microbial History, ppt baca aau anand, msc

history, spontaneous generation, abiogenesis, pasteurs ,

1. Spontaneous Generation Conflict Living organisms could developed from nonliving or decomposing matter Redi, Needham, Spallanzani Louis Pasteur – heated the necks of flasks and drew them out – see figure – (disprove spontaneous generation) The role of microorganisms in Disease Bassi – showed that silkworm disease was caused by a fungus Berkeley and Pasteur showed that Microorganisms caused disease Joseph Lister – developed system for sterile surgery Robert Koch (1843 – 1910) Bacillus anthracis and anthrax – Koch’s Postulates Injected Mice !!!!

2. Koch’s Postulates Microorganism must be present in every case of the disease but absent from healthy individuals The suspected microorganism must be isolated and grown in pure cultures The disease must result when the isolated microorganism is inoculated into a healthy host The same microorganism must be isolated from the disease host Isolation of the microorganism Growth on potatoes, solidify with gelatin Fannie Eilshemius – agar 100 °C to melt, 50 °C to solidify Role of Pasteur and Robert Koch in the development of Microbiology

3. The Three-Domain System Figure 10.1

4. WHITTAKER’S CLASSIFICATION

5. Fluorescent in situ hybridization (FISH) rRNA sequence most useful genetic taxonomic method :- • The DNA sequence that encodes rRNA is highly conserved among bacteria of common ancestry. • Sequence DNA encoding rRNA of different isolates, determine % identity, have a "molecular clock" –an accurate determination of phylogenetic distance. • Phylogenetic trees are now established on basis of rRNA sequence.

6. Genetics DNA fingerprinting: Number and sizes of DNA fragments (fingerprints) produced by RE digests are used to determine genetic similarities. Ribotyping : rRNA sequencing Polymerase chain reaction (PCR) can be used to amplify a small amount of microbial DNA in a sample. The presence or identification of an organism is indicated by amplified DNA. (see lab) Fig 10.14: Electrophoresis of RE digest of plasmid DNA

7. Nucleic Acid Hybridization Fig 10.15 Single strands of DNA or RNA, from related organisms will hydrogen-bond to form a double-stranded molecule; this bonding is called nucleic acid hybridization. Examples of Applications: Southern blotting, DNA chips, and FISH

8. METHODS IN BACTERIAL IDENTIFICATION Sources of Essential Nutrients a. Photoautotroph – are photosynthetic. They are eventually consumed by other organisms including humans. b. Chemoautotroph – have unusual nutritional adaptations that requires neither sunlight or organic nutrients. Usually live harsh environments like sulfur hot springs. This group use inorganic compounds for energy and derive their carbon from carbon dioxide. c. Photoheterotrophs – use the energy of sunlight and derive their carbon from organic compounds. d. Chemoheterotrophs – derive both carbon and energy from organic compounds. This group is the most common group associated with humans and other animals. 1. Saprobe – free living organisms that feed primarily on organic debris from dead organisms. 2. Parasite - live in or on the body of a host, which they usually harm to some degree.

9. Property  Gram-positive Gram-negative Thickness of wall  thick (20-80 nm) thin (10 nm) Number of layers  1  2 Peptidoglycan (murein) content  >50% 10-20% Teichoic acids in wall Present absent Lipid and lipoprotein content  0-3% 58% Protein content  9% Lipopolysaccharide content  0  13% Sensitivity to Penicillin G  Yes no Sensitivity to lysozyme Yes no

10. Azospirillum Phylum:  Proteobacteria Classis:  Alpha Proteobacteria Ordo :  Rhodospirillales Familia :  Rhodospirillaceae Genus:  Azospirillum Species:  A.  amazonense  - A.  halopraederans  -  A.  largimobile  -  A.  doebereinerae  -  A.  lipoferum  -  A.  brasiliense free-living in the soil or associated with the roots of cereal crops or grasses. N2 fixing organisms   Gram negative proliferates under both anaerobic and aerobic conditions but it is preferentially micro- aerophilic fix the considerable quantity of nitrogen in the range of 20- 40 kg N/ha in the rhizosphere in non- non-leguminous plants such as cereals, millets, Oilseeds, cotton etc. The efficiency of Azospirillium as a Bio-Fertilizer has increased because of its ability of inducing abundant roots in several pants like rice, millets and oilseeds even in upland conditions. Considerable quantity of nitrogen fertilizer up to 25-30 % can be saved by the use of Azospirillum inoculant .

