o They called this hypothetical process Spontaneous Generation.

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1 PHARMACEUTICAL MICROBIOLOGY JIGAR SHAH INSTITUTE OF PHARMACY NIRMA UNIVERSITY

2 History & Scope of Microbiology What is Microbiology? Study of living organisms of microscopic size, include BACTERIA, FUNGI, ALGAE, PROTOZOA & the infectious agents called VIRUSES. Study of their form, structure, generation, reproduction, physiology, metabolism & classification. Study of their distribution in nature, their relationship to each other & to other living organisms, their effect on human beings & on other animals & plants. Their abilities to make physical & chemical changes in our environment. Microorganisms are associated with health & welfare of human beings

3 HISTORY Before microorganisms were discovered, some investigators like Bacon, Fracstoro, Lucretius suggested that disease was caused by invisible living creatures. But there was no proof. In 1658, Kircher was first person to recognize significance of these living creatures in disease by referring them in decaying bodies, meat, milk & diarrhoeal reactions. However, the first person to observe & describe microorganisms accurately was Van Leeuwenhoek in He sent detailed letters describing his discoveries to the Royal Society of London. It was clear that he saw both bacteria & protozoa.

4 HISTORY o After van leeuwenhoek discovered the previously invisible world of microorganisms, the scientific community of the time became interested in origins of these tiny living things. o Upto the second half of the nineteenth century, many scientists and philosophers believed that some forms of life could arise spontaneously from nonliving matter. o They called this hypothetical process Spontaneous Generation.

5 HISTORY o The issue of spontaneous generation was resolved by French Scientist Louis Pasteur in o With a series of ingenious and persuasive experiments, Pasteur demonstrated that microorganisms are present in the air and can contaminate sterile solutions, but air itself does not create microbes. o Pasteur s views are 1) m.orgs can be present in non living matter, 2) microbial life can be destroyed by heat, 3) methods can be devised to block the access of airborne m.orgs. To nutrient environments.

6 HISTORY Important events in Development of Microbiology Muller produces first classification of bacteria 1798 Jenner introduces cowpox vaccination for smallpox 1857 Pasteur shows that lactic acid fermentation is due to a microorganism 1869 Miescher discovers nucleic acids 1876 Koch demonstrates that anthrax is caused by Bacillus anthracis 1880 Laveran discovers plasmodium, the cause of malaria. and so on till the date in future

7 SCOPE We live in age of Bacteria first living organisms on our planet - live virtually everywhere life is possible - more numerous, - Constitute largest component of earth s biomass - Whole ecosystem depends on their activities - Influence human society in countless ways - Microbiology has a great impact on fields such as medicine, agricultural & food sciences, ecology, genetics, biochemistry & Molecular biology. So MODERN MICROBIOLOGY is large discipline with many different specialities.

8 SCOPE Microbiology has been contributor to the rise of Molecular Biology Branch of Biology dealing with physical chemical aspects of living matter & its function. Microbiologists involved in study of genetic code & mechanism of DNA, RNA & protein synthesis, recombinant DNA technology & genetic engineering. Some microbiologists microorganisms: focus on specific group Virologists on viruses; mycologists on fungi; bacteriologists on bacteria; protozoologists on protozoa; algologists on algae. of

9 SCOPE Some work on microbial cytology, microbial physiology, microbial ecology, microbial genetics, microbial taxonomy, molecular biology. Some work on medical microbiology (human & animal disease study), food & dairy microbiology, public health microbiology (communicable diseases). Immunology Immune system against pathogens - Fastest growing area in science - E.g. production and use of monoclonal antibodies Agricultural Microbiology Impact of organisms on agriculture, combat plant diseases, to increase soil fertility, crop yields Microbial ecology Relationship between organisms & their living/nonliving habitats. Their contribution to C,N & S cycles in soil & freshwater. Study of pollution

10 FUTURE Microbiologists have to work against widespread & destructive new & old infectious disease like AIDS, SARS, Tuberculosis. To find ways to stop spread of established infectious diseases, Discover of new drugs, new vaccines, use of recombinant DNA technology, molecular biology M.organisms important in industry & environmental control like sometimes serve as sources of high quality food, their enzymes for industrial applications and increase agricultural productivity. To study of genome sequence, genome relation to cell structure, analysis of genome for advances in bioinformatics. Understanding of microorganisms help more effectively in control of pollution

11 BACTERIA Bacteria is a plural word. The singular for this word is bacterium (bacter = rod, staff). Bacteria are prokaryotes (Kingdom Monera), which means that they have no true nucleus. They do have one chromosome of double-stranded DNA in a ring. They reproduce by binary fission. Most bacteria lack or have very few internal membranes, which means that they don t have some kinds of organelles (like mitochondria or chloroplasts). Most bacteria are benign (benign = good, friendly, kind) or beneficial, and only a few are bad guys or pathogens.

