Vol. 8, May, 2004

viagra livraison express en france 1/8 PURIFICATION AND SOME PROPERTIES OF AN EXTRACELLULAR 1,4- cialis et femme b viagra wirkung wie schnell -ENDOGLUCANASE FROM TRICHODERMA LONGIBRACHIATUM

R.R. Saad and E.M. Fawzi

Department of Biological Sciences, Faculty of Education, Heliopolis, Cairo, Egypt

An extracellular endoglucanase (1,4-b-glucanohydrolase, EC 3.2.1.4) produced by kamagra oral jelly ou acheter Trichoderma longibrachiatum has been purified to homogeneity by ammonium sulfate (50%) precipitation, followed by gel filtration and ion-exchange chromatography on DEAE-cellulose. The enzyme was purified 9.96 folds with 9.72% yield and exhibited specific activity of 23.5 U mg-1 protein. Mr of the enzyme determined by SDS-PAGE was 45. The pH and temperature optima for the enzyme were pH 5.0 and 45°C, respectively. The Km towards CMC was 0.45 mg ml-1. The enzyme activity was enhanced by Mn2+, Fe2+ and Co2+ and inhibited by K+, Na+, Ca2+, Mg2+ and Hg2+.

kamagra super activ 2/8 ISOLATION, CHARACTERIZATION AND PHYSIOLOGICAL STUDIES ON CUCUMBER MOSAIC VIRUS ISOLATE FROM CUCUMBER GREENHOUSE PLANTS IN EGYPT.

E.T. Sayed; R.M. Taha* and R.M. Abd El-Hamid**

Botany Dep. Fac. Science , Cairo Univ., Cairo , Egypt.

*Botany Dep. Fac. Science ,Cairo Univ., El-Fayoum Branch , El-Fayoum ,Egypt.

**Botany Dep. Fac. Science , Helwan Univ., Cairo , Egypt.

An isolate of cucumber mosaic virus (CMV) was isolated from greenhouse cucumber plants. DAS-ELISA was applied for first virus identification .The virus was mechanically and aphid transmitted by a non persistent manner. A wide range of hosts were found to be readily infected by this isolate. Physical properties (thermal inactivation point, dilution end point and longevity ou trouver viagra a paris in vitro) were similar to that of CMV. Inclusion bodies in the form of amorphous, crystalline and round-shape were found in the cytoplasm of the virus-infected leaves. The virus causes cytopathological alternations in cell organelles including chloroplast deformations, mitochondrial aggregation, destruction of the nucleus as well as cytoplasmic vacuolation. Infected leaves showed increase in enzyme activity of polyphenol oxidase, peroxidase and amylase as well as increase in RNA, ammonia and soluble carbohydrate content. SDS-PAGE showed a similarity between polypeptides of CMV-infected leaves in relation to symptoms development. The polypeptide content accompanying the three symptoms (net mosaic, mild mosaic and malformation) may change quantitatively and qualitatively.                    

3/8 BIODEGRADATION OF CYCLOSULFAMURON BY THE FUNGUS ASPERGILLUS FLAVUS

H.M.S. Khalifa

Plant Protection Department, Faculty of Agriculture, Al-Azhar, University, Cairo

Biodegradation of the herbicide cyclosulfamuron by the fungus viagra 1 comprimé Aspergillus flavus Link, was determined in complete and in nitrogen or carbon-free Dox, s medium. Also, biodegradation was measured by using bioassay technique, whereas the herbicidal activity of cyclosulfamuron was assessed in peat moss either containing the fungus or not, and millet as indicator plant. The results indicated that the fungus could grow in complete medium containing cyclosulfamuron up to 300 µg/ml. achat rapide cialis Aspergillus flavus grew slightly in carbon-free medium, interestingly; it grew well in carbon-free medium containing different concentrations of the herbicide, 15 days after incubation at 25oC. Increment of the fungal growth in carbon-free medium increased as the concentration of the herbicide increased up to 50 µg/ml. However, increasing the concentration of the herbicide resulted in less increment of the fungal growth. On the other hand, the fungus did not grow in nitrogen-free medium containing different concentrations of the herbicide. This result showed that achat viagra par paypal A. flavus biodegrades cyclosulfamuron as a source of carbon as indicated by its growth in carbon-free medium containing the herbicide. The bioassay technique showed that cyclosulfamuron greatly reduced the growth of millet in peat moss, especially the root system. This effect was reduced by adding cialis 20 mg generique A. flavus to peat mass containing the herbicide, and this was more pronounced with increasing the incubation period up to four weeks .

4/8 VAGINAL MICROBIOLOGICAL STUDIES ON WOMEN USING DIFFERENT METHODS OF CONTRACEPTION

*R.H. El-Yamani and **M.A.B. Gamal.

* Obst. & Gynaec.Dep. Faculty of Medicine and **Microbiology Dept Faculty of Pharmacy, El-Margeb Univ., the Great Socialist People’s Libyan Arab Jamahiriya

Cervical microbial flora has been investigated in five comparable age groups, 20 women each. The first group using oral contraceptive pills (Microvolar), The second group using vaginal contraceptive pills (Anovolar) and the third group using injectable contraceptive (Depoprovera). All those contraceptives contained steroidal drugs. The fourth group using inserted intrauterine contraceptive device (Copper-T 380 A, IUCD). The control group included non contracepting women of comparable age (group V). Four endocervical swabs were collected from each women during the tenth through twelfth day of the second menstrual cycle of using contracertive method. The swabs used for culture and identification of aerobes, anaerobes, candida and mycoplasma. Lactobacilli were the dominant organisms in all groups, but more frequently higher in women using steroidal contraception than those using IUCD and the noncontracepting control women. Candida albicans was also more frequently isolated in all groups studied relative to the control group. The other isolated microorganisms showed no significant differences between those groups using steroidal contraceptives. However there was increased incidence of isolation of some bacterial isolates from women using IUCD than that of the control group. Concerning the detection of Ureaplasma urealyticum in all groups studied, was more or less the same and there was no significant differences between all groups except for those using IUCD, where higher detection rate of Ureaplasma urealyticum was illustrated. The use of IUCD as contraceptive method increase the likelihood of vaginal infection relative to the use of steroidal contraceptives.


