Microbes and biotechnology

1. Microbes and biotechnology Option F.1

2. Assessment statements F.1.1 Outline the classification of living organisms into three domains. F.1.2 Explain the reasons for the reclassification of living organisms into three domains. F.1.3 Distinguish between the characteristics of the three domains. F.1.4 Outline the wide diversity of habitat in the Archae, as exemplified by methanogens, thermophiles and halophiles. F.1.5 Outline the diversity of Eubacteria, including shape and cell wall structure. F.1.6 State, with one example, that some bacteria form aggregates that show characteristics not seen in individual bacteria. F.1.7 Compare the structure of the cell walls of Gram-positive and Gram-negative Eubacteria. F.1.8 Outline the diversity of structure in viruses including: naked capsid versus enveloped capsid; DNA versus RNA; and single stranded versus double stranded DNA or RNA. F.1.9 Outline the diversity of microscopic eukaryotes, as illustrated by Saccharomyces, Amoeba, Plasmodium, Paramecium, Euglena and Chlorella.

3. The five kingdoms • Established in 1959 by Robert Whittaker • Bacteria: single-celled organisms with no organized nucleus and no membrane-bound organelles • Protista: single-celled organisms with an organized nucleus and organelles each surround by a membrane • Fungi: multicellular organisms which obtain their food using extracellular digestion and have cell walls of chitin • Plants: multicellular organisms which obtain their food by photosynthesis and have cell walls of cellulose • Animals: multicellular organisms which obtain their food by feeding on other organisms and have no cell wall

4. Five kingdoms grouped into two categories • Prokaryotes: bacteria which have no organized nucleus and no membrane-bound organelles • Eukaryotes: all of the other kingdoms which have an organized nucleus

5. The three domains • Attempt by Carl Woese to improve the accuracy of the classification system based on studies of rRNA Eubacteria: ‘true’ bacteria, prokaryotes with no organized nucleus and no membrane-bound organelles. • Archaea: archaebacteria or ‘ancient’ bacteria are also prokaryotes; most live in extreme environments • Eukarya: single-celled and multicellular organisms which all have their DNA contained in a nucleus; plants, animals, protists, and fungi

6. rRNA is a molecule common to all organisms Performs same function in all organisms Coded for by DNA By looking at variation in the sequence of rRNA, evolutionary relationships became apparent Eubacteria and Archaea have different molecules making up their cell walls E and A have different molecular structure of their cell membranes E and A have different sequences of nucleotides in their rRNA Reasons for reclassification into three domains

7. Characteristics of the three domains

8. Homework • What is the RNA world hypothesis? • Give supporting and refuting details for the hypothesis. • NO OPINIONS!!!

9. Diversity of habitat of Archaebacteria • Methanogens – use carbon dioxide to make methane; strict anaerobes; live in guts of termites and cattle, Siberian tundra, swamps rice fields, and in large intestines of dogs, pigs, and humans • Thermophiles – live in sulfur hot springs where the pH is very acidic and temperatures are up to 90°C; some live in hydrothermal vents up to 105°C • Halophiles – live in the Dead Sea, the Great Salt lake, and evaporated salt water ponds

10. Three main shapes Spheres (cocci) Rods (bacilli) Helices (spirilla) Varieties of shape Chain (strepto) Cluster (staphylo) Pair (diplo) Aggregation Vibrio fischeri emit light when in large groups, but not alone Bioluminescence caused by turning on of gene by increased amount of signal molecule Exist in light organs of squid Bacteria senses high density based upon amt. of signal present called quorum sensing Variety of cell wall Gram + Stain purple Gram – Stain pink Diversity of Eubacteria

11. Comparison of gram + and gram - bacteria

13. Diversity of structure of viruses • Are viruses alive? • Not cellular • Cannot reproduce without a host • Basic structure • Nucleic acid • DNA (double- or single-stranded) • RNA (double- or single-stranded) • Several enzymes • Protein coat (capsid) • Naked • Enveloped by membrane

14. Homework:Fill in chart

15. Saccharomyces (yeast) • Ferments carbohydrates in flour or malt and gain energy from this digestion • Carbon dioxide gas (in bread) and ethanol (in beer) are by-produces • Fungi • Secrete enzymes outside their cells and absorb the produces of digestion back into the cell • Have chitin in cell wall

16. Amoeba • Fluid state of cytoplasm enables it to change its shape easily • Pseudopodia wrap around a prey in order to trap it in a food vacuole for intracellular digestion