11. Family- Pseudomonadaceae Straight or slightly curved rods Motile by polar flagella Catalase positive and usually oxidase positive Can grow aerobically. Pseudomonas aeruginosa  produces soluble blue-green pigment and can be problematic in weakened hosts; some pseudomonads produce fluorescent pigments; pseudomonads produce a large number of enzymes that break down chemicals such as pesticides; problematic in hospitals; can be highly resistant to antibiotics; pseudomonads can be  denitrifiers  which takes nitrate as an electron acceptor and produces nitrogen gas potentially under anaerobic conditions Include pseudomonas, Xanthomonas and Zoogloea .

12. Cells are large blunt rods, oval or cocci . Motile Saprophytes occurs in soil, water and at plant rhizosphere . Fix N 2 under aerobic conditions. used as a Bio-Fertilizer for all non leguminous plants especially rice, cotton, vegetables etc. Role of liquid Azotobacter as a Bio-control agent found to produce some antifungal substance which inhibits the growth of some soil fungi like Aspergillus , Fusarium , Curvularia , Alternaria , Helminthosporium , etc. Family- Azotobacteraceae Scientific classification Domain: Bacteria Kingdom: Bacteria Phylum: Proteobacteria Class: Gammaproteobacteria Order: Pseudomonadales Family: Pseudomonadaceae / Azotobacteraceae Genus: Azotobacter

13. Family- Rhizobiaceae Scientific classification Kingdom: Bacteria Phylum: Proteobacteria Class: Alphaproteobacteria Order: Rhizobiales Family: Rhizobiaceae Genus: Rhizobium Frank 1889 Gram-negative,  motile, non- sporulating  rods The first species of rhizobia ,  Rhizobium leguminosarum , was identified in 1889 able to colonize the legume roots and fixes the atmospheric nitrogen symbiotically. highly specific to form nodule in legumes

14. Facultative anaerobic Gram-negative bacteria -Medically important group Family  Enterobacteriaceae  ( Enterics ) - inhabit the intestinal tract of mammals. Some are permenant residents and some are found in the diseased state. Most are active fermenters of glucose and other carbohydrates. Biochemical tests are used to identify them as well as serological techniques. Motile and nonmotile species; some have fimbriae for adhering to mucous membranes; some produce  bacteriocins  that lyse closely related species. Includes  Escherichia, Salmonella, Shigella , Klebsiella , Serratia , Enterobacter , Proteus, Yersinia   and   Erwinia .

15. Anaerobic, Gram-negative, straight, curved and helical rods Bacteroides -   Straight rods live in the human intestinal tract in numbers reaching 1 billion per gram of feces. Some species reside in anaerobic environments such as the gingival crevice or deep tissue infections.  Bacteroides  are non-spore forming and non-motile. Fusobacterium -   are long and slender with pointed ends rather than blunt .   found in the gingival crevice and may be responsible for abscesses.

16. Dissimilatory sulfate-reducing or Sulfur reducing bacteria Desulfovibrio the best known species. Obligately anaerobic that use oxidized forms of sulfur, e.g., sulfate or elemental sulfur, as an electron acceptor during  anaerobic respiration . The end product is hydrogen sulfide which is released into the atmosphere. These organisms are found in anaerobic muds and sediments, the intestinal tract of animals. The reduced sulfur is an important substrate of a number of chemolithotrophs and anoxic photosynthetic bacteria.

