Vol. 38, May, 2014.

1/38 EPIDEMIOLOGIC STUDY ON THE PREVALENCE OF OCCULT HEPATITIS B VIRUS AMONG CANCER AND NON-CANCER HCV AMONGEGYPTIAN PEDIATRIC PATIENTS

Heba E. Raouf, Aymen S. Yassin*, Amira H. Soliman**, Mohamed T. Mansour***, SalwaA. Megahed*and Mohamed S. Ashour

Department of Microbiology and Immunology, Faculty of Pharmacy, Modern Sciences and Arts University, 6th of October City, Giza, Egypt.

*Department of Microbiology and Immunology, Faculty of Pharmacy, Cairo University, Cairo, Egypt.

**Department of Clinical Pathology, National Cancer Institute, Cairo University, Cairo, Egypt.

***Virology and Immunology Unit, Cancer Biology Department, National Cancer Institute, Cairo University, Cairo, Egypt.

Occult hepatitis B infection (OBI) is characterized by the presence of hepatitis B virus (HBV) DNA in the serum in the absence of hepatitis B surface antigen (HBs-Ag). Locally reported incidence of hepatitis C virus (HCV) infections in Egypt is among the highest incidence in the world. This study is designed to assess the prevalence of OBI in two groups of Egyptian pediatric patients (HCV cancer patients and HCV non-cancer patients), who gave negative result for the presence of HBs-Ag.All patients were subjected to clinical, biochemical and virological assessment. HBVparameters (HBV DNA, anti-HBc, anti-HBs) were determined. Nested-PCR and real time PCR techniques were used for the detection of HBV-DNA,while real time PCR was used for detection of HCV-RNA. OBI was not observed in the HCV non-cancer pediatric patients but was found in 31% of HCV pediatric cancer patients. There arestatistical significant differences between cancer and non-cancer groups regarding the transaminases ALT, AST and total bilirubin levels, as they are higher among cancer group. Levels of HBc-Ab and HBs-Ab indicated no significant difference among the HCV cancer patients and non-HCV cancer patients. OBI in this study was found to be more frequent in patients with acute lymphocytic leukemia.In conclusion, these results showed that HBs-Ag negativity is not sufficient to completely exclude HBV-DNA presence.

2/38 MOLECULAR CHARACTERIZATION OF EXTENDED SPECTRUM b LACTAMASES (ESBLS) GENES IN PHENOTYPICALLY ESBLS PRODUCING ESCHERICHIA COLI AND KLEBSIELLA PNEUMONIAEISOLATES

Kawthar I. Mohamed, Mona A. Khattab*, Nevin M. Al-azhary** and

Ayat R. Abdallah***

Microbilogy and Immunology, Faculty of Medicine, Ain Shams University, Cairo, Egypt. Faculty of Medicine, Taibah University, Almadinah Almunawarah, KSA.

*Clinical Pathology Department, National Cancer Institute, Cairo University, Cairo, Egypt. Faculty of Medicine, Taibah University, Almadinah Almunawarah, KSA.

**Public Health and Community Medicine, National Liver Institute, Egypt.

The precise identification of the genes involved in extended spectrum β lactamases (ESBLs) –mediated resistance is necessary for surveillance; epidemiological studies of their transmission and to determine appropriate antibiotic policy measures. K. pneumoniae and E. coli are the most prevalent among ESBL-producing microorganisms. ESBLs are encoded by mobile genes; the most frequently encountered ESBLs genes belong to the blaCTX-M, blaSHV, and blaTEM families. The aim of the present study to determine the prevalence of ESBL producing K. pneumoniae and E. coli phenotypically and study the molecular characterization of blaCTX-M, blaSHV, and blaTEM genes among phenotypically ESBLs producers, using conventional PCR. Using phenotypic methods; out of 60 E. coli and K. pneumoniae isolates (22 and 38 respectively), 44 isolates (73%) were potential ESBLs (13 E. coli and 31 K. pneumoniae isolates) and showed multidrug resistant pattern especially to amoxicillin, ciprofloxacin and gentamicin. We found that 82% (36/44) of ESBLs producing E. coli and K. pneumoniae were positive for single gene and (34% and 47.7 %) were positive for two or more genes. BlaSHV gene was the commonest detected either alone or in combination (69.2%, 67.7% among E. coli and K. pneumoniae respectively), followed by blaTEM and blaCTX-M. blaSHV gene was detected alone among 38.4%, and 29% of E. coli and K. pneumoniae isolates respectively. Rrgarding E. coli isolates 30.8% of ESBL positive E. coli were found to harbor two genes (either blaCTX-M and blaSHV or blaCTX-M and blaTEM), and 15.4% were positive for the three genes. Regarding K. pneumoniae isolates 35.5% of ESBL positive K. pneumoniae were found to harbor two genes at same time (either blaCTX-M and blaSHV, blaCTX-M and blaTEM or blaTEM and blaSHV), and 12.9 % were positive for the three genes.In conclusion a high percent of ESBL was detected among K. pneumoniae and E. coli isolates and the presence of blaTEM or blaTEM and blaSHV was associated with ESBL phenotype. Our results indicate that routine ESBL detection should be made mandatory. We concluded that a genotypic detection of ESBLs genes is needed to identify these genes and study their epidemiology and transmission in hospitals. As well as phenotypic methods are only screening methods for detection of ESBLs in a routine laboratory. The genotypic methods help us to confirm the genes responsible for ESBL production. Furthermore, Irrational use of third generation cephalosporins must be discouraged to reduce multidrug resistant bacteria to increase patients. compliance and to make an antibiotic policy.