5/8 DIAGNOSTIC APPROACH TO EXTRA PULMONARY TUBERCULOSIS

E.A.El-Sherbini, A. Ghazal and G. Fadaly*

Microbiology Department, * Pathology Department, Medical Research Institute, Alexandria University.

The paucibacillary nature of extra pulmonary tuberculosis makes its diagnosis not easy. In this work, nested PCR using IS6110 as a target and pathozyme TB complex plus kit that utilizes M. tuberculosis highly specific 16 KDa and 38KDa antigens were studied for the rapid detection of M. tuberculosis in extra pulmonary tuberculosis specimens. A total of 60 specimens (30 clinical specimens and 30 blood samples) obtained from 30 patients suspected clinically to have extra pulmonary tuberculosis were examined by microscopy, PCR and ELISA. Ten percent of the samples were positive by microscopy and ELISA, while 60% were PCR positive and cytopathological examination of these cases was in favor of tuberculosis. PCR gave rapid positive results in 55.6% of microscopy negative cases and 71.4% of cases presented with lymph node swelling could be diagnosed by PCR compared to 10% only when ZN or ELISA test was considered.

6/8 PRESERVATION OF MINIMALLY PROCESSED FRESH JEW’S MALLOW BY GAMMA IRRADIATION

S.M.A. Taha and H.M.H. Swailam

Microbiology Department, National Center for Radiation Research and Technology, P. O. Box 29 Naser City, Cairo, Egypt.

The possibility to extend the shelf-life of minimally processed fresh Jew’s mallow “Moulokeia” by gamma irradiation was studied. Changes that took place in microbial populations, chemical and physical properties as a result of irradiation and cold storage of Jew’s mallow were also investigated. Application of gamma irradiation greatly reduced the microbial load of Jew’s mallow (total bacterial counts and molds) and the reduction was proportional to irradiation dose. Gamma irradiation was very effective in inactivatingcoliforms, E. coli and other pathogenic bacteria such as Aeromonas hydrophila, Listeria monocytogenes, Staphylococcus aureus and Salmonella species. The results showed that Aspergillus species was the most predominant among the others genera isolated from irradiated and unirradiated Jew`s mallow at zero time and during storage. No clear differences were found between unirradiated and irradiated samples with different doses (up to 4.0 kGy) of moisture content, fiber, ash, acidity, pH and vitamin c. A clear increase was observed in chlorophylls and carotenoids in irradiated samples. The consistency index of Jew’s mallow samples decreased with increasing irradiation dose. Meanwhile, the flow index increased with increasing irradiation dose. The results clearly indicated that samples of Jew’s mallow irradiated with dose 4.0 kGy were organoleptically acceptable until 35 days of storage at 5 ºC.      


7/8 EFFECT OF GAMMA RADIATION ON THE MICROBIAL CONTAMINATION AND SOME BIOCHEMICAL PROPERTIES OF POLLEN GRAINS

A.S. Bashandy and S.M.A. Taha

Microbiology Department, National Center for Radiation Research and Technology,

P. O. Box 29, Naser City, Cairo, Egypt.

Pollen grains are used as health food ingredients, but may be subjected to microbial contamination as a result of extraction and handling. So, the present study was to evaluate effects of gamma radiation on the microbial contamination and some biochemical properties such as amino acid composition and mineral contents. Pollen grains have high levels of total aerobic bacteria, spore-forming bacteria, total molds and yeasts. The results showed that most samples were of unsatisfactory microbiological quality in the view of the coliforms and E. coli levels. Aeromonas hydrophila and Salmonella species were detected in 40% of pollen grains samples, whereas Clostridium species, Staphylococcus aureus and Streptococcus faecalis were detected in 50, 60 and 60 %, respectively. Listeria monocytogenes was not detected in any of the pollen grains samples. The present study showed that gamma irradiation of pollen grains at 5-10 kGy reduced total aerobic bacteria, spore-forming bacteria, fungi, coliforms and most pathogenic bacteria were below detection levels. Also, the results showed that glutamic acid (39.2 mg/g protein) was the most prevalent amino acid followed by aspartic acid (37.6 mg /g), then leucine (19.2) and serine (17.6 mg /g protein). The exposure of pollen grains to gamma radiation at 8 and 10 kGy caused variable increase in the levels of all amino acids except for methionine. Data of mineral content (mg /g) of pollen grains showed that the predominant mineral elements were potassium, manganese and sodium. However, the results also showed that mineral content was not markedly changed by gamma irradiation.

