ANTIBIOTIC SENSITIVITY REPORTS WITH DISK DIFFUSION METHODS (Believe it or Not) Dr.T.V.Rao MD 
A TOPIC FOR POST GRADUATE DISCUSSION
The greater strengths of Bacteriology lies with optimal isolation of the clinical pathogens and differentiating the Normal flora from infectious pathogens, The goals of testing are to detect possible drug resistance in common pathogens and to assure susceptibility to drugs of choice for particular infections We have several methods to perform and evaluate Antibiotic sensitivity patterns in our isolates as DILUTION METHODS, E-TEST, AUTOMATED ANTIMICROBIAL SUSCEPTIBILITY TESTING SYSTEMS MECHANISM-SPECIFIC TESTS, GENOTYPIC METHODS however we all do the DISK DIFFUSION METHOD as economical, technically not demanding method to evaluate our isolates Because of convenience, efficiency and cost, the disk diffusion method is probably the most widely used method for determining antimicrobial resistance in private and public Institutions .
 A growth medium, usually Mueller-Hinton agar, is first evenly seeded throughout the plate with the isolate of interest that has been diluted at a standard concentration (approximately 1 to 2 x 108 colony forming units per ml). It can be created with Barium sulphate using the basic dilution of the compound to obtain the desire concentration and compared with growth of isolate. Today we all use commercially prepared disks, each of which are pre-impregnated with a standard concentration of a antibiotic, are then evenly dispensed and lightly pressed onto the agar surface. The test antibiotic immediately begins to diffuse outward from the disks, creating a gradient of antibiotic concentration in the agar such that the highest concentration is found close to the disk with decreasing concentrations further away from the disk. After an overnight incubation, the bacterial growth around each disc is observed. If the test isolate is susceptible to a antibiotic, a clear area of “no growth” will be observed around that particular disk.
 The zone around an antibiotic disk that has no growth is referred to as the zone of inhibition since this approximates the minimum antibiotic concentration sufficient to prevent growth of the test isolate. This zone is then measured in mm and compared to a standard interpretation chart used to categorize the isolate as susceptible, or resistant. The word of Intermediate resistance is deceptive to take scientific decisions many do not report the isolate is INTERMEDIATE SENSITIIVTY, However MIC measurement cannot be determined from this qualitative test, which simply classifies the isolate as susceptible, resistant. Presence of zone of inhibition is not automatically interpreted as susceptibility to the antibiotic; the zone width must be measured and compared against a reference standard which contains measurement ranges and their equivalent qualitative categories susceptible or resistant. The advantages of the disk method are the test simplicity that does not require any special equipment, the provision of categorical results easily interpreted by all clinicians, and flexibility in selection of disks for testing. The disadvantages of the disk test are the lack of mechanization or automation of the test. The zones of growth inhibition around each of the antibiotic disks are measured to the nearest millimeter. The diameter of the zone is related to the susceptibility of the isolate and to the diffusion rate of the drug through the agar medium. The zone diameters of each drug are interpreted using the criteria published by the Clinical and Laboratory Standards Institute (CLSI, formerly the National Committee for Clinical Laboratory Standards or NCCLS)
It is important that the tables used for susceptibility test interpretations represent the most current criteria. Indeed, the CLSI documents are reviewed and updated frequently, usually once per year. Use of old or outdated information from the original editions of FDA-approved drug labels or older CLSI tables could represent a serious shortcoming in the reporting of patients' results.
The greater concerns to many Seniors and Technically knowledged Microbiologists- many of our laboratories in particular COMMERCIALLY RUNNING PRIVATE LABORATOREIS do not invest in isolation of true pathogen and randomly process all possible BACTERIAL ISOLATES, above all the ignorant clinicians want Sensitivity to all bacteria to be tested for all Available Antibiotics in store IT IS CERTAINLYY KILLING THE SPIRIT AND SCIENTIFIC ROLE MICROBIOLOGY STANDARDS AND IN FUTURE KILL MANY PATIETNS AND CAUSE OF SPREAD OF ANTIBIOTIC RESISTANCE SPREADING ALL OVER THE CLINICAL LOCATIONS . I WISH THERE SHOULD BE MORE EDUCATION TO MICROBIOLOGISTS AND CLINICANS HOW TO DEAL AND IDENTIFY STANDARDS IN MICROBIOLOGY AND PROCESS TO INITIATE ANTIBIOTIC POLOCY NEVER FORGET MANY ANTIBIOTICS CAN BE SENSITVE IN VITRO (IN THE LABOTORY 0 AND NEVER WORK IN VIVO THAT IS HUMAN BODY.
I WISH THE CLINICIANS SHOULD THINK ONCE BEFORE BELEIVING EVERY REPORT ARISING OF MANY VAGARIES AND SHOULD THINK TWICE BEFORE PRESCRING A ANTIBIOTIC.
I wish many young Microbiologists to learn from following well written articles
 1 Antimicrobial Susceptibility Testing: A Review of General Principles and Contemporary Practices. Barth Reller, Section Editor and Melvin Weinstein, Section Editor – Clinical infectious diseases Clin Infect Dis. (2009) 49 (11): 1749-1755.