3/38 IN VITRO ANTIBACTERIAL ACTIVITY OF DRUGS, RIVANOL AND HONEY AGAINST BACTERIAL STRAINS ISOLATED FROM CUTANEOUS LEISHMANIASIS

Khaled Nasher Qhatan

Department of Biology Chemistry, Faculty of Education Radfan, Aden University, Yemen

Microbial pathogens have developed resistance to every antibiotic introduced into clinical practice. Most bacteria that cause infections are resistant to at least one antibiotic, and some are resistant to all commonly employed antibacterial drugs. The samples have been collected at the laboratory of the Faculty of Education Radfan, Aden University, Yemen. The Cutaneous Leishmaniasis (CL) was diagnosed clinically Parasitologically, hematologically, and culturing by the parasites. The microbes which are found around the lesions were diagnosed microbiologically. Moreover the susceptibility of the microbes for antibiotics and the new compound 2-ethoxy-6, 9-diamino acridiniumbenzen sulphohydrazidooxalylglycinate (RIV) has been studied. Besides, the effect of Yemeni Honey (Sider and Summer) has been studied. The ages of patients ranged between 12- 68 years and 17/32 of them (53.13%) were males and the other 15/32 (42.87%) were females. In total (91) bacterial isolates were isolated from 32/40 (80.00%) of patients. Staphylococcus aureus isolated from most of the CL lesions 63 (69.23%) and Escherichia coli 7 (53.85%). Results of antibiotic sensitivity test revealed that S. aureus and β-Hemolytic streptococci showed 27.27% and 36.36% sensitivity respectively. Gram negative isolates. Citrobacter freundii, E. coli and Klebsiella spp., were sensitive to all antibiotics except Imipenem, Streptomycin with 81.81% sensitivity. All Gram negative isolates were sensitive to Meropenem and Trimethoprim and resistant to Streptomycin. All Gram positive isolates were sensitive to Trimethoprim and resistant to Ciprofloxacin and Penicillin-G. The two kinds of Honey showed wide spectrum of antibacterial activity against different bacterial species including C. freundii, E. coli, Klebsiella spp., Pseudomonas aeruginosa, S. aureus and β-Hemolytic streptococci. Besides, it was revealed that the six isolates tested were sensitive to a 100% v/v solution of Yemeni (Sider and Summer) honey in both agar (well and disc) diffusion assay, but none showed sensitivity to a 25%, 50%, and rarely 75% solution of honey. The results obtained showed also that Acridine RIV is somewhat active like honey and it is more active when is mixed with honey. Besides, Acridine RIV is active against five strains of bacteria namely C. freundii, E. coli, P. aeruginosa, S. aureus and β-Hemolytic streptococci. Our results show that in microbiology and clinical tests, honey offers advantages in controlling the bacteria growth and in the treatment of certain Health problems. The topical dressing of CL ulcers with honey is very effective and helps in the treatment in a short term as compared with RIV treatment. However, honey and RIV are more effective together than if they are used alone. The important step of treatment for CL is local care along with anti-leishmania therapy.