8/8 INFLUENCE OF GAMMA IRRADIATION ON THE MICROBIAL QUALITY AND ANTIBACTERIAL ACTIVITY OF SOME EGYPTIAN HONEY TYPES

S.M.A. Taha

Microbiology Department, National Center for Radiation Research and Technology (NCRRT), AEA, P. O. Box 29, Naser City, Cairo, Egypt

Different types of honey (clover, sweet marjoram and black cumin) were evaluated for their natural microbiological quality. They were also examined for the presence of coliforms, E. coli, Aeromonas hydrophila, Listeria monocytogenes, Staphylococcus aureus, Salmonella species and Streptococcus faecalis. Most tested honey types had an average total aerobic plate counts (TAPC) and spore-forming bacteria (SFB) values ranging from 1.2x102 to 7.0x103 and 1.2x102 to 6.8x102 cfu/g, respectively and the total mold and yeasts ranged from 7.0x10 to 2.0x103 cfu/g. Coliforms and E. coli and pathogenic bacteria were not found in all honey samples. The results showed that spore-forming bacteria was more resistant to gamma irradiation. Also, the irradiation dose of 5 kGy reduced most mesophilic counts to well bellow the maximum value legally allowed. Effect of gamma irradiation on antibacterial activity ofdifferent types of honeywas investigated in this study. The results showed that Streptococcus faecalis was the most resistant to different concentrations of tested honey followed with Listeria monocytogenes, Proteus species and Aeromonas hydrophila while Staphylococcus aureus was the most sensitive. Data also exhibited that black cumin honey was the most effective one while both sweet marjoram and clover honey were almost the same. Also, the results showed that there was no marked loss in the antibacterial activity of all tested honey before and after exposure to 1, 3, 5 and 25 kGy gamma irradiation.

 

9/8 DEFENCE AMENDMENTS OF CARROT PLANTS AGAINST POWDERY MILDEW DISEASE BY FERN EXTRACT AND AUXIN TREATMENT

E.A. Ewais and A.A. Farrag

Botany and Microbiology Department, Faculty of Science, Al-Azhar University, Cairo, Egypt.

Field experiments were conducted to investigate the role of fern (Adiantum capillus veneris) and auxin (Indole acetic acid) treatment for improving defence mechanism of carrot (Daucus carota L.) plants against powdery mildew disease. Aqueous extract of Adiantum and auxin treatments obviously reduced the infection rate upon carrot plants naturally infected down to (23 & 27%) for susceptible variety and (13 & 20%) for resistant variety. Interestingly all the growth parameters and yield of the treated plants improved more than untreated plants (control). Mass spectrum was consulted to confirm the resistance mechanism to fungal disease in both the resistant and susceptible varieties of carrot plants. This response was related to the activities of both peroxidase and catalase enzymes. Finally the results suggested the use of these treatments as strategy for controlling the outbreak of carrot powdery mildew disease.

10/8 THE INCIDENCE OF TOXOPLASMA GONDII AMONG CHILDREN WITH CHRONIC TONSILLITIS.

M.A. Moawad and A.T. Moawad

National Center for Radiation Research and Technology,Atomic Energy Authority, Cairo, Egypt.

Toxoplasmosis is a very common disease of worldwide distribution. It is caused by Toxoplasma gondii, which is an obligate intracellular protozoan. The study aims to identify the role of T. gondii as a cause of chronic tonsillitis. Forty children were included in this study, divided into two groups, a study group of 20 children presenting with non-suppurative chronically enlarged tonsils and a control group of 20 healthy children of matched age and sex. Children were examined clinically and parasitologically. Complete blood picture (CBP) and erythrocyte sedimentation rate (ESR) were done. Sera of all cases were screened for T. gondii IgG antibodies by indirect haemagglutnation test (IHT). Histopathological examination of available tonsils removed during tonsillectomy was performed. The results revealed 50% and 5% seropositivity for Toxoplasma IgG antibodies in the two groups respectively. In the study group, there was a high statistically significant increase in the red blood cells count and white blood cell count. A significant difference was in the mean value of Hb between the study group and control group. A biopsy was taken which proved the histopathological picture of Toxoplasma infection. The results showed that T. gondii infection can be considered in the differential diagnosis of chronic tonsillitis especially when WBC count and ESR are within normal range.

11/8 ENZYME INHIBITORS AND ANTIBIOTICS AS BIO-MODULATORS OF BACTERIAL PATHOGENS

B.M. Haroun, M.S. Ammar, H.M. El-Berry and S.T. Hamad

Department of Microbiology, Faculty of Science, Al-Azhar University,

Nasr City, Cairo, Egypt.

The study investigated the effect of different combinations of antibiotics on the prostatic borne bacterial pathoens (PBBP) in chronic bacterial prostatitis. Application of certain enzyme inhibitors has been also studied. The most effective antibiotics were Ofloxacin followed by Rifampicin and Togamycin, whereas Cd2+ followed by Zn2+and Cu2+ exhibited considerable growth inhibition through their enzyme inhibitng activities. Phosphate, Sn+2 and EDTA had no effect on the tested PBBP.

12/8 BACTERIOLOGICAL EVALUATION OF CHRONIC PROSTATITIS

H.M. El-Berry; B.M. Haroun; M.S. Ammar and S.T. Hamad

Department of Microbiology, Faculty of Science, Al-Azhar University,

Nasr City, Cairo, Egypt.

The current study focuses on assessment of the bacteriological causes of chronic prostatitis. Specimens of expressed prostatic secretions from twenty cases of bacterial prostatitis were obtained over six months. Bacteria were isolated on a single antibiotic-containing media, a total of 13 antibiotics were applied in the study as prescribed by the referring physicians. A total of 155 bacterial isolates were identified, 100 (64.5%) were Gram positive, whereas, 55 (35.5%) were Gram negative. The identified bacterial isolates were belonging to the genera Bacillus 31.6%, Staphylococcus 30.3%, Hemophilus 17.4%, Pseudomonas 8.5%, Neisseria 5.1%, Citrobacter 2.6%, Hafni 1.3%, Bacteroids 1.3%, Stomatococcus 1.3%, Proteus, Pasteurella, Listeria, Brochothrix and Micrococcus were 0.65% each.