Dr.T.V.Rao MD Professor of Microbiology Freelance writer

A TOPIC FOR POST GRADUATE DISCUSSION
The greater strengths of Bacteriology lies with optimal isolation of the clinical pathogens and differentiating the Normal flora from infectious pathogens, The goals of testing are to detect possible drug resistance in common pathogens and to assure susceptibility to drugs of choice for particular infections We have several methods to perform and evaluate Antibiotic sensitivity patterns in our isolates as DILUTION METHODS, E-TEST, AUTOMATED ANTIMICROBIAL SUSCEPTIBILITY TESTING SYSTEMS MECHANISM-SPECIFIC TESTS, GENOTYPIC METHODS however we all do the DISK DIFFUSION METHOD as economical, technically not demanding method to evaluate our isolates Because of convenience, efficiency and cost, the disk diffusion method is probably the most widely used method for determining antimicrobial resistance in private and public Institutions .
 A growth medium, usually Mueller-Hinton agar, is first evenly seeded throughout the plate with the isolate of interest that has been diluted at a standard concentration (approximately 1 to 2 x 108 colony forming units per ml). It can be created with Barium sulphate using the basic dilution of the compound to obtain the desire concentration and compared with growth of isolate. Today we all use commercially prepared disks, each of which are pre-impregnated with a standard concentration of a antibiotic, are then evenly dispensed and lightly pressed onto the agar surface. The test antibiotic immediately begins to diffuse outward from the disks, creating a gradient of antibiotic concentration in the agar such that the highest concentration is found close to the disk with decreasing concentrations further away from the disk. After an overnight incubation, the bacterial growth around each disc is observed. If the test isolate is susceptible to a antibiotic, a clear area of “no growth” will be observed around that particular disk.
 The zone around an antibiotic disk that has no growth is referred to as the zone of inhibition since this approximates the minimum antibiotic concentration sufficient to prevent growth of the test isolate. This zone is then measured in mm and compared to a standard interpretation chart used to categorize the isolate as susceptible, or resistant. The word of Intermediate resistance is deceptive to take scientific decisions many do not report the isolate is INTERMEDIATE SENSITIIVTY, However MIC measurement cannot be determined from this qualitative test, which simply classifies the isolate as susceptible, resistant. Presence of zone of inhibition is not automatically interpreted as susceptibility to the antibiotic; the zone width must be measured and compared against a reference standard which contains measurement ranges and their equivalent qualitative categories susceptible or resistant. The advantages of the disk method are the test simplicity that does not require any special equipment, the provision of categorical results easily interpreted by all clinicians, and flexibility in selection of disks for testing. The disadvantages of the disk test are the lack of mechanization or automation of the test. The zones of growth inhibition around each of the antibiotic disks are measured to the nearest millimeter. The diameter of the zone is related to the susceptibility of the isolate and to the diffusion rate of the drug through the agar medium. The zone diameters of each drug are interpreted using the criteria published by the Clinical and Laboratory Standards Institute (CLSI, formerly the National Committee for Clinical Laboratory Standards or NCCLS)
It is important that the tables used for susceptibility test interpretations represent the most current criteria. Indeed, the CLSI documents are reviewed and updated frequently, usually once per year. Use of old or outdated information from the original editions of FDA-approved drug labels or older CLSI tables could represent a serious shortcoming in the reporting of patients' results.
The greater concerns to many Seniors and Technically knowledged Microbiologists- many of our laboratories in particular COMMERCIALLY RUNNING PRIVATE LABORATOREIS do not invest in isolation of true pathogen and randomly process all possible BACTERIAL ISOLATES, above all the ignorant clinicians want Sensitivity to all bacteria to be tested for all Available Antibiotics in store IT IS CERTAINLYY KILLING THE SPIRIT AND SCIENTIFIC ROLE MICROBIOLOGY STANDARDS AND IN FUTURE KILL MANY PATIETNS AND CAUSE OF SPREAD OF ANTIBIOTIC RESISTANCE SPREADING ALL OVER THE CLINICAL LOCATIONS . I WISH THERE SHOULD BE MORE EDUCATION TO MICROBIOLOGISTS AND CLINICANS HOW TO DEAL AND IDENTIFY STANDARDS IN MICROBIOLOGY AND PROCESS TO INITIATE ANTIBIOTIC POLOCY NEVER FORGET MANY ANTIBIOTICS CAN BE SENSITVE IN VITRO (IN THE LABOTORY 0 AND NEVER WORK IN VIVO THAT IS HUMAN BODY.
I WISH THE CLINICIANS SHOULD THINK ONCE BEFORE BELEIVING EVERY REPORT ARISING OF MANY VAGARIES AND SHOULD THINK TWICE BEFORE PRESCRING A ANTIBIOTIC.
I wish many young Microbiologists to learn from following well written articles
 1 Antimicrobial Susceptibility Testing: A Review of General Principles and Contemporary Practices. Barth Reller, Section Editor and Melvin Weinstein, Section Editor – Clinical infectious diseases Clin Infect Dis. (2009) 49 (11): 1749-1755.
Dr.T.V.Rao MD Professor of Microbiology Freelance writer