4/38 IMPLANTED MEDICAL DEVICES AND THEIR IMPACT AS A RESERVOIR FOR RECURRENT MICROBIAL INFECTION

Mohamed A. Mohamed, Amr Hanora* and Marwa Rezk

Microbiology Dept. Faculty of Science Suez Canal University and * Microbiology and Immunology Dept. Faculty of Pharmacy, Suez Canal University

Modern medical biotechnology leads to more and more biomedical devices use and consequently rise in device related infection. The present study highlight the role of these devices in recurrent microbial infection. One hundred ninety one samples constituting 14 types of commonly used medical devices were collected from patients residing at Suez Canal and Suez Hospitals .Collected sample were cultured on nutrient agar plates for bacteria and Sabaurauds dextrose agar plates for yeasts.The growing microbial colonies were recorded, purified and identified using the conventional methods. Isolates were characterized by randomly amplified polymorphic DNA (RAPD-PCR) for further molecular characteristics definition. PCR reactions were conducted using 5 arbitrary 10-mer   primers. Antibiotic susceptibilitytests were performed using 15 different antibiotics for bacterial isolates (Escherichia coli, Staphylococcus epidermidis, Proteus mirabilis and Pseudomonas spp.) using the disc agardiffusion method.          


5/38 CENTRAL VENOUS CATHETERS; A RISK FACTOR FOR BACTEREMIA IN NEONATAL INTENSIVE CARE UNIT (NICU); ALMAZAH HOSPITAL FOR NEONATAL CARE; CAIRO; EGYPT

Ibrahim M. Al-Hosiny and *Ihab I. Sror

Microbiology and Immunology and *Pediatrics Departments, Faculty of Medicine, Al–Azhar University.

Central venous catheters (CVCs) are widely used in neonatal intensive care units (NICUs), unfortunately, catheter-related bacteremia is a frequent life-threatening complication. The current study was conducted to detect the incidence of bacteremia among newborns admitted in NICU at Almazh Neonatal Hospital, to study some risk factors influencing it including the insertion of CVCs, to identify the causative organisms, and its biogram. During the period from January 2013 to October 2013,120 newborns with central venous lines were selected from Neonatal Intensive Care Unit (NICU), 48 male and 72 female, of age ranging from 2 to 35 days, with periods of admission > 48 hours duration (group: 1). A control group including 120 newborns without CVCs, matched with group: 1 for gender, gestational age, birth weight, and periods of admission was also included in the study(group:2). All newborns included in the study developed signs of sepsis. Complete blood count with positive neutrophil indices (Immature/Total neutrophils >0.2) or thrombocytopenia and elevated C-reactive protein were positive in all patients. Blood samples (2.0 to 3.0 ml) were obtained from peripheral venous blood for blood culture. Any growth was fully identified. Determination of antimicrobial susceptibility for each isolate was done by means of the disc diffusion method on Mueller Hinton agar. The incidence of bacteremia was 21.7% in neonates with CVCs (group: 1) compared to 10.0% in neonates without CVCs (group: 2). The incidence of bacteremia was much higher among neonates with gestational age < 32 weeks (31% in group: 1 and 12.0% in group: 2) compared with the rate of infection in those with gestational age >36 weeks (13% and 10.0%).Birth weight was inversely proportional to the duration of CVL insertion in NICU, where the highest percentage of infected patients was in Very Low Birth Weight (VLBW) group (32.5% in group:1 and 15.0% in group: 2), followed by Low Birth Weight (LBW) group (19.1% and 8.5%). The Normal Birth Weight (NBW) group shows the lowest percentage (12.1% and 6.0%).The rates of bacteremia were directly proportional to durations of CVCs insertion (12.1% in <1 week of insertion group, 19.1% in 1 – 2 weeks of insertion group, and 32.5% in>2weeks of insertion group) as shown in table (4). The organisms causing bacteremia isolated from patients with CVCs were S. epidermedis, S. aureus, E. coli, Candida albicans, Klebsiella pneumonia, and Proteus vulgaris with percentages of 42.3%, 30.7, 11.5%, 7.7%, 3.8%, and 3.8% respectively compared to those isolated from patients without CVCs including S. aureus, E. coli, S. epidermedis, and Strept. Pneumoniae with percentages of 50.0%, 25.0%, 16.6%, and 8.3% respectively. Antibiogram of the isolates showed that the isolated strains were sensitive to Imipenim, Amikacin, Sulbactam/Ampicillin, Vancomycin, Cefotaxime, Clindamycin, and Ceftriaxone with percentages of 94%, 86%, 72%, 72%, 69%, 53%, and 50% respectively.


6/38 CANDIDA ALBICANS AS AN IMPORTANT CAUSE OF ORAL HUMAN INFECTION AND ITS EFFECT ON SALIVA PROTEIN PROFILE.

