5. Fundamental principal of microbioology (2).pptx

Social Pharmacy ER 2020

1. 4. Fundamental principal of microbiology By, Argade V. P. Asst. Prof. PRES, Institute of Pharmacy Loni . Social Pharmacy (20115)

2. microbiology Microbiology can be defined as the study of living organisms of microscopic size which include bacteria, fungi, algae, protozoa & viruses. Some microbes are beneficial & others are harmful to man.

3. Scope of Microbiology Aero-Microbiology — helps in the study of food-prone diseases, and their ultimate prevention . 2 . Beverage Microbiology — making of beer, brandy, wine, and a variety of alcoholic beverages e.g .,whisky , brandy, rum, gin, vodka etc. 3. Exo -microbiology — to help in the exploration of life in the outer-space. 4. Food Microbiology — making of cheese, yogurt 5. Geochemical Microbiology — to help in the study of coal, mineral deposits, and gas formation; prospecting the deposits of gas and oil, coal, recovery of minerals from low-grade ores.

4. 6 . Industrial Microbiology — making of ethanol, acetic acid, lactic acid, citric acid, glucose syrup, highfructose syrup . 7. Medical Microbiology — helps in the diagnostic protocol for identification of causative agents of various human ailments, and subsequent preventive measures. 8. Pharmaceutical Microbiology — making of life-saving drugs, ‘antibiotics’ e.g., penicillins , ampicillin, chloramphenicol , ciprofloxacin, tetracycline's, streptomycin. 9. Soil and Agricultural Microbiology — helps in the maintenance of a good farm land by keeping and sustaining a reasonable and regular presence of microbes in it. 10. Waste-Treatment Microbiology — treatment of domestic and industrial effluents or wastes by lowering the BOD and COD

5. Classification of microorganism Prokaryotics :Microorganism with relatively smaller & simpler form of cells are known as Prokaryotics . It does not contain nucleus e.g . bacteria Eukaryotics : Microorganism with well developed cell structure are known as Eukaryotics . It contain well defined nucleus e.g . Algae, Fungi, Protozoa Virus is neither Prokaryotics nor Eukaryotics because of its not well define structure.

6. 1. bacteria

7. Bacterial Cell structure Capsule : Cell surrounded by gelatinous or slimy material forming a protective covering over a cell wall. I t is made up of polysaccharides.   It protects the organism against environmental change.   Cell wall: I t is a rigid structure that surrounds the cytoplasmic membrane. I t is made up from polysaccharides molecule. T he wall give the shape and protect it from the environmental change.

8.   Cytoplasm: I t is a gel like Matrix composed of about 80% water, enzyme, carbohydrates, proteins, nucleic acid, lipids, inorganic ions and contain cell structure such as ribosomes and nucleoid.   Cytoplasmic membrane: I t is a semipermeable membrane beneath the cell wall. I t is composed of phospholipid and proteins. It control the passage of nutrient and waste product into and out of the cell.

9. Nucleus : Nucleus is not well define, but nuclear material is present near the center of the cell. I t contains DNA.   Ribosomes : Ribosomes are present in cytoplasm and it is a site of protein synthesis   Flagella : T hese are hair like structure that extend from cytoplasmic membrane through the cell wall. I t gives mobility to the bacterial cell.   Pili : It is small hair like structure present over outer surface of cell wall.

10. Virus V irus is non cellular infective agent that consists of nucleic acid molecule and it can able to multiply only within the living cell of host.   It causes following disease: common cold, influenza, measles, AIDS, smallpox, polio and rabies.   It can be classify depending upon presence of nucleic acid in them: 1 . DNA virus : it contains DNA in them. example poxvirus and hepatitis   2. RNA virus : it contain RNA in them. example influenza, Cholera, rabies, AIDS, rubella

11. A bove figure consists of nucleic acid surrounded by protein covering called capsids. T he capsid together with nucleic acid called the nucleo -capsid .  the capsid is form by a large number of capsomeres .   This virons may be envelope or non envelope. T he Spikes are present on the surface of the envelope are called peplomer .

12. Fungi Fungi  is a eukaryotic organism that can be unicellular or multicellular. Disease caused by fungi are: 1. Candidiasis 2. Otomycosis 3. Dermatophytosis 4. Blastomycosis

13. Classification of Fungi This classification depends on sexual spore formation of fungi and is divided into 1. Lower fungi : Phycomycetes 2. Higher fungi : Ascomycetes Basidiomycetes Fungi imperfecti

14. Protozoa Classify Protozoa. Flagellates , or Mastigophora - e.g. Giardia lamblia Amoebae or Sarcodina -e.g. Entamoeba histolytica Sporozoans , or Sporozoa -e.g. Plasmodium knowlesi C iliates , or Ciliophora e.g. Balantidium coli

15. Isolation of pure culture Culture : a method of multiplying microorganism in laboratory scale is known as culture. Colony : when microorganisms are grown on a solid medium from a single cell is known as Colony.   Pure culture : culture contain only one type of microorganism is known as pure culture. Mixed culture : when culture contains several type of microorganism is known as mixed culture.