17. Rickettsias and Chlamydias Obligate intracellular parasites. Rickettsias   are rod shaped or coccobacilli gram negative bacteria. Most are transmitted to humans by insects and ticks except  Coxiella burnetii  which causes Q-fever. It is transmitted by aerosols or contaminated milk.  Chlamydias are gram-negative coccoid bacteria. Transmitted by interpersonal contact or by airborne respiratory routes.  Chlamydia  species cause blindness in humans, nongonococcal urethritis , a common STD

18. Gram-positive cocci 1) Aerobic cocci They posses cytochromes . able to respire with oxygen. some may able to fermentation. high resistance to gamma and UV rays. Staphylococcus   occur in grapelike clusters.  S. aureus  is one of the more important species.   grow well under conditions of high osmotic pressure and low moisture  produces many toxins to aid in its pathogenicity . common in nosocomial infection and more resistant to many of the common antibiotics.   produces enterotoxins that cause vomiting and nausea a common food poison.

19. 2) Aerotolerant fermentative cocci - Do not posses cytochromes . cells are arranged in pairs, chains, or tetrades . Have only fermentative type of metabolism. catalase - negative. Streptococcus   are spherical gram-positive bacteria that usually occur in chains.  S. pneumoniae  occurs as a diplococci . May be aerotolerant or obligately anaerobic. Cause a diseases such as scarlet fever, sore throat, and pneumococcal pneumonia. Classified according to their action on blood agar plates: Alpha-hemolytic strains produce alpha hemolysin that reduces hemoglobin to methemoglobin which is greenish. Beta-hemolytic species produce hemolysins that lyse the red blood cells and zones of clearing are produced. Gamma-hemolytic species produce no reaction on blood agar plates

20. Endospore forming Gram-positive rods and cocci Important medically and in the food industry Their spores are extremely resistant to heat and many chemicals. Range from aerobic to strict anaerobes. Bacillus  and  Clostridium  are two important genera.

21. Bacillus anthracis  causes anthrax, a disease of cattle, horses and sheep that can go to humans. Nonmotile , facultative anaerobe, rod-shaped. Endospore is centrally located. Bacillus   thuringiensis  is a insect pathogen. Produces a protein that is toxic to insects and causes them to have a bad stomach ache. Sold commercially as Bt Toxin under the brand names such as Dipel . Bacillus Scientific classification Domain: Bacteria Division: Firmicutes Class: Bacilli Order: Bacillales Family: Bacillaceae Genus: Bacillus

22. Clostridium  are obligate anaerobes. Rod-shaped, resemble large, straight or slightly curved rods with rounded ends. consists of around 100 species Form endospores that appear swollen.  In their active form, these bacteria secrete powerful exotoxins that are responsible for such diseases as tetanus (lockjaw), botulism, and gas gangrene etc. The four most notable species of Clostridium are  C. tetani , C. difficile , C. perfringens , and C. botulinum C . tetani  causes tetanus;  C. botulinum  causes botulism; and other species cause gas gangrene. Clostridium

23. Regular nonsporing Gram-positive rods Lactobacillus   Lack a cytochrome system, therefore ferment to yield lactic acid. The acidity creates an ecological niche suited for the lactobacilli. Found in intestinal tract, and oral cavity. Used in food industry to make sauerkraut, pickles, buttermilk and yogurt. Listeria monocytogenes   a pathogen of this group. Found in food especially dairy products. survives within a phagocytic cells grows in refridgeration . Can cause stillbirths in pregnant women.

24. corynebacteria   Pleomorphic . Includes  Corynebacterium diptheriae  which causes diptheria . Another species is  Propionibacterium acne  that is common on skin and is implicated in acne. Mycobacteria  are fungus-like" as their name implies, because they have a filamentous growth pattern occassionally . Cell wall similar to gram-negative bacteria but the outermost layer, LPS, is replaced with mycolic acid, which forms a waxy, water-resistant layer. Resistant to desiccation and many drugs because of the mycolic acid layer. Includes  M. tuberculosis  which causes tuberculosis and  M. leprae  which causes leprosy. Irregular nonsporing Gram-positive rods

25. HALOPHILES Live in extremely SALTY conditions. Can grow in water that is up to ten times saltier than seawater. Such as the great salt lake in Utah, and dead sea . These organisms use salt to generate ATP. Most halophilic and all halotolerant organisms expend energy to exclude salt from their cytoplasm to avoid protein aggregation (‘ salting out ’). They maintain osmotic balance with their surroundings by building up the solute concentration within their cells. Example:- Pyrococcus furiosus , C hromohalobacter beijerinckii .

