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Introduction Testing for antibiotic susceptibility can be used for drug development, epidemiology, and therapeutic outcome prediction. The antibiotic susceptibility test can determine the inhibition of bacterial growth in the specific antibiotics. These three are the most important method in determining antimicrobial susceptibility. Antimicrobial susceptibility testing (AST) is a laboratory procedure performed by medical technologists (clinical laboratory scientists) to identify which antimicrobial regimen is specifically effective for individual patients. On a larger scale, it aids in the evaluation of treatment services provided by hospitals, clinics, and national programs for the control and prevention of infectious diseases. Recently, researchers have had to implement continuous surveillance activities for resistance patterns due to the mutations in bacterial DNA. Types of infections caused by E.coli, S.aureus and pseudomonas aereginosa E.col E coli is a common bacterium that is found in the guts of humans.It produces harmful toxin which destroys the inner lining of small intestine.Also results in bloody diarrhoea.But in other words it keeps alimentary system healthy.The type of infections caused by it is cholocystis,infections of urinary tract traveler's diarrhoea and infection of natal meninges.The results shown by it are it is are gram negative (the cell wall is thinner than any other gram positive bacterium), besides that it is beta hemolytic. S.aureus Staphylococcus aureus upregulates the genes of virulence.Type of infection caused by it are if we are in direct contact with any contaminated person or infectious droplets or dispersion by sneezing of by cough.This bacterium shows positive results to grams stain as the cell wall becomes purple. Pseudomonas aereginosa Pseudomonas aereginosa is categorised under mechanisms of type 3 secretion.The weak immune system is taken advantage for the occurence of infection.It infects pulmonary system, infections of soft tissues, infection of dermal region, gastrointestinal infection and also affects joint regions(where two bones meet).The results shown by this bacterium is gram negative.It is cultured on the swab of wounds. mode of action and efficacy of citrus (Gram positive vs. Gram negative) The antimicrobial action of Quaternary ammonium compounds which are cationic salts of organically substituted ammonium compounds is rather wide-ranging and they are more effective for gram-positive bacteria. They work much less well against bacterial spores. Sometimes QACs are categorized as surfactants. For instance, benzalkonium chloride. The non-oxidizing disinfectants with the highest usage in the pharmaceutical sector are QACs. They work on the cell membrane, where they interact with phospholipids to cause cytoplasm leakage and coagulation. QAC (Quaternary ammonium compounds) work on the cell membrane, where they interact with phospholipids to cause cytoplasm leakage and coagulation. Testing for antibiotic susceptibility can be used for drug development, epidemiology, and therapeutic outcome prediction. The antibiotic susceptibility test can determine the inhibition

of bacterial growth in the specific antibiotics. These three are the most important method in determining antimicrobial susceptibility. 1. Agar disk-diffusion method – A standard inoculum of the test microorganism is used to inoculate agar plates. The test substance is then put in the desired concentration on 6 mm-diameter filter paper discs, which are then placed on the agar surface. The Petri dishes are incubated in the proper environments. An antimicrobial agent inhibits the growth of microorganisms that diffuses into the agar, and the diameters of the inhibitory growth zones are then determined. 2. Antimicrobial gradient method – The MIC value is calculated using the antimicrobial gradient approach, which combines the principles of diffusion and dilution procedures. It is predicated on the potential for establishing a gradient in the concentration of the antimicrobial agent examined in the agar medium. 3. Agar diffusion – It entails the antimicrobial agent being transferred by diffusion from the chromatogram such as thin- layer chromatography to an agar plate that has already been inoculated with the pathogen under investigation. The chromatogram is removed after a few minutes or hours to allow for diffusion, and the agar plate is then incubated. Where the antimicrobial chemicals come into contact with the agar layer is where the growth inhibition zones occur. Materials and Methods (Write in paragraph form. Can use subtitles.) Methods of MIC producer The MIC is determined by preparing of the chemical in vitro at increasing concentrations, incubating the solutions with separate batches of cultured bacteria, and measuring the results using agar dilution or broth microdilution. MIC Materials The basic design of a moving-coil dynamic microphone involves the following: Diaphragm with conductive coil attached. The diaphragm is typically made of polyester film (Mylar is a common brand) while the conductive coil is typically copper. Magnets and pole pieces . Method of MBC producer The MBC is identified by determining the lowest concentration of antibacterial agent that reduces the viability of the initial bacterial inoculum by a pre-determined reduction such as ≥99.9% . MBC detection uses a broth micro-dilution method similar to MIC . The detection process is as follows: First, a pure culture of a specific microorganism is grown overnight, and then diluted to an appropriate concentration in a growth-supporting broth (usually Mueller Hinton broth).

Material Then we took one double strength test tube with 2 ml MHB(mueller-hinton broth) in it. and 9 single strength test tubes with 2 ml MHB Results (Introductory paragraph for each table) Observation & Results for MIC Tube Dil. ½ ¼ 1/8 1/16 1/3 2 1/6 4 1/12 8 1/25 6 1/51 2 1/102 4 Result -ve -ve -ve -ve -ve -ve -ve +ve +ve +ve Mic value = 1/128 –ve Class Data – for E.coli Sr no. MIC MBC 1 1/512 1/512 2 1/1024 1/256 3 1/128 ----- 4 1/64 1/64 5 1/128 1/128 6 1/128 1/128 7 1/128 1/128 8 1/512 1/512 Calculations for MIC & MBC Citrus (1.6%) – MIC – 20 west conc. Highest dilution of stock to kill bacteria SS MHB No overuse of antibiotic, microbes will become resistant to it so use with minimum one  2 stocks cone bottle Find conc, which bacteria grow dilution  chance of TST group me ds MHB = 24 X 2ml = 48ml = 52.8ml = 60ml dsMHB 21g/L SS MHB 21/1000 X 60= 126/100 = 1.26g/60ml dh 2 O ds MHB 2 X = 2X1.26 = 2.52 g/ml SSMHB = 21/100X550 = 11.55g/550 ml SSMHB =14X2=28+2.8=30.8ml MIC = turlrdity MBC = minimum bactericidal concentration killing bacterial static (refrigerator) Now conditions -ve might to +ve on media MBC = 24X20ml = 480ml+48ml = 528ml = 55ml

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MHB = 21g/L = 11.55g + Agar 15g/L MBC = 15/1000ml X 550 = 8.28gn/550 Media 20ml dH 2 O + 2052g +add 20ml dH 2 O Clear solution on it hotplate 2 X 24 48 ml citrus +6ml citrus = 54ml + 5.4 = 60ml Means 30 ml for each organism 90ml  Table with control results /2   Table with conclusive Individual MIC & MBC results /2  Calculation of % Citrus for Individual MIC & MBC /0.5  Table with Class average only for MIC & MBC of each organism (**combined table) /2.5

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