Biotechnology: Theory and Application - page 47

Woese, C.R. and Fox, G.E. (1977) Phylogenetic structure of the prokaryotic
domain: the primary kingdoms. Proceedings of the National Academy of Sci-
ences of the United States America , 74 , 5088-5090.
Woese, C.R., Kandler, O. and Wheelis, M.L. (1990) Towards a natural system
of organisms: proposal for the domains Archaea, Bacteria, and Eucarya.
Proceedings of the National Academy of Sciences of the United States America ,
87 , 4576-4579.
CaseStudy2.1 MicrobialEco-ToxicologyTesting(Scotland/Sweden)
Assessment of eco-toxicant effects has often been made conventionally by extrap-
olating from straightforward chemical analysis of the soil, water or other medium
being investigated, but while this is useful information perse it gives little clue as to
their impact on biological systems.
Although there is an obvious potential in extending microbial assays to the
problem, many of the available techniques have made use of individual species,
typically depending on a single genotypic character, such as the Lux operon in
the bio-luminescent bacterium, Vibrio fischeri. As a consequence, though the tests
developed are swift and well established, they are undoubtedly open to criticism
for their narrow basis. However, with support from Scottish Enterprise and the UK's
EUREKA scheme, together with Sweden's Karolinska Institute, the Aberdeen-based
biotech company, NCIMB, has recently succeeded in expanding the concept to
embrace a diverse, multi-species complement of microbes.
The Microbial Assay for toxic Risk Assessment (MARA) kit uses 11 genetically
diverse microbe species (1 yeast and 10 bacteria from the alpha, beta and gamma
proteobacteria) lyophilised in situ in a 96-well micro-titre plate. In use, the micro-
organisms are first reconstituted at 30 ◦ C for 4 hours before being inoculated with
a range of sample dilutions and incubated, again at 30 ◦ C, for 18 hours. At the end
of this stage the plate is scanned and dedicated image analysing software used to
quantify microbial growth or inhibition by measuring the level of reduction of a
redox dye incorporated in the plate's substrate medium.
This enables a microbial toxic concentration (MTC) value to be calculated
for each individual microbial species as well as a mean MTC for the entire 11
micro-organisms, yielding a 'toxic fingerprint' for the sample, which could prove a
particularly useful indicator in routine monitoring applications. The system also has
considerable investigative potential to help diagnose unidentified contaminants in
the environment, by enabling a comparative dendogram to be produced to allow
the unknown chemicals to be evaluated against known standards.