Difference between Gram Positive and Gram Negative Bacteria

Difference between Gram Positive and Gram Negative Bacteria

1. GROUP 9 The contrast between gram positive and gram negative Group members : Siteri Donu -20170086 Synthia Achari -20170175 Kavinesh - 20170007 Rizwan Khan - 20160119 Akash Prasad - 20170038 Rutvi Kalavadiya - 20170112 Riya Patel - 20170153

2. Introduction Gram staining is the use of stains to differentiate between the two major types of bacteria Is called a differential stain for that very reason Helps to understand different characteristics of bacteria Has been perfected by modification of the original experiment over the years Widely used from high school experiments to clinical microbiology laboratories

3. History Method invented by Hans Christain Gram in 1884 Was originally devised to make bacteria more visible He observed that some bacteria destained after he washed it with ethanol (typhus bacillus) This unstaining of certain bacteria was due to certain bacteria having thin cell walls(He called those gram negatives) The stained ones were named gram positive His methods were revised and now we stain both gram negative and gram positives for better visual under the microscope .

4. Purpose Gram staining is used to differentiate between two different groups of bacteria The classification helps to identify some of the characteristics the bacteria may have Method was adopted and is widely used as it is cost effective, easy and the results are reliable Method should not be used to diagnosis or for specific identification as it only distinguishes one large group to another and not specific species of bacteria

5. The Gram Stain Procedure

6. Step 1 - Prepare a Smear “Bacteria” Suspend some of the material to be stained in a drop of water on a microscope slide, spread the drop to about the size of a nickel. Allow to air dry. Heat fix by gently warming above a flame or other heat source.

7. The Gram Stain Procedure Step 2 - Apply the Primary Stain Flood the Smear with Crystal Violet Allow to stand 30 sec to 1 min Rinse with water to remove excess stain

8. The Gram Stain Procedure Step 3 - Apply the Fixing Agent Flood the Smear with Iodine solution Allow to stand 30 sec to 1 min

9. The Gram Stain Procedure Step 4 - Rinse Rinse with water to remove excess Iodine

10. The Gram Stain Procedure Step 5 - Decolorize Drip 90% Alcohol across the slide about 5 sec The effluent should appear pale or clear

11. The Gram Stain Procedure Step 6 - Rinse Rinse with water to remove excess alcohol

12. The Gram Stain Procedure Step 7 - Counterstain Flood the slide with Safranin solution Let stand 30 sec

13. The Gram Stain Step 8 - Rinse, Dry and Observe Gram-Positive Gram-Negative Rinse with water to remove excess stain Blot dry Observe under Oil Immersion

14. DIFFERENCES BETWEEN THE GRAM POSITIVE BACTERIA AND GRAM NEGATIVE BACTERIA

15. Gram Positive Retain crystal violet dye and stain blue or purple Gram Negative Can be decolorized to accept counterstain (safranin) and stain pink or red Gram Reaction

16. Gram Positive Thick (multilayered) Gram Negative Thin (single-layered) Peptidoglycan Layer

17. Gram Positive Cell Wall is 20-30 nm thick. The wall is Smooth Gram Negative Cell Wall is 8-12 nm thick. The wall is wavy. Cell Wall

18. Gram Positive Present in many Gram Negative Absent Teichoic Acids

19. Gram Positive Absent Gram Negative Present Periplasmic Space

20. Gram Positive Absent Gram Negative Present Outer Membrane

21. Gram Positive Absent Gram Negative Occurs in Outer Membrane Porins

22. Gram Positive Virtually None Gram Negative High Lipopolysaccharide (LPS) Content

23. Gram Positive Low (acid-fast bacteria have lipids linked to peptidoglycan) Gram Negative High (because of presence of outer membrane) Lipoprotein Content

24. Gram Positive Quite Prominent Gram Negative Less Prominent Mesosomes

25. Gram Positive 2 rings in basal body Gram Negative 4 rings in basal body Flagellar Structure

26. Gram Positive Exotoxins Gram Negative Endotoxins or Exotoxins Toxin Produced

27. GRAM POSITIVE BACTERIA GRAM NEGATIVE BACTERIA Resistance to Physical Disruption high Low Isoelectric Range pH 2.5-4.0 4.5-5.5 Cell Wall Disruption by Lysozyme High Low (requires pretreatment to destabilize outer membrane) Susceptibility to Penicillin and Sulfonamide High Low Susceptibility to Streptomycin, Chloramphenicol and Tetracycline Low High Inhibition by Basic Dyes High Low Susceptibility to Anionic Detergents High Low Resistance to Sodium Azide High Low Resistance to Drying High Low Rendering They can rendered Gram - ve by increasing acidity They can rendered Gram + ve by increasing alkalinity