26. METHANOGENS Live in oxygen-free environments (anaerobic conditions) and produce methane gas. Harvesting energy by converting H 2 and CO 2 into methane gas. Can live only in anaerobic conditions, such as the bottom of swamps and in sewage. Example:- Methanococcoides burtonii Methanobacterium thermoautotrophicum .

27. The methane produced by methanogens living in the waters of SWAMPS, SEWAGE , or MARSHES is called SWAMP GAS.   The reduction of carbon dioxide into methane in the presence of hydrogen can be expressed as follows: CO 2 + 4 H 2 → CH 4 + 2 H 2 O Methane produced in the DIGESTIVE TRACTS of many animals including humans is called INTESTINAL GAS. They are used in INDUSTRY to treat SEWAGE and to help PURIFY WATER. Economic importance

28. Viruses can be placed in one of the seven following groups: I: dsDNA viruses (e.g. Adenoviruses, Herpesviruses , Poxviruses) II: ssDNA viruses (+)sense DNA (e.g. Parvoviruses ) III: dsRNA viruses (e.g. Reoviruses ) IV: (+) ssRNA viruses (+)sense RNA (e.g. Picornaviruses , Togaviruses ) V: (−) ssRNA viruses (−)sense RNA (e.g. Orthomyxoviruses , Rhabdoviruses ) VI: ssRNA -RT viruses (+)sense RNA with DNA intermediate in life-cycle (e.g. Retroviruses) VII: dsDNA -RT viruses (e.g. Hepadnaviruses )

29. Prions Prions , named for their description as " proteinaceous and infectious particles", lack any detectable (as of 2002) nucleic acids or virus-like particles. They resist inactivation procedures that normally affect nucleic acids.   Mammalian prions : Agents of spongiform encephalopathies Fungal prions : PSI+ prion of Saccharomyces cerevisiae URE3 prion of Saccharomyces cerevisiae RNQ/PIN+ prion of Saccharomyces cerevisiae Het-s prion of Podospora anserina

30. The lytic and lysogenic cycles of phage  , a temperate phage Many cell divisions produce a large population of bacteria infected with the prophage. The bacterium reproduces normally, copying the prophage and transmitting it to daughter cells. Phage DNA integrates into the bacterial chromosome, becoming a prophage. New phage DNA and proteins are synthesized and assembled into phages. Occasionally, a prophage exits the bacterial chromosome, initiating a lytic cycle. Certain factors determine whether The phage attaches to a host cell and injects its DNA. Phage DNA circularizes The cell lyses, releasing phages. Lytic cycle is induced Lysogenic cycle is entered Lysogenic cycle Lytic cycle or Prophage Bacterial chromosome Phage Phage DNA Figure 18.7

31. The importance of extremophiles Extremophiles have tremendous potential in biotechnology because of their ability to produce unique stable proteins with high technological value. A number of enzymes, such as ones involved in DNA processing, from several different extremophiles are already on the market. The importance of extremophile research to biology should not be underestimated. It is only through the search for and elucidation of extremophilic microorganisms that we have the universal phylogenetic tree proposed by Carl Woese in 1990. Molecular studies of RNA sequences from the extremophilic Archaea showed that these microorganisms were distinct from bacteria, giving rise to the three domains of life that exist today: Bacteria, Archaea and Eucarya

32. Definition - Foodborne Infection Disease organism is carried through food to the host. Illness occurs after eating a food containing living, harmful microorganisms. An invasion and multiplication of microorganisms in body tissues, typically in the intestines Symptoms - usually gastrointestinal, may take many hours or days to appear

33. Clostridium botulinum - botulism Rare but very serious , often fatal or long-term recovery. Symptoms: double vision, constipation; difficult eye movement, swallowing, speech, loss of autonomic nervous system function Spore forming, anaerobic - organism becomes vegetative and produces toxin Vegetative and spore forms found in soil, water, intestinal tracts of animals, fish

34. Electron Transport Chain

35. Copyright © McGraw-Hill Companies, Inc. Permission required for reproduction or display. 36 Figure 6.1 no increase maximal rate of division and population growth population growth ceases decline in population size