13/8 SCREENING FOR GALACTOSIDASES, GLUCOSIDASES AND INVERTASE PRODUCED BY ISOLATED FUNGAL STRAINS

S.A. Ismail and A.F. Sahab*

Department of Chemistry of Natural and Microbial Products, National Research Centre, Dokki, Cairo, Egypt

*Department of Plant Pathology, National Research Centre, Dokki, Cairo, Egypt

Nineteen fungal strains isolated from molasses were primary identified, and tested for their ability to produce invertase, α- and β-galactosidases and α- and β-glucosidases, in three different culture media. A beet and cane - molasses were used individually in culture medium. The cultures were tested at two different periods in rotatory shaker (180 rpm) at 30o. All the tested microorganisms possessed intracellular and / or extracellular enzymes in different distinct values. The results indicated that the type of the medium as well as the incubation period have strong influence on the production of all tested enzymes produced by the different fungal strains.

14/8 MICROBIAL QUALITY OF OREOCHROMIS NILOTICUS (BOLTI) AND WATER OF RIVER NILE AND EL-EBRAHEMYAH CANAL AT ASSUIT CITY

U. M. Abdul-Raouf,

Botany and Microbiology Department. Faculty of Science, Al-Azhar University, Assuit 71524, Assuit, Egypt.

Associated microflora of Oreochromis niloticus (Bolti) and, water of River Nile and El-El-Ebrahemyah canal at Assuit City, Egypt, in which the fish were caught and determined. Standard methods were used to determine aerobic, anaerobic and Enterobacteriaceae counts. The presence of selected foodborne pathogens in fish and water samples were also investigated. For all counts, no significant differences were observed in water samples and the intestinal contents of El-Ebrahemyah canal fish. The fish from El-Ebrahemyah canal are the most contaminated (6.33x107), while, River Nile fish specimens (Gills) yielded the lowest numbers (2.7x103). Motile and non-motile aerobic and anaerobic gram-negative bacteria together with Bacillaceae and Enterobacteriaceae are the most predominant flora of water and fish samples. One strain of Vibrio cholerae was isolated from El-Ebrahemyah canal water. Two strains of Listeria monocytogenes were isolated from the   water and fish intestine of El-Ebrahemyah canal. One strain of Escherichia coli O157: H7 was isolated from El-Ebrahemyah canal intestine fish. In conclusion, the fish and water samples of El-Ebrahemyah canal, which analyzed in this study, were of unacceptable quality and safety.

15/8 PURIFICATION OF CHITINASE AND ENDO-b-1,3-GLUCANASE FROM TRICHODERMA HARZIANUM

M.S. El-Katatny

Botany Department, Faculty of Science, El-Minia University 61519, Egypt

Four isolates of Trichoderma spp. were tested for the production of chitinase and b-1,3-glucanase in presence of chitin or laminarin as a sole carbon source. Trichoderma harzianum was the most potent isolate for production of these enzymes. Addition of glucose (5, 10 g/l) repressed enzyme production measured in the presence of R. solani dried mycelium (5 g/l) as a carbon source. Chitinase and b-1,3-glucanase were purified from the culture filtrates of T. harzianum grown on chitin as a sole carbon source. The 43 kDa chitinase was purified by a single hydrophobic chromatography (HIC) step, which maintained its activity when tested on chitin SDS-PAGE. Purification of b-1,3-glucanase using either (HIC) or Ion-exchange chromatography (IEX) has led to partially purified protein. The partial purified b-1,3-glucanases obtained by the two purification techniques was studied by active-stained native PAGE and was about 74 kDa. Different concentrations of glucano-d-lactone did not inhibit b-1,3-glucanase activity. Therefore, b-1,3-glucanase has been classified as an endo-b-1,3-glucanohydrolase. The isoelectric focusing (IEF) for chitinase and b-1,3-glucanase was 4.0 and 6.2 respectively. The possible involvement of these enzymes in the antagonistic interaction between Trichoderma and phytopathogenic fungi is discussed.


16/8 THE STATUS OF FAS AND FAS LIGAND EXPRESSION ON PERIPHERAL MONONUCLEAR CELLS IN CHRONIC HEPATITIS C VIRUS INFECTION AND HEPATOCELLULAR CARCINOMA

M.M. Darwish, S.F. Hafez, M.A. Mohammed, NM. Elaish and A.M. El Zayadi*

Microbiology & Immunology Department, Tropical Medicine Department*,

Faculty of Medicine, Ain Shams University.

The immune response triggered by Hepatitis C virus (HCV) infection, its underlying mechanisms and its relation to disease progression and response to antiviral therapy are still poorly understood and are still to be clarified. The aim of the present study was to evaluate the possible role of Fas mediated apoptosis of peripheral blood mononuclear cells (PBMCs) in the persistence of HCV infection, development of chronic HCV infection, response to antiviral therapy and the development of hepatocellular carcinoma (HCC) through studying the status of expression of Fas and Fas ligand (FasL) on these cells. Also to find out the relation between the status of expression of Fas/ Fas L on PBMCs and histopathological changes in liver tissues and the possibility of utilizing this test as a noninvasive tool to monitor disease progression and predict disease outcome. The present study was carried on 55 chronic hepatitis C (CHC) patients; 20 patients complicated by HCC and 35 chronic hepatitis C (CHC) patients without HCC; 13 of them were under combined interferon and ribavarin treatment, 12 recently diagnosed patients and 10 patients with manifestations of portal hypertension. Fifteen apparently healthy subjects were enrolled in the study as a control group. The percentage of Fas and Fas-ligand expressing PBMCs was assayed by flowcytometry using whole blood technique.Histological activity index (grading) and staging were determined in liver biopsies taken from CHC patients receiving antiviral treatment and the recently diagnosed patients. The mean percentage of FasLexpressing PBMCs was significantly diminished when comparing patients with HCC to patients with CHC and the control group (P<0.001). Also patients with HCC showed a significant decrease in the mean percentage of Fasexpressing PBMCs in comparison to patients with CHC (P<0.05). No significant difference was detected between CHC patients or their different subgroups and the control group as regard mean percentage of either Fas or FasL expressing PBMCs. A significant positive correlation was found between Fas expressing PBMCs in CHC patients and the levels of ALT, AST and direct bilirubin (P<0.001). No significant correlationwas found between the mean percentage of Fas and Fas ligand expressing PBMCs and the histological grading (HAI) and staging of liver disease in the biopsies taken from CHC patients.These results suggest that there is induction of apoptosis of immune cells during HCV infection via the Fas signaling pathway, thus facilitating HCV persistence and with the decrease in FasL expression by PBMCs there is liability of development of HCC. Monitoring FasL expression on PBMCs might provide a reliable tool to predict the tendency to develop HCC and might open challenging therapeutic strategies in HCV infection through manipulation of Fas/FasL expression on hepatocytes and immune cells.