28. EXAMPLES OF GRAM POSITIVE AND GRAM NEGATIVE BACTERIA

29. GRAM POSITIVES Staphylococcus Cells are very round and deeply stained. Normal flora of skin and nose

30. Streptococcus Are most in a form of long chains. Normal Flora of mouth and GI tract.

31. Clostridium Anaerobic. Eubacteria Can cause Botulism, diarrhea.

32. Bacillus Gram positive rods – large , box car shaped, sometimes spores are visible. Sometimes they over-decolorize and appear Gram negative

33. Nocardia Delicate Gram positive branching rods - faintly staining. stain red with modified acid fast stains

34. ENTROBACTERIA GRAM NEGATIVE Escherichia Coli Normal Flora of human intestine. Facultatively anaerobic. Rod-shaped bacteria.

35. ENTROBACTERIA Salmonella Rod-shaped bacteria Causes Food Poisoning. Neisseria Neisseria gonorrhoeae causes gonorrhoea

36. Vibrio Curved or comma shaped gram negative rods

37. Helicobacter Helical shape Helicobacter pylori can cause peptic Ulcer and if not treated can lead to gastric cancer.

38. Gram-negative bacteria Gram-negative bacteria can cause many types of infections and are spread to humans in a variety of ways. Several species, including  Escherichia coli , are common causes of foodborne disease.  Vibrio cholerae —the bacteria responsible for cholera—is a waterborne pathogen. Gram-negative bacteria can also cause respiratory infections, such as certain types of pneumonia, and sexually transmitted diseases, including gonorrhea.  Yersinia pestis , the Gram-negative bacterium responsible for plague, is transmitted to people through the bite of an infected insect or handling an infected animal.

39. Gram-positive bacteria Bacillus anthracis (+ rod)-Anthrax Streptococcus pyogenes (+ coccus)-Rheumatic fever Corynebacterium diphtheriae (+ rod)-Diphtheria Clostridium botulinum (+ rod)-Botulism Streptococcus pyogenes (+ coccus)-Septic Sore Throat (pharyngitis) Staphylococus aureus (+ coccus)-Boils Clostridium tetani (+ rod)-Tetanus Clostridium perfringes (+ rod)-Gas gangrene Staphylococcus aureus (+ coccus)-Toxic shock syndrome

40. An investigation by Montie et al. has proposed that the bacterial cell divider may not be traded off by ROS from plasma, in spite of the fact that it enables ROS to infiltrate by dissemination, and just the cell layers (cytoplasmic film and external film) are defenseless against physical disturbance by plasma. Be that as it may, the dissemination over a thick Gram positive cell divider would probably still be slower than over a thin Gram negative cell divider. Journal article on gram positive and gram negative bacteria

41. Conclusion QUIZ

42. What is the effect of iodine on crystal violet ? what does alcohol do to the pepetidoglycan layer of the gram positive bacteria ? Which gram bacteria has two flagella rings in their basal body ? What is the ph difference between gram positive and gram negative bacteria ? What gram negative bacteria causes gonorrhoea? Which bacteria causes anthrax ? Which enterobacteria causes food borne diseases?

43. Reference   Mai- Prochnow , A., Clauson, M., Hong, J. and Murphy, A. (2016). Gram positive and Gram negative bacteria differ in their sensitivity to cold plasma.  Scientific Reports , 6(1). Claus, D. (1992). A standardized Gram staining procedure.  World Journal of Microbiology & Biotechnology , 8(4), pp.451-452. Malanovic , N. and Lohner , K. (2016). Gram-positive bacterial cell envelopes: The impact on the activity of antimicrobial peptides.  Biochimica et Biophysica Acta (BBA) - Biomembranes , 1858(5), pp.936-946.   Team, E. (2018).  Differences Between Gram Positive and Gram Negative Bacteria | LaboratoryInfo.com . [online] LaboratoryInfo.com. Available at: https://laboratoryinfo.com/differences-between-gram-positive-and-gram-negative-bacteria/ [Accessed 24 Feb. 2018].   Vlab.amrita.edu. (2018).  Gram Stain Technique (Theory) : Microbiology Virtual Lab I : Biotechnology and Biomedical Engineering : Amrita Vishwa Vidyapeetham Virtual Lab . [online] Available at: http://vlab.amrita.edu/?sub=3&brch=73&sim=208&cnt=1 [Accessed 24 Feb. 2018].