17. Plasmodium • Parasitic heterotroph • Mosquitoes carry and infect humans causing malaria

18. Paramecium • Ciliated heterotroph • Intracellular digestion • Food taken into oral groove and passes to gullet

19. Euglena • Both autotrophic and heterotrophic • Contains chlorophyll • Has eyespot which facilitates movement towards light • Can absorb food from outside the cell • Has flagellum

20. Chlorella • Single-celled green algae • Non-motile • Cell wall of cellulose

21. Microbes and biotechnology Option F.2

22. Assessment Statements • F.2.1 List the roles of microbes in ecosystems, including producers, nitrogen fixers and decomposers. • F.2.2 Draw and label a diagram of the nitrogen cycle. • F.2.3 State the roles of Rhizobium, Azotobacter, Nitrosomonas, Nitrobacter and Pseudomonas denitrificans in the nitrogen cycle. • F.2.4 Outline the conditions that favour denitrification and nitrification. • F.2.5 Explain the consequences of releasing raw sewage and nitrate fertilizer into rivers. • F.2.6 Outline the role of saprotrophic bacteria in the treatment of sewage using trickling filter beds and reed bed systems. • F.2.7 State that biomass can be used as raw material for the production of fuels such as methane and ethanol. • F.2.8 Explain the principles involved in the generation of methane from biomass, including the conditions needed, organisms involved and the basic chemical reactions that occur.

23. Role of microbes in ecosystems • Producers • Change inorganic molecules into organic molecules • Algae and some bacteria use chlorophyll to trap sunlight • Chemosynthetic bacteria use chemical energy • Nitrogen fixers • Bacteria remove nitrogen gas from the atmosphere and fix it into nitrates which are useable by producers • Decomposers • Breakdown detritus and release inorganic nutrients back into the ecosystems

24. Mutualistic nitrogen fixation: bacteria forms symbiotic relationship with a host plant and fix nitrogen for it (Ex.Rhizobium) Free-living nitrogen fixation: live in soil (Ex. Azotobacter) Industrial nitrogen fixation: burning of fossil fuels to produce fertilizer Nitrification: ammonia turned into nitrites by bacteria (Ex. Nitrosomonas) and to turn nitrites into nitrates (Ex. Nitrobacter) Active transport of nitrates: nitrates taken in by roots Plants and animals: plants use nitrates to make their own proteins; animals feed on plants, digest and rearrange proteins to make their own proteins Death and excretion: products of digestion and dead bodies contain molecules which contain nitrogen Putrefaction: decomposers break down complex proteins and release nitrogen gas into the atmosphere Denitrification: bacteria remove nitrates and nitrites and put nitrogen gas back into the atmosphere (Ex. Pseudomonas denitrificans) The nitrogen cycle

25. Nitrification Carried out by Nitrosomonas (ammonia into nitrate) Carried out by Nitrabacter (nitrite into nitrate) Available oxygen Neutral pH Warm temperature Denitrification Carried out by Pseudomonas denitrificans (nitrates into nitrogen gas) No available oxygen High nitrogen input Conditions which favor nitrification and denitrification

26. Consequences of releasing raw sewage and nitrate fertilizer into rivers • High nitrates and phosphates fertilize the algae present in water • Increased growth of algae • Algae are decomposed by aerobic bacteria which use up the oxygen in the water (biochemical oxygen demand – BOD) • Water becomes low in oxygen and fish and other organisms which need oxygen die

27. Sewage treatment by saprotrophic bacteria • Stages of sewage treatment: • Inorganic materials are removed and organic matter is left • 90% of the organic matter is removed by saprotrophic bacteria

28. Trickling filter system • Bed of stones 3-6 feet wide • Saprotrophic bacteria adhere to the stones and act on the sewage trickled over them until it is broken down • Cleaner water trickles out of the bottom of the bed • This flows to another tank where the bacteria are removed • The water is further treated with chlorine to finish the disinfectant process

29. Reed bed • Waste water provides water and the nutrients to the growing reeds • Reeds are then harvested for compost • Breakdown of organic waste is again accomplished by saprotrophic bacteria • Nitrate and phosphates released as a result of bacterial action are used as fertilizer by the reeds • Can only handle small sewage flow

31. Water Alert!

32. Homework • Fill in the blank: The three wastewater treatment plants in Mobile have a total design flow of _____ million gallons per day. • Read the following article: • http://www.usaid.gov/our_work/environment/water/us_japan_init.html • What has been done in the countries aided by this project?