12 Difference between procaryotes and eucaryotes Property Procaryotes Eucaryotes Size Small cells 1-10mm10mm Large cells mm100mm Organisms Oxygen tolearation Cell division Bacteria Anaerobic, aerobic No mitosis, by binary fission Fungi, protozoa, algae, virus aerobic Variuos forms of mitosis,

13 BACTERIA There are two different ways of grouping bacteria. They can be divided into three types based on their response to gaseous oxygen. Aerobic bacteria require oxygen for their health and existence and will die without it. Anerobic bacteria can't tolerate gaseous oxygen at all and die when exposed to it. Facultative aneraobes prefer oxygen, but can live without it. The second way of grouping them is by how they obtain their energy. Bacteria that have to consume and break down complex organic compounds are heterotrophs. This includes species that are found in decaying material as well as those that utilize fermentation or respiration. Bacteria that create their own energy, fueled by light or through chemical reactions, are autotrophs.

14 BACTERIA

15 BACTERIA Bacteria Cell structure: Surface Structure: Beginning from the outermost structure and moving inward, bacteria have some or all of the following structures: 1) Capsule: Protective covering, made up of polysaccharides, - Important role to keep the bacterium from drying out and to protect it from phagocytosis (engulfing) by larger microorganisms. - The capsule is a major virulence factor in the major disease-causing bacteria, such as Escherichia coli and Streptococcus pneumoniae. 2) Cell Wall: Composed of peptidoglycan (polysaccharides + protein), the cell wall maintains the overall shape of a bacterial cell. The three primary shapes in bacteria are coccus (spherical), bacillus (rod-shaped) and spirillum (spiral). Mycoplasma are bacteria that have no cell wall and therefore have no definite shape. - Protect cyctoplasmic membrane from the environment. - also helps to anchor appendages like the pili and flagella.

16 3) Cytoplasmic membrane: A layer of BACTERIA of Phospholipids & proteins -Encloses interior of bacterium, Regulating flow of materials in & out of cell. Internal structure: Bacteria have a very simple internal structure, and no membrane-boundbound organelles. 1) Nucleoid: The nucleoid is a region of cytoplasm where the chromosomal DNA is located. It is not a membrane bound nucleus, but simply an area of the cytoplasm where the strands of DNA are found. Most bacteria have a single, circular chromosome that is responsible for replication, although a few species do have two or more. Smaller circular auxiliary DNA strands, called plasmids, are also found in the cytoplasm.

17 BACTERIA 2) Ribosomes: Ribosomes give the cytoplasm of bacteria a granular appearance in electron micrographs. Though smaller than the ribosomes in eukaryotic cells, these inclusions have a similar function in translating the genetic message in messenger RNA into the production of peptide sequences (proteins). 3) Storage granules: (not shown) Nutrients and reserves may be stored in the cytoplasm in the form of glycogen, lipids, polyphosphate, or in some cases, sulfur or nitrogen. 4) Endospore: (not shown) Some bacteria, like Clostridium botulinum, form spores that are highly resistant to drought, high temperature and other environmental hazards. Once the hazard is removed, the spore germinates to create a new population.

18 Appendages: Bacteria may have the following appendages: BACTERIA 1) Pili: Many species of bacteria have pili (sing. pilus), small hairlike projections emerging from the outside cell surface. Assist the bacteria in attaching to other cells and surfaces, such as teeth, intestines, and rocks. Without pili, many disease-causing bacteria lose their ability to infect because they're unable to attach to host tissue. Specialized pili are used for conjugation, during which two bacteria exchange fragments of plasmid DNA. 2) Flagella: Flagella (singular, flagellum) are hairlike structures that provide a means of locomotion for those bacteria that have them. They can be found at either or both ends of a bacterium or all over its surface. The flagella beat in a propeller-likelike motion to help the bacterium move toward nutrients; away from toxic chemicals; or, in the case of the photosynthetic cyanobacteria; toward the light.

19 BACTERIAL CLASSIFICATION Classification means the ordering of groups into higher units. The basic unit, i.e. the pure culture of the isolated bacterium, is the STRAIN. Strains are collected into SPECIES, and species are collected into GENERA, these are collected in to FAMILIES, families collected into ORDER, order collected into CLASS, class collected into PHYLUM, phylum collected into KINGDOM. Such classification required adequate description of strains and proceeds by comparing the properties of various groups. Phyletic or Phylogenetic classification is based on evolutionary relationships. Because of lack of a good fossil record, this classification has proven difficult for bacteria.