17/8 COMPARATIVE DISSOCIATION OF CAMPYLOBACTER JEJUNI ANTIBODIES BY TWO CHAOTROPIC AGENTS

H.M. El-Berry, T.F. Ismail, M.S. Ammar and M.O. Wasfy

Faculty of Science, Al-Azhar University

Antibody avidity and immune maturation are well known as important traits of the immune response to antigenic stimulants. Campylobacter jejuni infection occurs at a relatively high level and is the cause of acute illness in developed countries. Eighty-four (84) serum samples were collected from individuals suffering from diarrhea; in addition to twenty-two (22) sera from control subjects were used in this study. Enzyme-linked immunosorbent assay (ELISA) was employed to detect C. jejuni reactive antibodies in the serum samples. Eighty-one percent of the diarrheal population had no, or little, Campylobacter reactive antibodies as in the healthy controls. The remaining group of diarrheal patients (19%) showed significantly elevated titers for antibodies reactive with C. jejuni. The Complement Fixation (CF) test was performed as another specific, but less sensitive, technique to confirm the previous results.   Avidity of antibodies has been expressed as the concentration of chaotropic agent required to disrupt an antigen-antibody complex. Avidity of the C. jejuni reactive antibodies (IgA, IgM and IgG) in this study was relatively low, However, IgG seemed to be more avid than IgA and IgM. This study shows that the low incidence and mild symptoms of campylobacteriosis in Egypt could be a function of low-avidity antibodies.

18/8 BACTERIOLOGICAL STUDY ON NILE WATER AT CERTAIN LOCALITIES AT CAIRO

H.M. Mahdy and M.E. Zain

Botany and Microbilogy Department, Faculty of Science, Al-Azhar University, Nasr City, P.O. Box 11884, Cairo, Egypt.

The investigation concerning isolation of E. coli strains from the River Nile water in Cairo and testing their pathogenicity with other reference strains. Forty seven isolates isolated from 3 localities viz. Rod El-Farag, El-foastat and Helwan. Seven isolates caused diarrhoeal diseases in mice; six isolated from Helwan and one from El-Foastat. Lactobacilli prevented pathogenicity of E. coli. E. coli strains were multidrug resistant. The most common protein responsible for pathogenicity was of molecular weight 20, 12 and 6.5 Kda. Ca2+ recorded 88± 0.4 mg/L in Helwan, Na+ recorded 111 mg/L in Rodh El-Farag and El-Foastat and Cl- recorded 32±0.9 mg/L in Helwan.

19/8 DETECTION OF HETEROGENEOUS INTERMEDIATE VANCOMYCIN RESISTANCE(HETERO-VISA) AMONG METHICILLIN RESISTANT S. AUREUS (MRSA)BLOOD ISOLATES

N.S.Omar, W.K. Mowafy and M.S.Rizk

Microbiology & Immunology Dept., Faculty of Medicine, Mansoura University

Background: In May 1996, Hiramatsu et al. described the first clinical isolate of S. aureus with reduced susceptibility to vancomycin (hetero-VISA). Hetero-VISA refers to isolates for which the MIC of vancomycin for one or more subpopulations is >4 mg/L, whereas the overall MIC is <4 mg/L Recent reports from many countriessuggesting that hetero-VISA is responsible for the failure of vancomycin therapy, and a potential source of VISA subclones in patientsgiven vancomycin for a prolonged period. Aim Of The Work: Patients with MRSA bacteremia at Mansoura University Hospitals were investigated to determine the prevalence of the isolated strains of hetero-VISA ,to investigate their molecularepidemiology, and clinical significance. Also assay the effect of pre-incubation of the isolates with vancomycin on the expression of vancomycin resistance. Materials & Methods: MRSA bacteramic isolates were screened for reduced susceptibility to vancomycin and then confirmed hetero-VISA status by demonstrating MICs above the susceptible range (i.e., >4 mg/L).Testing the effect of pre-incubation with vancomycin on the expression of vancomycin resistance by the bacteria was done .Epidemiological typing by using pulsed-field gel electrophoresis (PFGE) was performed for the isolated hetero-VISA strains. Results: Among 82 S.aureus blood isolates, 20 MRSA (24%) were isolated , the prevalence of confirmed hetero -VISA strains among the 82 S. aureus isolates were 3.6%, or among MRSA were 15%. The examined hetero -VISA isolates were closely related genotypically as the DNA patterns were very similar. Conclusion &Recommendation: Attention to rational prescribing of antibiotics is required, avoiding broad-spectrum agents whenever possible. Vancomycin should be used only when necessary: when there is resistance to other agents, and only when infection is present. Colonisation is not an indication to use glycopeptides, nor is minor allergy to β-lactams. Clinicians and laboratories should consider hetero-VISA if a patient with a proven MRSA infection fails to respond to vancomycin. It is a challenge to establish a laboratory screening protocol for hetero-VISA and to determine what resources should be allocated to screening for hetero -VISA.