16. methods of isolation of pure culture Streak plate method Spread plate method Pour plate method

17. 1. Streak plate method In this method small amount of sample transferred on a nutrient agar media in petriplate by using Nichrome -wire loop ( innoculating needle) which is previously sterilized by heating directly on the flame. This innoculating needle is streak across the surface of nutrient agar medium. Streaking can be done by following methods:  Square method  Four quadrant method  Zig-zag method The petridish are incubated at 37oC for about 18-24 hrs , after that individual colonies can be observed on the agar surface.

18. 2. Spread plate method Place a drop of diluted sample of pure culture on the surface of nutrient agar medium in a petridish . This drop is spread over the entire surface using sterilized glass rod. The petridish are incubated at 37oC for about 18-22 hrs , after that individual colonies can be observed on the agar surface.

19. 3. Pour plate method In this method the sample is diluted to concentration of about 100 microbes/cm3. The diluted sample (1 ml ) is pour in to the petry plate with nutrients agar medium & move it gently in the direction shown in fig. I ncubate at 37oc for 18 to 20 hrs & observe on agar surface .

20. staining Definition : Staining is defined as imparting color to the specimen with the purpose of its identification. Stains : Stains are the dye or reagent use for coloring the bacteria to observe them more clearly. Eg . Crystal violet, Methylene blue. Purpose of Staining: For greater visualization of cells For identification bacterial cell structure To differentiate the cell To classify the bacteria

21. techniques of staining Simple staining : In this method only one stain (dye) is used for identification of bacteria . 2 . Differential staining : In this method more than one stain (dye) is used for identification of bacteria. a ) Gram staining b) Acid fast staining c ) ZiehlNeelsen

22. 1. Simple staining: It is also called as monochrome technique . In this method only one stain (dye) is used . It is used to study morphology, size, shape of microbes. Procedure:  Smear is fixed, stain is put, stain is allowed to react for 30 sec to 3 min, wash smear with stream of cool water, dry and examine under oil immersion lens.

23. 2. Differential staining a ) Gram staining Grams staining method is a differential staining method for bacteria . Use to differentiate between gram positive & gram negative bacteria. Procedure : Smear is prepared on clean glass slide. Smear is air dried or fixed by gentle heating. Crystal violet solution is applied on smear as primary stain for about 1 to 2 min. Excess stain is remove by washing with running tap water . Slide is washed using Alcohol (95% soln.) till no colour is remove & observed under microscope. Bacteria is completely destained ( stain comes out) these are gram negative bacteria , If color will be same ie violet then it will be gram positive bacteria . Finally slide immerse in 0.2% aqueous saffranin solution so that gram negative bacteria which lost there color during destaining may shown as pink. Observation : Gram positive bacterial cells appear violet colored while Gram negative bacterial cells appear pink colored.

24. Gram Positive & Gram Positive Difference Sr. No. Gram Positive Sr. No. Gram Positive 1 Cell wall is simple 1 More complex cell wall 2 Outer membrane is absent 2 Outer membrane is present 3 Lipids are absent 3 Lipids are present 4 Cell wall is more thick 4 Cell wall is less thick 5 They retain crystal violet 5 They retain Saffranin 6 Appear in violet color 6 Appear in Red/Pink color

25. b) Acid fast Staining This method differentiate bacteria acid fast or non acid fast Dyes used- melachite green, methylene blue Procedure: Smear is prepared on clean glass slide. Smear is air dried or fixed by gentle heating. Methylene blue solution is applied on smear as primary stain for about 1 to 2 min. Excess stain is remove by washing with Acid & alcohol . If bacteria is not decolourised & retain a stain of dye such bacteria are known as acid fast bacteria . Observation:  Acid fast- not decolorized by acid and alcohol  Non acid fast- loose stain, decolorized by acid and alcohol

26. c) Ziehl-Neelsen stain method The Ziehl-Neelsen stain is a type of differential bacteriological stain used to identify acid-fast organisms, mainly Mycobacteria tuberculosis and M. Leprae Procedure: Smear is prepared on clean glass slide. Smear is air dried or fixed by gentle heating. Ziehl’s carbolfuchsin dye is applied on smear for about 10min . Heat to dry & rinse with water Decolonization is done by 20 % sulfuric acid Observe under oil immersion lenses. Tubercle bacilli are stained red & are rod in shape . Observation: Acid fast bacteria- appear pink or red Non acid fast bacteria appear- blue.

27. REFERENCE: 1.Social Pharmacy, By Jagtap D.B., Gaikwad V.S. S,V. Bhise , Brilliant publication 2.SOCIAL PHARMACY (First Year FY Diploma Pharmacy - PCI's ER 2020), By  Dr. S.B. Bhise ,  Mrs. M.S. Bhise  , Nirali Prakashan . 3.Social Pharmacy, By Dr. Virendra Kumar , Thakur Publication.

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