الفهرس 
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Abstract
The main objective of this work was to study the prevalence of gram negative, gram-positive, candida and H. pylori in gastritis patients that may lead to failure of eradication therapy.
Resultsobtainedinthis studycanbe summarizedasfollows: -
1- Fifty-six stool and blood samples were collected from patients suffering from gastritis, and fifteen samples from patients without any symptoms of gastritis (as a control group), in Zagazig City, Sharkia Governorate.
2- Out of 56 samples 15 H. pylori isolates were obtained, identified by morphological characters and biochemical reactions. These isolates were recovered as follow: (26.7%) positive H. pylori, 5 isolates (8.9%) were recovered on Columbia blood agar supplemented with 5-7% deliberated sheep blood, 2(3.5 %) isolates on Brain heart infusion agar supplemented with 5-7% deliberated sheep blood, 1(1.7%) isolates recovered on chocolate agar media, and 7(12.5%) isolates recovered on Egg yolk emulsion with Columbia agar base. while in the control group all samples gave negative culture.
3- Gram negative bacteria were identified by morphological characters on MacConkey agar. Pink color colonies (lactose fermenters) and yellow colonies (none lactose fermenters) were picked and subjected to biochemical identification. Out of 56 samples, 54 (96.4%) were positive for Gram negative bacteria by biochemical tests The infected patients had multiple coexistence of H. pylori and species of Gram negative bacteria 54( 96.4%),11(19.6%) patients had H.pylori and E. coli , 9(16.6) patients had H. pylori and Enterobacter earogens, 2(3.7%) patients had H. pylori and Enterobacter cloacae, 3(5.5%), patients had H. pylori and K. pneumonia, 1(1.8%) patient had H. pylori and K. oxytoca, and 10(17.8%) patients had H. pylori andProteus mirabilis, 4(7.1%), patients had H. pylori and Proteus vulgaris ,6(10.7%) patients had H. pylori and Providenciarettgeri, 2(3.7 %) patients had H. pylori and Serratia liquefaciens, 5(9.2%) patients had H. pylori and Citrobacter freundii, 1(1.8%) patient had H. pylori and Citrobacter diversus, isolates were identified. While in control group samples negative in H. pylori were tested for the presence of gram negative, the results revealed that out of 9 samples 4(44.4%) E.coli,2(22.2%) K. pneumonia, and 3(33.3%) Proteus mirabilis
4- Gram positive bacteria were identified by morphological characters on Manitol agar. Yellow colonies (coagulase positive) and white colonies (coagulase negative) were picked and subjected to biochemical identification. Biochemical tests of the isolates revealed that out of 56 stool samples, 10 (35.7%) patients have H. pylori and S. aureus isolates and 8(28.5%) patients have H.pylori and S. epidermidis ,4(14.5%) patients have H. pylori and S. xylosus ,5(17.8%) patients have H. pylori and S. intermedius 1(3.6%) patient have H. pylori and S. saprophyticus were identified. While in control group only 2 samples were positive for gram positive bacteria 1(11.1%) S. intermedius,and 1(11.1%) S.saprophyticus
5- Yeasts were identified on sabouraud, s dextrose agar. colonies (were picked and subjected to biochemical identification, Out of 56 patient , 9 (30.9%) patients have H. pylori and C.albicans,3 (10.3%) patients have H. pylori and C. parapsilosis,2(6.8%) patients have H. pylori and C. krusei, 4(13.7%) patients have H. pylori and C. tropicalis, 2(6.8 %) patients have H. pylori and C.glabrata ,7(24.1 %) patients have H. pylori and Saccharomyces cerevisiae,1(3.4 %) patients have H. pylori and Rhodotorula glutinis,1(3.4 %) patients have H. pylori and Rhodotorula mucilaginosa (rubra) by biochemical tests . While in control group only 3 samples gave positive yeast culture all were Saccharomyces cerevisiae (30.0)%
6- By direct microscopic examination for the stool samples from gastritis patient it was revealed that there are11(19.6%)patient have H. pylori and E.histolyica trophozoit, 8 (14.2%) patients have H. pylori and giardia lambelia cyst, 4 (7.1%)patients have H. pylori and Strongyloides stercoralis, and 33 (58.9%) patientshave H.pyloriandNo parasites, While in control group only 3(33.3%) have E.histolytica while 6 patients have no parasites.