20/8 ENTEROTOXICITY, SLIME PRODUCTION AND GENOTYPING OF COAGULASE NEGATIVE STAPHYLOCOCCI ISOLATED FROM FOOD HANDLERS

W. Mowafy, S. Taher*, W. Badawy** and Bernedette Bernard***

*Deparments of Microbiology and Immunology, Faculty of Medicine, Mansoura University, **Microbiology and Immunology, Mansoura Student Hospital, ***Community Medicine, Faculty of Medicine, Mansoura University

Staphylococcal food poisoning (SFP) caused by enterotoxigenic staphylococci is one of the main food-born diseases. Coagulase negative staphylococci (CNS) were isolated from the hands of food handlers in Mansoura University Hospital (MUH) restaurants and studied for the production of enterotoxins, slime production, antibiotic resistance profile and genotyping. Out of 170 hand swabs, 56 CNS isolates were detected (32.9%). Seventeen (30.4%) of CNS isolates produced slime and 6 (10.6%) produced enterotoxins. Staph enterotoxins (SEA, SEB SEC and SED) were assayed by reversed passive latex agglutination test (RPLA). There was no relation between toxin and slime production with exception of one isolate which produced both slime and enterotoxin. As regard antibiotic resistance profile, there was insignificant increase of resistance to some antibiotics of CNS slime producing strains and also for CNS enterotoxin producing isolates than isolates negative for production of slime and enterotoxins. Genotyping of the isolates was performed by pulsed field gel electrophoresis (PFGE), we detected genetic relatedness for different enterotoxin producing isolates. On the other hand, isolates did not produce enterotoxins were unrelated genetically to enterotoxin producing strains. Isolation of enterotoxigenic CNS from hands of food handlers suggests that they may contribute to food poisoning if food is contaminated by them and held in conditions that allow their growth and elaboration of enterotoxins. So enterotoxigenic CNS should be considered in suspected cases of staphylococcal food poisoning.

21/8 METABOLIC STUDIES ON RESPONSES OF RESISTANT AND SUSCEPTIBLE SOYBEAN SEEDLINGS INFECTED WITH PHYTOPHTHORA MEGASPERMA ROT DISEASE

A.A. Farag and E.A. Ewais

Botany and Microbiology Department, Faculty of Science, Al-Azhar University, Cairo, Egypt.

The effect of Phytophthora megasperma as a pathogen on growth and metabolism of resistant and susceptible soybean seedling were studied in vitro. The disease incidence was severe in susceptible (var. Clark) than resistant (var. Giza 82) plants. The total soluble carbohydrate and total phenolics contents were higher in both infected than healthy plants. However, protein contents were higher in resistant infected than control plants. On the contrary, DNA and RNA as will as mitotic index were obviously decreased in infected plants. The least rot infection in soybean resistant tissues was generally accompanied with an obvious increase in endogenous contents of hormones auxin (IAA), abscisic acid (ABA) and gibberellic acid (GA3). The infection in soybean susceptible tissues was associated with a great decrease in IAA content, while GA3 and ABA contents were slightly and markedly increased, respectively. In addition, electrophoretic profiles of soybean proteins showed quantitative differences between major bands obtained in healthy and infected plants.

22/8 ISOLATION OF CERTAIN CLOSTRIDIUM SPECIES FROM DIFFERENT SOURCES AND PREPARATION OF TETANUS TOXOID FROM TOXIGENIC CLOSTRIDIUM TETANI

M.S. Ashour, M.H. Hosny*, W.N. El -Tayeb, F.M. Gomaa.

Microbiology Department, Faculty of Pharmacy, Alazhar University, and

* VACSERA, Cairo, Egypt.

Tetanus continues to be a threat to public health especially in developing countries. One million tetanus cases occur worldwide. Immunization is the only way to prevent this deadly disease. The aim of the present work was to isolate one or two species of the toxigenic Clostridium species as well as preparation of toxoid from such toxigenic species. In the present study, 51 samples including twenty three soil samples, twenty five horse faeces samples and three goat excreta were investegated for isolation of Clostridium species. A total of 65 microbial isolates of Clostridia were recovered. The recovered species were; C. sporogenes, C. bifermentans, C. sordellii, C. tetani, C. botulinum, C. subterminale, C. perfringens, C. septicum, C. tertium, C. histolyticum, C. cadaveris, C. baratii, C. novyi type A, C. novyi type B and C. sphenoides. Thirty one isolates out of the 65 isolates were toxigenic, four C. tetani were selected for preparation of tetanus toxin. The toxin produced by the four isolates were evaluated by flocculation test and determining the Minimal Lethal Dose for mice. The two most toxigenic isolates were then chosen for toxoid production. The produced toxoids evaluated according to WHO recommendations were sterile, innocuous, non-toxic and not reverted to toxin. The titer of the toxoid of the two strains was 100 Lf / ml and 80 Lf / ml.

23/8 SUSCEPTIBILITY OF NOSOCOMIAL PATHOGENS TO CERTAIN ANTIBIOTICS AND BIOCIDES COMMONLY USED IN EGYPTIAN HOSPITALS

A.A. Salama, A.M.S. Hosny, A.M. Ahmady, A.A. Abdel-Hamid and A.S.M. Hussien

Department of Microbiology and Immunology, Faculty of Pharmacy,

Cairo University.