33. Sewage treatment video

34. Biomass can be used as raw material for the production of fuels such as methane and ethanol • To make biogas, manure and cellulose are put into a digester without oxygen • Anaerobic decomposition is performed by bacteria which occur naturally in the manure • Manure and cellulose are broken down into organic acids and alcohol • Organic acids and alcohol are broken down into carbon dioxide, hydrogen, and acetate • Finally two type of bacteria work on these to produce methane which can be used to run electrical machinery • Ammonia and phosphate are byproducts and can be used as fertilizer

35. Conditions to be kept constant in digester No free oxygen Constant temperature of 95 degrees F pH (not too acidic) Bacteria required for methanogenesis Acidogenic bacteria convert organic matter to organic acids and alcohol Acetogenic bacteria make acetate Methanogenic bacteria create the methane

36. Topic F.3

37. Assessment Statements F.3.1 State that reverse transcriptase catalyses the production of DNA from RNA. F.3.2 Explain how reverse transcriptase is used in molecular biology. F.3.3 Distinguish between somatic and germ line therapy. F.3.4 Outline the use of viral vectors in gene therapy. F.3.5 Discuss the risks of gene therapy.

38. Background information • Genetic material of virus can be RNA or DNA • An RNA virus is called a retrovirus • Flow of genetic information in a retrovirus is from RNA back to DNA • The enzyme which enables backwards transcription is reverse transcriptase

39. HIV life cycle • HIV attaches to a host cell • RNA of the virus and the enzyme reverse transcriptase enter the host cell • Reverse transcriptase copies viral RNA into cDNA (complementary DNA) • cDNA makes a second strand which is a complement to the first strand of DNA; viral RNA is destroyed • New double-stranded viral DNA enters the nucleus of the host cell • If HIV is active, it will use this DNA to make more HIV viruses; they will then burst out of the cell and infect other cells

40. How reverse transcriptase is used in molecular biology • To make therapeutic proteins such as insulin and somatostatin • Human DNA molecule is taken from a pancreas cell • mRNA copies the DNA without the introns • Reverse transcriptase produces a new single strand of DNA called cDNA • The single strand replicates to make double-stranded DNA using DNA polymerase • Inserted into plasmid • Bacteria cell is stimulated to take up plasmid • Bacteria multiply and produce insulin • Insulin is harvested and used by diabetics

41. somatic and germ line therapy • Gene therapy aims to replace defective genes with effective ones which give the message to make the correct protein • Genes are delivered by vectors which are viruses that have been genetically engineered to infect certain cells in the patient • Somatic therapy– affects only the patient involved; may be possible to cure single-gene defects such as cystic fibrosis and hemophilia • Germ line therapy – changes gamete DNA, which would be passed to offspring

42. use of viral vectors in gene therapy • Treatment of a disease called severe combined immune deficiency disease (SCID) • Children do not produce enzyme ADA and thus have no immune system • In 1998 stem cells were withdrawn from kids with SCID • Cells mixed with a virus carrying normal form of gene that produces ADA • Virus transferred normal copy of gene into stem cells • Stem cells then infused back into bone marrow • Immune systems were restored

43. risks of gene therapy • Virus vector may enter another cell by mistake • Virus vector might put gene in wrong place and cause mutation • Genes could be over-expressed and too much protein produced • Virus vector might stimulate immune reaction • Virus vector might be transferred from person to person • Children might be more sensitive

44. Benefits of gene therapy • Possibility of curing a disease caused by a single-gene or multiple genes

45. F.4 Microbes and food production

46. Assessment Statements F.4.1 Explain the use of Saccharomyces in the production of beer, wine and bread. F.4.2 Outline the production of soy sauce using Aspergillus oryzae. F.4.3 Explain the use of acids and high salt or sugar concentrations in food preservation. F.4.4 Outline the symptoms, method of transmission and treatment of one named example of food poisoning.

47. The successful making of bread, beer, and wine • Yeast was discovered as the fungus responsible for products • Saccharomyces cerevisae • Yeast uses sugars for energy and reproduces quickly by ‘budding’ • Bud breaks off and forms a new yeast cell • Turns glucose into two molecules of ethanol and two molecules of carbon dioxide gas as waste product

48. Beer • Source of glucose: grain • Sweet liquid wort is made from malt • Hops are added and liquid is boiled and cooled • Fermentation by yeast produces beer containing ethanol and carbon dioxide

49. Wine • Crushed grapes and yeast are put into a tank • Ethanol stays in the tank, while carbon dioxide escapes

50. Bread • Fermentation of sugars in the dough by yeast • Carbon dioxide makes the dough rise • Baking in the oven kills the yeast, stops fermentation and evaporates the ethanol