7- After the isolation and identification procedures we notice that gastric microbes were increased in the presence of H.pylori (gastritis patients) than in the absence of H.pylori(non-gastritis patients)
8- Stool samples from 56 gastritis patients and 15 non-gastritis patients were cultured to calculate CFU revealed that there were abundance in microbial growth in case of gastritis patient than in non-gastritis patients.
9- After isolation and identification of isolates from samples of gastritis patients and non-gastritis patients we found that there was an increase in the number of species isolated from gastritis patients than in non-gastritis patients, as the results revealed( 8) gram negative species in case of gastritis but only (3)species from non-gastritis patients.
10- H. pylori diagnosis with stool antigen rapid test is very sensitive and revealed that all patients suffering from gastritis symptoms were positive in stool antigen rapid test 56 of 56 (100%),While in control group only 6 from15 show positive antigen for H. pylori.
11- H. pylori diagnosis with serum antibody rapid test is sensitive and revealed that all patients suffering from gastritis symptoms were positive in serum antibody rapid test 56 of 56 (100%), While in control group only 6 from15 show positive antibody for H. pylori(the same sample that gave positive antigen).
12- Fresh blood samples were tested for H. pylori line, cagA was positive in 16 isolates (29.6%), vacA was positive in 16 isolates (29.6%), ureA was positive in 28 isolates (51.9%), P25 was positive in 50 isolates (92.5%), P30 was positive in 46 isolates (85.1%) and P19 was positive in 21 isolates (38.8%).While in control samples with positive antibody were tested for H. pylori line and the result showed that no sample had virulent gene neither cagA nor vacA but cytotoxins were present.
13- Stool samples with confirmly positive antigen and positive virulent gene in H. Pylori line serological test were subjected to amplification of ureA gene, 5 from 7 samples were positive for ure A .using (reference strain of code no. ATCC 43504)
14- RT-PCR with ureA primers detected 5 of 7 (71.4%). CagA mRNA was detected by RT-PCR in 5 of 5 PCR of genomic DNA for the presence of the cagA gene in the corresponding bacterial isolates correlated absolutely with cagA gene expression in fecal samples. VacA s1 mRNA was detected by RT-PCR in 2 of 5 and vacA s2 mRNA was detected by RT-PCR in 2 of 5 and dupA mRNA was detected by RT-PCR in 3 of 5 , vacAm1 mRNA was detected by RT-PCR in 2 of 5 vacAm2 mRNA was detected by RT-PCR in 2 of 5 as shown in figures (2,3,4,5&6).
15- H. pylori isolates were tested for their antibiotic susceptibility to 4 antibiotics by the disc diffusion method. According to the zone diameter break points, the most common resistance of isolates was observed against ciprofloxacin (30.6%) and metronidazole(23.2%) . While, sensitive isolates were observed againsr clarithromycin (82.6%) and levofloxacin (78.7%).
16- Gram negative isolates were tested for their antibiotic susceptibility to 13 antibiotics by the disc diffusion method. According to the zone diameter break points, the most common resistance of gram negative isolates was observed against cefixim (100%) and Trimethoprime (100%). While, high level of sensitivity was observed to imipenem (100%) and Meropener (100%).
17- Staphylococcus spp. were tested for their antibiotic susceptibility to 10 antibiotics by the disc diffusion method, According to the zone diameter break points, , the high level of sensitivity was observed to Imipenem
18- Yeast isolates were tested for their antifungal susceptibility to three antifungal agents by the disc diffusion method . According to the zone diameter break points, all yeast species show high sensitivity to all antifungal.
9- Conclusion
1- H.pylori is considered the major cause of gastritis.
2- Virulent genes were found only in 28.5% this may explain why there were avarity in symptoms.
3- Gastricmicrobiome show agreat diversity and more abundance in case of gastritis and show aresistance to different types of antibiotics, this may be areason for failure of eradication therapy.
4- Candida speacies show abundant growth in patients with multiple severs symptoms.
5- We conclude that the failure of eradication duo tomultiple reasons e.g the type of H.pylori strain, type of microbes coexist with H.pylori and its susibtability to different antibiotics, the hosts themselves, their history of taking antibiotics and their immunity.