A total of one hundred isolates were recovered from clinical specimens collected from hospitalized patients with nosocomial infections The etiological agents encountered were enteric gram-negative rods (38%) mainly Proteus mirabilis (16%), Pseudomonas aeruginosa (30%), gram-positive cocci (29%) mainly Staphylococcus aureus (20%) and Candida tropicalis (3%). Antibiotic susceptibility tests showed that for staphylococcal isolates, rifampicin was the most effective antibiotic (93%) followed by vancomycin and sulfamethoxazole/ trimethoprim (66% each), gentamicin (62%), amoxycillin/clavulanate and chloramphenicol (52% each). Cefamandole, clindamycin, tetracycline, erythromycin, cefoperazone, cefotaxime, ceftriaxone, and polymyxin B showed activity against less than 50% of these isolates. Ninety percent of the isolates were resistant to methicillin and all isolates were resistant to ampicillin and penicillin. As for isolates of Enterobacteriaceae, imipenem was the most active antibiotic (95%), followed by ofloxacin (87%), amikacin (68%), gentamicin (61%), ciprofloxacin (58%), and piperacillin (50%). Sulfamethoxazole/trimethoprim, aztreonam, cefoperazone, cefotaxime, ceftriaxone, ampicillin, chloramphenicol, nalidixic acid, tetracycline, and polymyxin B showed activity against less than 50% of the isolates. As for P. aeruginosa, all isolates were sensitive to polymyxin B and resistant to ampicillin, cefotaxime, chloramphenicol and nalidixic acid. Imipenem was the second most active antibiotic (90%) followed by piperacillin (77%), amikacin (73%), aztreonam and ciprofloxacin (70% each), ofloxacin (67%), cefoperazone and gentamicin (60% each). Sixty two percent, 58% and 73% of staphylococcal, Enterobacteriaceae and P. aeruginosa isolates, respectively, were found to be multi-drug resistant. All isolates of Candida tropicalis were susceptible to fluconazole. The assessment of the antimicrobial activity of five commercial biocide preparations against the recovered nosocomial isolates demonstrated that povidone-iodine (Betadine) exhibits the highest biocidal activity, followed by sodium dichloro-isocyanurate (Presept), a combination of chloroxylenol and dichloroxylenol (Dekol), a combination of cetrimide and chlorhexidine (Cetridine Fort), and finally saponified phenol (Abdeen Phenol). Povidone-iodine and sodium dichloro-isocyanurate had the most rapid biocidal activity followed by Cetridine Fort, Dekol and finally Abdeen Phenol. The activity of Abdeen Phenol was not affected by the presence of organic matter while that of sodium dichloro-isocyanurate was markedly reduced. Increasing the inoculum size did not significantly affect the activity of all biocide preparations at the dilutions recommended by the manufacturers. This study reveals the high rate of resistance to antibiotics among nosocomial pathogens in Egyptian hospitals and shows that biocides remain an effective tool against these pathogens. Their use according to the dilution and contact time recommended by the manufacturers offers a high activity level against almost all isolates. The possible link between antibiotic and biocide resistance needs to be considered further in a clinical context.

24/8 HUMAN PARVOVIRUS B19 INFECTION IN PATIENTS WITH SYSTEMIC LUPUS ERYTHEMATOSUS: CORRELATION WITH SYSTEMIC LUPUS ACTIVITY MEASURE

W.K. Mowafy, O.M. Saleh* and Z. Abdel Azim**

Microbiology and Immunology, *Internal Medicine and **Dermatology and Venerology Departments, Faculty of Medicine, Mansoura University

Human parvovirus infection and its correlation with systemic lupus erythematosus (SLE) clinical and laboratory parameters was investigated in this study. The study included 34 female patients with SLE and 14 female healthy subjects of matched age. Detection of parvovirus B19 DNA in the sera was done using nested PCR technique. Clinical examination and other laboratory investigations were done including routine laboratory investigations, c-reactive protein (CRP), anti-nuclear antibody (ANA), anti-double stranded DNA (ds-DNA), and serum complement components (C3 and C4). Systemic lupus activity measure (SLAM) was estimated using a scoring system. B19 DNA viremia was found in 17.6% of patients and none of the controls. An association between B19 positivity and SLAM was found. All B19 positive patients were CRP positive. It was concluded that there is increased prevalence of B19 infection among SLE patients which coincides with SLE exacerbation. So, B19 infection should be ruled out in SLE exacerbation and CRP positivity might be suggestive of the presence of infection in SLE patients including parvovirus infection.

25/8 PURIFICATON AND PROPERTIES OF THE EXTRACELLULAR LIPASE(S) OF BACILLUS CEREUSSH6-HEH/CAND BACILLUS STEAROTHERMOPHILUS SH5-HEH/NC

W.A. Hassanein, Y.A. El-Zawahry, A.A. El-Mougith and A.M. El-Shawadfy

Botany Department, Faculty of Science, Zagazig University, Egypt.

All tested soil samples contained considerable numbers of lipolytic bacteria, however, the highest mesophilic and thermophilic lipolytic bacterial populations (1.69x106 c.f.u./g soilDW) and (2.9x104 c.f.u./g soilDW) were recorded in soil samples no. 7 (Shrweedah-Sharkia, non-cultivated) and no. 6 (Hehia-Sharkia, cultivated), respectively. Out of 153 mesophilic and 52 thermophilic lipolytic bacterial isolates, 16 mesophilic and 6 thermophilic lipolytic isolates had the highest lipase activity on the agar plate containing olive oil as a substrate. The most lipolytic ten isolates (7 mesophilic & 3 thermophilic) were selected by using tributyrin-hydrolysis test. The most lipolytic active mesophilic (lipase activity [LA]; 37.5 U/ml & specific lipase activity [SLA]; 17.1 U/mg protein) and thermophilic (LA; 34.5 U/ml & SLA; 15.5 U/mg protein) isolates were identified as Bacillus cereusSh6-Heh/C and Bacillus stearothermophilusSh5-Heh/NC, respectively. Optimum incubation temperature, incubation period and pH value for lipase production by B. cereus Sh6-Heh/C and B. stearothermophilus Sh5-Heh/NC were 35˚C for 3 days at pH 7 and 55˚C for 2 days at pH 7.5, respectively. The maximum lipase activity of both isolates was achieved by using olive oil (0.3 % w/v) as a carbon source, ammonium sulphate (0.3%) as a nitrogen source and in the presence of 0.1% K2HPO4, 0.05% MgSO and 0.05% CaCl2 in the growth medium. The addition of different carbohydrates, amino acids, antibiotics and EDTA decreased the lipolytic activity for both organisms. The lipases of B. cereus Sh6-Heh/C and B. stearothermophilus Sh5-Heh/NC were extracted and purified with 27.8 & 13.8 % yield and 78.1 & 83.2-fold purification, respectively. The molecular masses of the purified lipases from B. cereus Sh6-Heh/C and B. stearothermophilus Sh5-Heh/NC were 50 & 30 kDa, respectively. The optimum conditions for the activity of purified B. cereus Sh6-Heh/C and B. stearothermophilus Sh5-Heh/NC lipases were at incubation temperature 35 & 45˚C for 2 hours; pH 7.0 & 8.0 and substrate conc. of 12.0 & 8.0 % , respectively.

