Microorganisms in the Environment and Microbial Biotechnology

1. Microorganisms in the Environment and Microbial Biotechnology M J Larkin

2. INTRODUCTION • Microorganisms in the environment • Where are they found? • How diverse are they? • Role in geochemical nutrient cycles. • How do they grow and what are their requirements for growth and biodegradation? • Microorganisms in waste treatment: Biodegradation and environmental clean up. • Microbial production and products in industry • The genomic – metagenomic future DIRECTED READING: Prescot. Ch40 microorganisms as components of the environment Ch 44 Industrial microbiology and Biotechnology.

3. Larkin Lab Research – Biological Sciences and QUESTOR http://questor.qub.ac.uk • Basic theme is molecular biology and biochemistry of microorganisms that mediate global processes and remediation of the environment. • Currently: • Function of dioxygenases – structure and biochemistry • Bioproducts – chiral chemicals for pharmaceutical use • Diversity of biodegradative genes in environment – evolution from Archaea • Metagenomic approaches • Funded by UK government – BBSRC, EC and Industry Overview from keynote lecture at: http://www.qub.ac.uk/mlpage/researchoverview/overview.ppt Perception: http://www.qub.ac.uk/mlpage/researchoverview/space.ppt Web of Knowledge – name and address function

4. Microorganisms in the environment; Challenging conventional views of life. • Sagan and Margulis (1998) “Garden of Microbial Delights”. • “ALL of the elements crucial to global life- oxygen, nitrogen,phosphorus, sulfur, carbon- return to a usable form through the intervention of microbes… Ecology is based on the restorative decomposition of microbes and molds, acting on plants and animals after they have died to return their valuable chemical nutrients to the total living system of life on earth” • Gould (1996) “Life’s Grandeur” The Power of the Modal Bacter. • The first multicellular organisms do not enter the fossil record until about 580 million years ago - this is after about five sixths of life’s history have passed. Bacteria have been the stayers and keepers of life’s history.

5. Where are they found?Diverse environments • Virtually every environmental niche • Extremes of pH and salinity • Extremes of temperature and pressure • Without air (Anaerobic) • Growth on many chemical substrates • Attached to surfaces in biofilms • Geothermal vents and subterranean deposits

6. Where are they found?Biomass on the planet. • Most culturing analysis misses over 99% of the microbial population. • Molecular techniques now reveal hidden diversity • Heterotrophs - 5-20% biomass in sea waters - up to 80% of the primary production • Rich bacterial communities in sub-surface strata (600 m deep) - up to 2 x 1014tons - more than all flora and fauna -equivalent to 2 m layer over planet! • see:http://www.stephenjaygould.org/library/gould_bacteria.html

7. How diverse are they? Plants & Animals • Diverse range of species • Earliest life on the planet • Anaerobic then aerobic • Three Kingdoms • Eukaryote Plants & Animals • Eubacteria • Archaebacteria • Exteme living bacteria Eubacteria Archaea 3 billion years

8. How diverse are they? Diversity of bacteria in soil 16s rRNA sequences reveal true diversity in soil DNA

9. Genomics and Metagenomics Pyrosequencing “454” direct sequencing of single strands – 300 bps per read – but rapid. Use in analysis of RNA transcipts Use for rapid analysis of ALL DNA in environment – metagenomics Screening environment for useful genes. Expression requires suitable host E.Coli not always suitable Other hosts more useful – e.g Rhodococcus – used in many industrial processes Chain termination sequencing used for genomes to date – 800 bps per read

10. Microbial genome sequencing links • Sanger Institute UK • http://www.sanger.ac.uk/Projects/ • Lists bacterial pathogens sequenced and ongoing • Joint Genome Institute USA • http://www.jgi.doe.gov/ • Many environmental microorganisms and metagenomic projects Belfast connection: Pathogen Bacteroides fragilis – unprecedented gene switching mechanisms see: http://www.sanger.ac.uk/Projects/B_fragilis/ Rhodococcus – analysis of largest bacterial genome at 9.7 mB Gene rearrangements and adaptation see: http://www.rhodococcus.ca/

11. Role in geochemical nutrient cycles. • Microorganisms play a role as: • PRIMARY PRODUCERS • BIODEGRADERS AND CONSUMERS • Critical role in cycles of many elements; • Carbon and Oxygen cycle – oxygenases and oxygen fixation • Nitrogen cycle – nitrogenase - denitrification • Sulfur cycle – sulphate reduction • Phosphorus cycle

12. How do they grow: requirements for biodegradation? • Nutrients • Carbon, Nitrogen, Phosphorus, Sulfur • Many chemicals supply these • Micronutrients/ trace metals/ vitamins • Electron acceptors - usually O2 • Converts / burns carbon substrate to CO2 • Energy and biomass ie GROWTH

13. 2.0m SINGLE BACTERIUM Biodegradation GROWTH - CELL DIVISION INCREASE IN BIOMASS O2 consumption ORGANIC POLLUTANT AND NUTRIENTS (C,P,N,O,Fe,S……) CO2 evolved Controlled release of energy Slow Burning!

14. Oxygen and Electron Acceptors: crucial for Biodegradation reactions in the environment. Electron acceptor 2H+ H2O O2 SUBSTRATE ADP Pi METABOLISM ATP H2/2e- ENERGY CARBON GROWTH/Biomass CO2

15. O2 NO3- SO42- Fe3+ NO2- N2 H2O H2S Fe2+ 0.814V -0.214V -0.185V 0.741V FAST GROWTH SLOW GROWTH Role of electron acceptors; rate of biodegradation

16. Anaerobic growth and biodegradation Fermented Acetic Acid Organic matter + H2 CO2 Methanogenesis CH4 CO2 H2O

17. Fixation of oxygen as a first step in biodegradation – the key step – biodegradion – complex biochemistry Cell membrane Cell Biomass Further degradation CO2

18. Biological waste treatment; Managing microorganisms for environmental cleanup • 10 x 106Chemicals • 8 x 106Xenobiotic • 1 x 106Recalcitrant • 0.4 x 106 traded at over 50 tonnes per year • Toxicological/ biodegradative data on only around 5000-6000 • Municipal waste-water treatment • Biodegradation of industrial wastes • petrochemicals, bulk chemical processes • textiles, leathers • metals • Remediation of contaminated land in situ

19. Biological waste-water treatment: The activated sludge process.

20. EFFLUENT FREE OF POLLUTANT WASTE -WATER CONTAINING POLLUTANTS Biological waste treatment; Advanced industrial membrane reactor.

21. Cultivation of microorganisms for industrial use.

22. Cultivation of microorganisms for industrial use. Advanced laboratory fermenters in the Questor Centre

23. Products from Microorganisms: Overview of range of examples. • Various foods and drinks • Enzymes for varied uses (GM enzymes); biocatalysts • Engineered proteins ( antibodies ) • Vaccines and antibiotics (secondary metabolites) • Primary metabolites and bulk chemicals (amino acids (glutamic acid) and organic acids (acetic acid) • Pharmaceuticals and novel chiral chemicals • Recovery of metals in bioleaching • Biosensors (use of enzymes to specifically detect chemicals in medical and )

24. Microbes are everywhere ! ----- “where the bee sucks, there suck I in a cowslip’s bell I lie” ------ Ariel in “The Tempest” proclaiming his ubiquity in all manifestations of life