There is currently a shortage in the literature on the antibiotic susceptibilities of fish pathogens. In this review I cover a paper whose authors compare three different methods in the hope of examining and evaluating antibacterial susceptibility; (i) the broth microdilution method (ii) the Etest and (iii) the disk-diffusion method
Francisella noatunensis subsp. orientalis and F. noatunensis subsp. noatunensis are small pathogenic bacteria with capabilities of causing piscine francisellosis (an acute to chronic disease) across a wide geographical range of different fish species. 10 bacterial isolates were obtained from two different fish species from four geographic regions between 2006 and 2010 (Escherichia coli ATCC 25922 was used as a control). To begin, the genetic homogeneity of all the isolates and the control were tested via PCR mediated genomic fingerprinting. Electrophoretic profiles established great levels of homogeneity between all 10 isolates and a significant difference from the E. coli ATCC 25922 control. It was concluded the isolates from different fish and different regions shared the same genotype.
When Soto et al. evaluated the use of the broth microdilution method, they found all the minimum inhibitory concentrations for substances (which reference values at 28°C are available) fell within the range given by the Clinical and Laboratory Standard Institute (CLSI), indicating this method is suitable way to gage the antimicrobial susceptibility of the of the two above mentioned bacteria and indeed any fastidious organism.
When the authors evaluated the use of the Etest, they first appraised Cystine Heart Agar supplemented with bovine haemoglobin (CHAH) as a potential media and found that it was indeed suitable. The MICs were very similar to the broth microdilution method, and although there isn’t currently any publication for testing bacteria isolated from aquatic animals, the Etest appears to be a suitable method for determining antibiotic susceptibility. It is pointed out, however, that further exploration surrounding this method is necessary.
As with the Etest, when it came to evaluating the use of the disk-diffusion method, CHAH was again considered as a potential media. Clear zones of inhibition were observed for the quality control in the CHAH for numerous antibiotics, but when these zone diameters were compared to those observed on Mueller–Hinton agar supplemented with 5% sheep blood (MHB) and the ranges given by the CLSI, only the zones from a few of the tested antibiotics fell within the ranges. Although not what was expected, the results from the disk-fusion method were consistent and repeatable and together with the low MICs for these antibiotics it is reasonable to conclude the bacterial isolates are intermediately/highly susceptible to these few antibiotics (florfenicol, tetracyclin, nitrofurantoin, gentamicin and erythromycin).
The paper concludes by giving a long list of antibiotics the bacterial isolates are susceptible too (18 in total), and also a long list of antibiotics that they are resistant too (26 in total).
This paper provides a good baseline of data for future research monitoring the development of antibiotic resistance among the bacterial isolates, as well as being a starting point for the development of potential therapeutics to bacterial diseases. The article was well structured and not overly complex to read and understand. Although I didn’t feel like I’d learnt much after reading it, I don’t feel that was the aim of the authors, the aim was to provide a very good and essential platform for further research.
Francisella noatunensis subsp. orientalis and F. noatunensis subsp. noatunensis are small pathogenic bacteria with capabilities of causing piscine francisellosis (an acute to chronic disease) across a wide geographical range of different fish species. 10 bacterial isolates were obtained from two different fish species from four geographic regions between 2006 and 2010 (Escherichia coli ATCC 25922 was used as a control). To begin, the genetic homogeneity of all the isolates and the control were tested via PCR mediated genomic fingerprinting. Electrophoretic profiles established great levels of homogeneity between all 10 isolates and a significant difference from the E. coli ATCC 25922 control. It was concluded the isolates from different fish and different regions shared the same genotype.
When Soto et al. evaluated the use of the broth microdilution method, they found all the minimum inhibitory concentrations for substances (which reference values at 28°C are available) fell within the range given by the Clinical and Laboratory Standard Institute (CLSI), indicating this method is suitable way to gage the antimicrobial susceptibility of the of the two above mentioned bacteria and indeed any fastidious organism.
When the authors evaluated the use of the Etest, they first appraised Cystine Heart Agar supplemented with bovine haemoglobin (CHAH) as a potential media and found that it was indeed suitable. The MICs were very similar to the broth microdilution method, and although there isn’t currently any publication for testing bacteria isolated from aquatic animals, the Etest appears to be a suitable method for determining antibiotic susceptibility. It is pointed out, however, that further exploration surrounding this method is necessary.
As with the Etest, when it came to evaluating the use of the disk-diffusion method, CHAH was again considered as a potential media. Clear zones of inhibition were observed for the quality control in the CHAH for numerous antibiotics, but when these zone diameters were compared to those observed on Mueller–Hinton agar supplemented with 5% sheep blood (MHB) and the ranges given by the CLSI, only the zones from a few of the tested antibiotics fell within the ranges. Although not what was expected, the results from the disk-fusion method were consistent and repeatable and together with the low MICs for these antibiotics it is reasonable to conclude the bacterial isolates are intermediately/highly susceptible to these few antibiotics (florfenicol, tetracyclin, nitrofurantoin, gentamicin and erythromycin).
The paper concludes by giving a long list of antibiotics the bacterial isolates are susceptible too (18 in total), and also a long list of antibiotics that they are resistant too (26 in total).
This paper provides a good baseline of data for future research monitoring the development of antibiotic resistance among the bacterial isolates, as well as being a starting point for the development of potential therapeutics to bacterial diseases. The article was well structured and not overly complex to read and understand. Although I didn’t feel like I’d learnt much after reading it, I don’t feel that was the aim of the authors, the aim was to provide a very good and essential platform for further research.
A review of:
Soto, E., Griffin, M., Judy, W. and Hawke, J. P. (2012) Genetic analysis and antimicrobial susceptibility of Francisella noatunensis subsp. orientalis (syn. F. asiatica) isolates from fish. Veterinary Microbiology 154 407-412
Soto, E., Griffin, M., Judy, W. and Hawke, J. P. (2012) Genetic analysis and antimicrobial susceptibility of Francisella noatunensis subsp. orientalis (syn. F. asiatica) isolates from fish. Veterinary Microbiology 154 407-412
2 comments:
Hi Rachel,how the final report going?
I was just wondering how the broth microdilution method works? I'm guessing (as the name suggests) it involves growing the culture in broth rather than agar but how do they test it with antibiotics? Is it basicly the practical we did on antibiotics last year?
Hi Lee,
Everything is going well thanks, what about for you? Can you also tell me please, if you know, how to set up alerts when I receive comments or responses? As I seemed to miss some last term and I only happened to stumble upon this one. I’ve looked in setting but can’t seem to find it.
A quick response – Yes, similar to what we did last year, but plates with premade wells already contain antibiotics were used and the broth added before incubation.
In the broth microdilution method the MICs of 39 different antimicrobials to the isolates and control were tested using two different plate formats; (i) GN2F – sensititre gram negative plate format and (ii) AVIAN1F – avian one isolate MIC plate using the manufactures protocol.
To begin with the isolates and control were plated on CHAH agar (Cystine Heart Agar supplemented with bovine haemoglobin – as mentioned above for the Etest), for 96h or 24h respectively at 28˚. The inocula were prepared by suspending the colonies in 1L phosphate buffer saline, then diluted 100-fold (isolates) or 100-fold (control) in Mueller-Hinton II cation adjusted broth supplemented with 2% IsoVitaleX and 0.1% glucose. 50 µl was added to each well of the plates containing the different antimicrobials. These plates were incubated for 48 h (isolate) or 24 h (control) at 28˚C. Bacterial growth was visually checked at this time. The MICs were defined as the lowest concentration exhibiting no visible growth.
I hope this answers your question, if not, let me know.
Rachel
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