36. Copyright © McGraw-Hill Companies, Inc. Permission required for reproduction or display. 37 The Mathematics of Growth generation (doubling) time time required for the population to double in size mean growth rate constant number of generations per unit time usually expressed as generations per hour

37. Calculating growth rate from the exponential growth curve generation time (g) = t/n where t = elapsed time (hr) and n= number of generations n= 3.3(logN-logN ) N = number of cells at the end of the elapsed time N = number of cells at the beginning see sample calculation page 143. k=growth rate ( generations per hour) = 0.693/g = ln2/g

38. Copyright © McGraw-Hill Companies, Inc. Permission required for reproduction or display. 39 Solutes and Water Activity water activity (a w ) amount of water available to organisms reduced by interaction with solute molecules (osmotic effect) higher [solute]  lower a w reduced by adsorption to surfaces (matric effect)

39. Copyright © McGraw-Hill Companies, Inc. Permission required for reproduction or display. 40 The Chemostat rate of incoming medium = rate of removal of medium from vessel an essential nutrient is in limiting quantities Figure 6.9

40. Anaerobic Respiration: Glycolysis & Fermentation Glycolysis generates NADH which is converted back to NAD by the lose of a hydrogen. The hydrogen is added to pyruvate and, depending on the bacterial species, a variety of metabolic end-products are produced. NADH --> NAD Pyruvate ---------------------------> Short chain alcohols, (C3) fatty acids (such as lactic acid or ethanol) (C2-C4)

41. Aerobic Respiration: Glycolysis & the Tricarboxylic acid cycle (Krebs Cycle) Pyruvate is fully broken down to carbon dioxide (C1) and NAD is converted to NADH. NADH is generated from two sources (glycolysis and the Krebs cycle). Oxidative phosphorylation converts excess NADH back to NAD and in the process produces more ATP (stored energy). Ubiquinones and cytochromes are components of the electron transport chain involved in this latter process. Conversion of oxygen to water is the final step in the process.

42. Molecular-Based Methods -ecology Molecular techniques have been employed, basically involving the application of hybridisation probes, PCR amplification of rDNA genes and other DNA fingerprinting techniques. These include terminal restriction fragment length polymorphism (T-RFLP), amplified rDNA restriction analysis (ARDRA), amplified random intergeneric spacer analysis (ARISA), denaturing gradient gel electrophoresis (DGGE), temperature gradient gel electrophoresis (TGGE), oligonucleotide fingerprinting of rRNA genes or single-stranded conformation polymorphism (SSCP) and have been used frequently in combination with traditional techniques to analyses fungal community composition

43. PCR fingerprinting PCR fingerprinting can be accomplished by several different methods, all of the which are aimed at distinguishing differences differences in the genetic makeup of microbial population from different samples. DGGE and TGGE are identical in principle. Both techniques impose a parallel gradient of denaturing conditions along a polyacylamide gel.

44. Process for Making Alcoholic Beverages (1) Starch Material Steaming Saccharifying / Fermentation Saccharifying Saccharifying Fermentation Fermentation Batch Distillation Makgeolli Yakju Cheongju Soju (1) Kaoliang Beer Bourbon whisky Canadian whisky Vodka(1) Nuruk or Koji Malt Yeast α -amylase Glucoamylase Yeast Once or twice Scotch whisky Irish whisky Neutral Spirits Soju(2) Vodka(2) Steaming Steaming Batch Distillation Continuous Distillation Continuous Distillation Twice Yeast

45. Process for Making Alcoholic Beverages (2) Sugar Material Spirits-Based Fermentation Batch Distillation Continuous Distillation Yeast Liqueur Brandy Wine Wine spirits Rum Extraction / Blending / …. Fermentation Yeast Steaming Batch Distillation Tequila Twice Twice Fruits, Botanicals, etc .

46. Process for Making Wine Fruits Stemming Crushing/ Pressing Fermentation Press Settling Maturing Yeast Filtration Fruits Wine Sulfurous Acid

47. Process for Making Rice Wine Previous Treatment / Steaming Nuruk or Koji Yeast Seed Culture First Fermentation Press Filtration Blending Pasteurization Bottling Second Fermentation Sludge Ingredients Treated Water Rice Rice wine Nuruk