الفهرس 
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Abstract
S.aureus is a leading human pathogen and remains a major cause of infections
worldwide. Antibiotic resistance is a growing problem because of its effect on the rapid
spread of threatening diseases and infections and the inability to control them.
The antibiotic resistance in S.aureus began to be isolated and identified limiting the
use of several drugs over the years. The emergence of resistance to penicillin (MRSA)
followed by Vancomycin (glycopeptides) has turned the therapy of staphylococcal
infections into a global challenge. Unfortunately, S.aureus isolates with reduced
susceptibility to vancomycin (VRSA, VISA and hVISA) has been detected in clinical
S.aureus strains throughout the world and has been associated with vancomycin treatment
failure for S.aureus.
The present study was a trial to detect S.aureus with reduced susceptibility to
vancomycin in different types of clinical samples.and their genomic characterizations.
The study was carried out on 250 S.aureus isolates from different types of clinical
samples collected from patients admitted to various departments in the Alexandria
University Hospitals, Egypt from May 2014 to April 2015.
All S.aureus isolates were subjected to the following:
1- Phenotypic identification based on colony morphology and standard biochemical
tests as S.aureus and genotypic identification by the presence of nuc gene using
PCR.
2- Antibiotic susceptibility by disk diffusion method on Mueller-Hinton agar based
on CLSI guidelines.
3- Determination of MIC of vancomycin by broth microdilution method as standard
and by agar diltution method according to CLSI guidelines for detection of VISA
and VRSA.
4- Molecular detection of Van gene in S.aureus isolates and in the reference strain
Entercococcus faecalis (ATCC 51299) as positive control.
5- S.aureus isolates with reduced vancomycin susceptibility (VISA or hVISA) was
confirmed by the PAP-AUC approach using Mu3 (ATCC 700698) S.aureus
reference strain and by BHIA-V2, BHIA-V3 and BHIA-V4 agar screening.
6- Molecular typing of VISA and hVISA strains was done by the identification of
the agr specificity groups genes ( agrI, agrII, agrIII and agrIV) using multiplex
PCR.
The results of the present study revealed:-
1- The 250 S. aureus isolates were collected from different types of clinical
samples; 173(69.2%) from pus, followed by 26(10.4%) from sputum, 24(9.6%)
blood, 12(4.8%) from nasal swabs, C.V.C and urine (each 4(1.6%) and 7
isolates (2.8%) from body fluid (pleural fluid, ascitic and synovial fluid).
2- PCR amplification of the nuc gene (nuc gene positive) in the clinically isolated
strains yielded a 270 bp amplicon in 216(86.4%) out of the 250 S. aureus
isolates included in the study.
3- The antibiotic sensitivity of 250 S.aureus isolates to different classes of
antibiotics (12 antibiotics) revealed that.the most active agents by susceptibility
were consistently linezolid (100%), vancomycin (94.8%) and teicoplanin
(84.4%). On the other hand the isolates exhibited high level of resistance to
Oxacillin (72%), Cefoxitin (69.2%), and moderate level of resistance to
carbapenems (32.4%), aminoglycosides (32.2%), macrolides (38%),
levofloxacin (40.4%) and clindamycin (36%).
4- The prevalence of MRSA among the different types of clinical samples was
69.2% with a high prevalence among pus (70.8%) and blood samples (70 %).
5- Regarding susceptibility to clindamycin out of 250 S.aureus isolates 90 (36%)
were found to be resistant to clindamycin,of them 8% iMLSB and 28% cMLSB.
The rate of MLSB phenotypes among MRSA (10.4%, 38.2%) were higher in
comparison to MSSA (2.5%, &5.2%).
6- Using the broth microdilution method as a gold standard for screening of
S.aureus resistance to vancomycin, the sensitivities and specificities of the disc
diffusion and agar dilution methods found to be (13.6%, 95.6%) and (86.4% ,
100%) respectively.
7- No VRSA could be detected among the 250 S.aureus isolates using the broth
microdilution method and all the 250 S.aureus isolates were negative for vanA
gene.
8- Meanwhile 22 (8.8%) S.aureus isolates with reduced susceptibility to
vancomycin [3 (1.2%) VISA and 19 (7.6%) hVISA] was found, all of them
were MRSA.
9- Out of the 22 S.aureus isolates (VISA and hVISA) 19 (86.4) isolates were from
pyogenic infection (3 VISA+16/19 hVISA).
10- The dominant agr group among the 22 VISA and hVISA S.aureus isolates was
agr group I (45.5%) followed by agr group III (36.3%) then agr group II
(9.1%) . Two isolates (9.1%) were non-typable and none of the isolate was agr
group IV.
11- The agr specific group I had the highest rate of detection (45.5%) among the
22 S.aureus isolates with reduced sensitivity to vancomycin all of them were
MRSA isolated mostly from pyogenic infections.
In Conclusion
1- S.aureus isolates with reduced susceptibility to vancomycin were detected as
both VISA and hVISA (1.2% and 7.6%) respectively. On the other hand, no
VRSA could be detected.
2- All S.aureus isolates with reduced susceptibilityto vancomycin were MRSA.
3- Most of the VISA and hVISA (86.4%) were isolated from pyogenic infection
most of them were agr group I.
4- Agar dilution (BHIA-V2, BHIA-V3 and BHI-V4) method is a reliable method of
screening for VISA and hVISA.
5- Linezolid is the best alternative therapy for S.aureus isolates with reduced
susceptibilityto vancomycin.
6- A high prevalence (69.2%) of MRSA among the different types of clinical
samples with a high prevalence among pus and blood samples about 70% each.
7- D-test for detection of inducible resistance to clindamycin is a surrogate marker
for resistance to streptogramin B; a drug approved to be effective in treatment of
S.aureus isolates with reduced susceptibility to vancomycin
8- Low sensitivity (13.6%) of the disc diffusion method to detect S.aureus wth
reduced sensitivity to vancomycin (VISA &hVISA).