M. Abd El-Razik, Noha ELsayed Frag* and Mayada Mahmoud

Microbiology Department, Faculty of Science, Suez Canal University and*Human Physiology Department, Faculty of Medicine, Suez Canal University

The role of Candida albicans in eliciting oral human infections between various patient groups and the various factors predisposing for such infections was studied. Oral swab samples were collected, from oral mucosa, dorsum of the tongue and mucosa below denture of 115 patients. Patients were uptaking broad spectrum antibiotics, suffering from diabetes mellitus, heavily smokers and complete or partial denture wearing and other various underlying diseases leading to inefficient immunity. Candida albicans was mostly isolated from various collected samples. A representative collected saliva samples were electrophoretically analyzed for its protein profile. The obtained results confirmed the harmful role of the colonizing Candida albicans in changing the saliva normal composition and accordingly its vital function.

7/38 PHENOTYPIC DETECTION OF MULTIDRUG RESISTANT PSEUDOMONAS AERUGINOSA AND ACINETOBACTER BAUMANNII ISOLATED FROM URINARY TRACT WARD IN SOME EGYPTIAN HOSPITALS

Abdel Rahman K. Raslan1, Mai M. Helmy1,3, Moselhy S. Mansy2, Mohamed S. Ashour1,2

Faculty of Pharmacy, October University for Modern Sciences and Arts1,

Microbiology & Immunology Department, Faculty of Pharmacy - Al-Azhar University, Cairo, Egypt2,

Microbiology & Immunology Department, Faculty of Medicine Zagazig University3

Pseudomonas aeruginosa (P. aeruginosa) and Acinetobacter baumannii (A. baumannii) are important nosocomial pathogens in health care settings. Treatment is complicated by multi-drug resistance [MDR]. This study was undertaken to analyze the resistance mechanisms of P. aeruginosa and A. baumannii urinary isolates in some Egyptian Hospitals using simple, easy and inexpensive screening phenotypic methods. Antibiotic susceptibility profile for 31/205 (out of 186 urinary samples) isolates of P. aeruginosa (24) and A. baumannii (7) was determined by standard disc diffusion method. Isolates showing reduced susceptibility to3rd Cephalosporins, azetronam were tested for ESBL producing by double disc synergy test (DDST). Isolates showing resistant to carbapenem were tested for carbapenemases and metallo-β-lactamases producing using modified Hodge (MHT) and EDTA disc synergy tests (EDS) respectively, while ciprofloxacin resistant isolates were confirmed to be efflux pump producers by determining minimal inhibitory concentration of ciprofloxacin with and without efflux pump inhibitor (EPI) omeprazole (100 mg/L) by microtitre broth dilution method. Moreover biofilm formation ability of MDR isolates was determined by quantitative microtiter plate method. As regard P. aeruginosa, high level of antimicrobial resistance was noticed with ampicillin (100%), tetracycline, doxycycline (91.7%) and slightly low resistant level with imipenem and pipercillin/tazobactm (45.8% and 50% respectively). On the other hand A. baumannii isolates were resistant to ampicillin (100%), pipercillin, tetracycline (85.7%), doxycycline, cephalosporins, azetronam (71.4%), while pipercillin/ tazobactm, meropenem, imipenem, amikacin and ciprofloxacin were equally the most effective antibiotics (85.7%). Among 25 suspected ESBL isolates, 85% (17/20) of P. aeruginosa and 100% (5/5) A. baumannii were actually ESBL producer by DDST. Among 14 carbapenem resistant isolates, 38.5% (5/13) of P. aeruginosa were confirmed as carbapenemase by MHT and 84.6% (11/13) were metallo-β-lactamase by EDS. On the other hand the only A. baumannii carbabenem resistant isolate (1/1) were confirmed by both MHT and EDS. Among 12 MDR isolates (11/24 P. aeruginosa and 1/7 A. baumannii), omeprazole was able to potentiate the effect of ciprofloxacin in 41.7% (5/12) of isolates. Moreover 50% of MDR isolates (P. aeruginosa & one A. baumannii) were strong biofilm formation that potentiates the resistance mechanisms. MDR P. aeruginosa and A. baumannii are significant threat in hospitalized patients. It should be addressed with infection control measures, and new strategies needed for treatment and prevention of MDR.

8/38 RELATIONSHIP BETWEEN VIRULENCE FACTORS AND ANTIFUNGAL RESISTANCE OF DIFFERENT CANDIDA SPECIES IN CANCER AND NON CANCER PATIENTS

Soheir S. Maklad, Omnia A. El-Dydamoni,Laila H. Saleh and Jaylan A. Aly

Microbiology & Immunology Department, Faculty of Medicine for Girls, Al-Azhar University, Cairo, Egypt.

