الفهرس 
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Abstract
In this study, bacteriological examination of 400 different samples. Meat, offals, fish and luncheon which represents RTE foods, 100 sample from each. The examination was done according to ISO 11290 amendment 2004. The bacteriological examination revealed that: 2 isolates of L. monocytogenes were recovered from 100 meat samples, 2 isolates of L. monocytogenes were recovered from 100 offals samples, 1 isolate of L. monocytogenes was recovered from 50 pangasius fish samples and 1 isolate of L. monocytogenes was recovered from 50 shrimp samples.
Confirmation was done by Gram stain and biochemical tests including catalase, oxidase, sugar fermentation test, haemolysis on blood agar.
Serotyping for further confirmation of L. monocytogenes isolates by using Difco™ Listeria O Antiserum Type 1 and Difco™ Listeria O Antiserum Type 4. Serological examination revealed that: 2 isolates from meat: Listeriolysin o type 1, 2 isolates from offals: Listeriolysin o type 4, 1 isolate from pangasius fish: Listeriolysin o type 1, 1 isolate from shrimp: Listeriolysin o type 4.
Antimicrobial Susceptibility testing of L. monocytogenes was done for nine most commonly used antibiotics in veterinary and human therapy, using the standard disk diffusion method on Mueller Hinton Agar (MHA), following the procedures recommended by the Clinical and Laboratory Standards Institute. All L. monocytogenes isolates were resistant to Ceftriaxone, Ampicillin and Tetracycline. Only one isolate was resistant to Erythromycin. All L. monocytogenes isolates were susceptible to Amoxicillin-clavulanate, Ciprofloxacin, Rifampin, Vancomycin and Streptomycin. Five isolates were susceptible to Erythromycin.
Biofilm binding activity of L. monocytogenes was examined by using tissue culture plate assay and the results were as follow: 3 isolates showed strong binding activity, 2 isolates showed moderate binding activity and 1 isolate showed weak/non biofilm producer.
Genotypic characterization of L. monocytogenes isolates were done by using different techniques as:
Conventional PCR for detection of class-1 Integrons, Unfortunately, PCR couldn`t detect class-1 integrons in all 6 L. monocytogenes isolates.
Conventional PCR for detection of hlyA virulence gene, all six isolates of L. monocytogenes were examined for the presence of hlyA virulence gene. four isolates only could be detected positive for hlyA gene by conventional PCR, which were isolated from meat and fish. on the other hand, isolates which were isolated from offals were negative for hlyA gene by using conventional PCR.
Sequencing of L. monocytogenes hlyA gene isolated from meat and fish was detected by Applied Biosystems® 3500 Genetic Analyzers, then multiple sequence alignment analysis by using Clustal2.1 software then Identity percentage between L. monocytogenes (hlyA gene) sequence and other related sequences from Gene Bank database (NCBI) and Sequence identity matrix between different isolation sources (Meat and Fish) of L. monocytogenes (hlyA) created by Clustal2.1.
Confirmation of Listeria monocytogenes isolates by using Real time PCR (Sybr-green dye): The study revealed that all the six isolates are molecular identified and confirmed to be L. monocytogenes responsible for food poisoning using Real time PCR primers and Sybr-green dye that will amplify the gene hlyA (Listeriolysin) in L. monocytogenes.
The aim of this study is to determine the differences concerning phenotypic and genotypic characters of L. monocytogenes isolated from different sources (meat, offals, fish and RTE foods which is represented by luncheon) on the basis of pathogenicity and Antibiotic resistance to determine the difference between the isolates from different sources.
To achieve this aim, the following were carried out:
A-Phenotypic characterization B-Genotypic characterization
1-Isolation of L. monocytogenes.
2-Confirmation of L. monocytogenes by Biochemical tests.
3-Serotyping of L. monocytogenes.
4-Antimicrobial susceptibility testing for L. monocytogenes.
5-Detection of Biofilm binding activity. 1-Detection of class1 integron by conventional PCR.
2-Confirmation of pathogenicity by detection of the presence of hlyA gene.
3-Sequencing of the resulted PCR product.
4-Confirmation of results was done by SYBR green Real-time PCR.