20 BACTERIAL CLASSIFICATION So to classify bacteria satisfactory, phenetic classification system was accepted, available in Bergey s Manual of Systematic Bacteriology. In this classification system, characteristics used to define sections are normally features such as general shape and morphology, - Gram-staining properties, - oxygen relationship, - motility, - presence of endospores, - the mode of energy production,etc. The Bergey s Manual of Systematic Bacteriology places all bacteria in the kingdom procaryotae which in turn is divided into 4 divisions as follows:

21 Procaryotae Gracilicutes (thin skin) -Gram-negative Class I.Scotobacteria Nonphotosyn II.Anoxyphotoba cteria Photosyn, no O 2 III.Oxyphotobac teria Photosyn, evolve O 2 Firmicutes (strong skin) -Gram-positive Class I.Firmibacteria Rods or cocci II. Thallobacteria Branching cells (Actinomycetes) Tenericutes (Soft or tender skin) -Lacking of cell wall Class I.Mollicutes (mycoplasmas) Mendosicutes (skin with faults) -Unusual cell walls Class I Archaeobacteria

22 Filamentous Bacteria: Morphological Classification 1) Actinomyces: Gram positive, non-acid-fast, - Tend to fragment into short coccal and bacillary forms, anaerobic, e.g. A. israeli 2) Nocardia: similar to actinomyces, but aerobic, acid-fast e.g. N. farcinica 3) streptomyces: Vegetative mycelium, not fragmenting into short forms, e.g. S. griseus True Bacteria: (A) Cocci: 1) Staphylococcus & Micrococcus: - Arranged in irregular clusters, due to successive division occurring irregularly in different planes, gram positive, e.g. S. aureus

23 2) Streptococcus: - Arranged in chains, due to successive cell divisions ocurring in same axis, Gram positive, e.g., S. pyogenes 3) sarcina: - Arranged in cubical packets of eight, or multiple of 8, due to division occurring in 3 planes at right angles, Gram positive, e.g. S. lutea 4) Neisseria: - Arranged in pairs and slightly elongated at right angles to axis of pair, Gram negative, e.g. N. meningiditis. 5) Veillonella: - Very small cocci arranged in clusters and pairs, Gram negative, e.g. V. parvula. (B) Bacilli: 1. Acid-fast bacilli: Z.N. positive, e.g. M.tuberculosis

24 2. Gram positive spore forming bacilli: - Either aerobic (Bacillus) or anaerobic (clostridia) 3. Gram positive non sporing bacilli: -Corynebacteria club shaped, slightly curved - Erysipelothrix grow in chains or filaments - Lectobacilli grow in filaments but anaerobic - Listeria flagella, motile 4. Gram negative bacilli: e.g. pseudomonadaceae, achromobacteriacae, enterobactericeae, Brucellaceae and bacteridaceae. (C) Vibrios: Curved-rod-shaped cells, short, gram negative, motile, polar flagellum, e.g. V. Cholerae (D) Spirilla : spirally twisted, gram negative, e.g. S. minus (E) Spirochaetes: Slender, spiral filaments, Gram negative, motile without flagella

25 1. Borrelia: 8-12 fibrils, e.g. B. recurrentis 2. Treponema: Seen by dark-ground microscopy, 4 fibrils, e.g. T. pallidum 3. Leptospira: too many fibrils, e.g. L.interrogans Morphology of Bacteria: Size: Bacterium Length Disease Staphylococcus 0.8µ Carbuncles M. Tuberculosis 0.5-4µ Tuberculosis S. typhii 0.5-4µ Typhoid Vibrio comma 1-5.0µ Cholera N.Meningitidis 1.0µ Meningitis C.Tetani µ Tetanus P. Pestis 1-2µ Plague

26 Shapes and arrangements of bacterial cell 3 conventional shapes of the cells 1) Ellipsoidal or Cocci: appears like a minute sphere. Classify a)micrococci appears singly b)diplococci appear in pairs c)streptococci- appear in rows of cells or in chains d)tetrads arrange in square of four e)staphylococci arrange in bunches of grapes f)sarcinae arrange in cuboidal or packet arrangement 2) Cyllindrical or Rod Shaped: Bacilli - Appear like stick or rod, some times curved a) Diplobacilli: appear in pairs b) Streptobacilli: appear in chains 3) Spiral or helicoidal shaped: helical forms known as vibrio or spirilla. Appear like cork-screw.

27 Different shapes of bacteria and positions of flagella.

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