The relationship between virulence factors and antifungal resistance of different Candida species was investigated in 276 adult patients with different types of cancer (group I) and 50 non cancer patients (group II) who have clinical manifestations of Candida infections. Candida isolates were identified in clinical samples by the standard procedures. Antifungal susceptibility was evaluated by the Integral Yeast Plus System contains 11 antifungal agents. Germ tube (GT), secretory aspartic proteinase (SAP), phospholipase (PL), and biofilm formation (BF) were assessed by the standard procedures. The isolation rate of Candida spp. in cancer patients was significantly higher (79.3%) than non-cancer patients (20.7%). C. albicans was the most common spp. among both groups (44.8%) and (52%) respectively followed by C. tropicalis (25%); C. glabrata (21.9%) and C. krusei (8.3%) in group I and followed by C. krusei (28%), C. glabrata (12%) and C. tropicalis (8%); in group II. Most of Candida spp. showed high rates of resistance to econazole, clotrimazole, miconazole, itroconazole and fluconazole 16 ug. Significant higher frequencies (P<0.001), (P<0.05) of all virulence factors (GT, SAP, PL, BF) were detected in Candida isolates of group I (75.8%), (77.5%), (79.2%) and (79%) respectively than group II; (24.2%), (22.5%), (20.8%) and (21%) respectively and in systemic (58.9%) than mucocutaneous infections (41.1%). There were significant associations of resistance to some antifungals and virulence factors (P<0.001 and P<0.05). Conclusion: Although, C. albicans is the most frequent and virulent isolates in cancer patients yet non-albicans candida which are less virulent have emerged as important opportunistic pathogens. Significant associations of virulence factors and antifungal resistance may explain the increased rate of morbidity and mortality particularly in cancer patients.


9/38 A COMPARATIVE STUDY OF BACTERICIDAL AND POSTANTIBIOTIC EFFECTS OF MACROLIDES AND KETOLIDES AGAINST CLINICAL BACTERIAL ISOLATES COLLECTED FROM ALEXANDRIA, EGYPT

Mostafa A. El Nakeeb, Nadia M. El-Guink, Hamida M. Abou Shleib

and Nihal K. Moussa

Department of Pharmaceutical Microbiology, Faculty of Pharmacy,

Alexandria University, Egypt.

The present work is a comparative microbiological study to emphasize the main differences between macrolides: azithromycin (AZM), clarithromycin (CLR), erythromycin (E), roxithromycin (ROX), spiramicin (SP) and ketolides represented in telithromycin (T). A total of eighty three bacterial standards and isolates of different identities and susceptibilities with variable resistance patterns were used in this study. Among the different macrolides subjected to susceptibility testing, cross resistance was complete between the different members; T was not a subject for cross-resistance as it showed higher activity against Staphylococcus spp. with zero resistance. Minimum inhibitory concentrations of macrolides and T indicated relatively higher activity against Gram-positive organisms (MIC range: 0.0035->500µg/ml) than against Gram-negative ones (MIC range: 0.01->250 µg/ml). The highest activity was detected with AZM and T in Gram-negative and Gram-positive bacteria, respectively, while Pseudomonas aeruginosa isolates were resistant. Macrolides and T manifested considerable bactericidal activity against both Gram-positive and Gram-negative organisms, this effect was found to rely on different factors; among which are drug concentration and bacterial susceptibility. T showed a bactericidal activity that exceeded that of the macrolides in some tested strains.The postantibiotic effect (PAE) of macrolides and T was determined spectrophotometrically. PAE values ranged between 0 and 11.64 hr for all the tested macrolides and T after one hr exposure to 4 x MIC. Higher PAE values were observed against Gram-positive organisms than Gram-negative ones. The highest PAEs were obtained with Enterococcus faecium isolate (e101) using SP and ROX (11.64 and 10.88 hr, respectively), and that obtained with alphahemolytic Streptococcus viridans isolate (α sp) with all macrolides and T(>10.56 hr). No PAE was obtained with ROX against Staphylococcus epidermidis isolate (Se21). T showed the highest PAE against 40% of the strains. The effect of cefoperazone, ciprofloxacin, or trimethoprim on the antibacterial activity of macrolides and T was also determined turbidimetrically using the clinical isolates Escherichia coli (E8) and Staphylococcus epidermidis (Se3) as they showed high degree of resistance to all the tested macrolides. The effect of combination was insignificant. In conclusion, due to the considerable bactericidal activity obtained in this work, and contrary to their known spectrum of activity, macrolides and ketolides should be introduced in the treatment of infections caused by fastidious Gram-negative organisms. This is highly dependent on the concentration used and the susceptibility of the isolates. Also, whenever clinically possible, the high PAE of macrolides and ketolides should be considered in adjusting the time intervals of the doses to achieve the highest patient compliance.

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