Av Paragallinarum Case Study

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Materials and Methods
Overview
The whole study consists of four phases. To start with, the suitable media need to be identified for the growth of Av. paragallinarum. Secondly, the media used in disc diffusion and broth dilution testing need to be validated. This validation process is completed by performing disc diffusion and broth dilution testing of quality control strain on candidate agar or broth. Then, the antimicrobial sensitivity of Av. paragallinarum can be tested on validated media, by both disc diffusion and dilution method. Any antibiotic resistance shown in the tests will be recorded and analysed. Finally, PCR will be performed to identify any genetical reason underneath the resistance. This report only focus on growth media selection,
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The methods and interpretative ranges were based on CLSI handbook. Three agars were used in method validation, chocolate Mueller-Hinton agar (cMH), cation-adjusted Mueller-Hinton agar (CAMH) and BA/SN agar. cMH is the media for fastidious bacteria testing in CLSI handbook, which was prepared with 3.8% Mueller-Hinton agar (Oxoid), 1% BBLTM freeze-dried bovine haemoglobin (BD) and 1% BBLTM IsoVitaleXTM enrichment (BD) as nutrition supplement. CAMH agar was made with 3.8% BBLTM Mueller-Hinton II agar (cation adjusted, BD), supplemented with 0.0025% of NADH and 1% of heat inactivated chicken serum. The formula of this media is identical to the CAMHB, other than the agarose component. BA/SN agar was prepared with BBLTM Blood Agar Base (BD), with identical supplement formula as TMB …show more content…
For each repeat, the suspension was swabbed on a blood agar plate for sterility control. It is critical to use correct reading method in measuring inhibition diameter, as suggested by CLSI handbook. For some plates, multiple rings appeared, the reading of inhibition zone was based on the inner ring, which should be readily distinguished with unaided eyes (CLSI, 2013). The results of inhibition diameter was compared with QC ranges for each antibiotics provided in CLSI standards (2015). A media can be validated only if all tested antibiotics gave inhibition ranges that fall into the QC

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