26/8 OUTER MEMBRANE BARRIER AND PLASMID-MEDIATED RESISTANCE OF PSEUDOMONAS AERUGINOSA TO QUATERNARY AMMONIUM COMPOUNDS AND ANTIBIOTICS

M.M. Kh. Okasha

Department of Microbiology and Immunology, Faculty of Pharmacy,

Zagazig University, Egypt

Biocide – adapted Pseudomonas aeruginosa isolate exhibited a potent resistance to both cetyltrimethyl ammonium bromide (CTAB), benzalkonium bromide (BC) and many antibiotics in comparative with the parent strain ATCC15442 . Both the already isolated CTAB – degradable strain R/D CTAB and the adapted R CTAB derived from the parent one by standard broth dilution method, exhibited 13-, 6- and 2 – fold increase in MICs of polymyxin-B, ciprofloxacin and tobramycin, respectively. Three fold increase in MIC of CTAB, BC and chloramphenicol. In contrast, 2 –fold decrease in MIC was noted with B-lactams antibiotics (ceftazidime and imipenem) . Adapted resistance to CTAB which is commonly used as a disinfectant, was observed in P. aruginosa nosocomial infections. Analysis of the outer membrane proteins (OMPs) of the CTAB resistant strain by two dimentional polyacrylamide gel electrophoresis showed a significant increase in the level of expression of a protein (Opr R) whose molecular mass was approximately 26 KDa. A correlation between the level of expression of Opr R and the level of bacterial resistance to CTAB was observed by using a p A2800 gene mutant derived from the CTAB –resistant strain. The mutant recovered susceptibility not only to CTAB but also to BC. Lipopolysaccharides quantification by colorimetric assay of 2-keto-3-deoxyoctoate (KDO) revealed a slight increase (from 6 to 9 ug/2mg of outer membrane material) in KDO associated developing of adapted CTAB resistant strain. Furthermore, P. aerugiosa was examined for the presence of the antibacterial resistance genes qac E and qac E Delta 1. Strains that degrade CTAB as well as CTAB-adapted P.aeruginosa showed presence of 2 plasmids (33 – 60 Kb), instead of a small one (15 Kb) in the strain harboring p A2800 gene of CTAB – resistance . Restriction analysis indicates the presence of plasmid gene fragments 6.6 and 2.3 Kb in plasmids of the standard mutant harbor biocide resistance-gene and both the laboratory adapted and degradable nosocomial ones.


27/8 DETECTION OF GENETIC VARIATION IN FUSARIUM OXYSPORUM ISOLATES USING RAPD ANALYSIS

M.E. Zain, A.T. Abul-Hamd; H.M. Atta

Botany and Microbiology Dept., Faculty of Science, Al-Azhar University, Nasr City, Cairo, Egypt.

Fusarium oxysporum is a common and severe fungal pathogen responsible for wilt disease of plants. The RAPD-PCR technique was used to identify genetic correlation among 5 isolates of F. oxysporum. DNA polymorphism was detected among different isolates using 5 oligonucleotide universal primers. Two fungal isolates were found to be identical based on the RAPD correlated data.

28/8 EVALUATION OF THE ANTIBACTERIAL POTENTIAL OF BOVINE AND BUFFALO’S LACTOFERRIN UNDER HEAT DIFFERENT TREATMENTS

H.A. Mansour*, S. El Behery* and A.M. Abdel-Salam **

*Pharmacology Department (Medical Microbiology and Immunology)

* Food Evaluation & Food Science Dept., National Organization for Drug Control & Research. (NODCAR)

** Dairy Science Department, National Research Centre, Dokki, Egypt

This work was performed to study the effect of different heat treatments on the derived lactoferrin from the manipulated cow and buffalo’s milk. The milk was treated with   boiling, autoclaving and microwave, in the presence of amoxicillin residues. Antibacterial activity of lactoferrin (cow and buffalo`s milk) on E. coli, Shigella flexeneri, Salmonella typhi., Pseudomonas fluorescence and Staphylococcus aureus were studied. In addition, the majority of data have shown a highly significant antibacterial action of both heated bovine and buffalo’s lactoferrin against the five bacterial strains understudy. The heated manipulated samples incorporated with amoxicillin residues have shown the same trend of results. Concerning the effect of heat treatments on the Infra Red spectra of derived lactoferrin, mild change were noticed in case of microwave treatment. On the other hand no obvious changes were noticed in lactoferrin profile with the other treatments or in the incorporation with amoxicillin residues. The results of the spectrophotometric analysis showed obvious changes in the spectra with untreated one which can be attributed to a partial breakdown of protein fraction.

29/8 SCREENING AND GROWTH CHARACTERIZATION OF THE GREEN LIFE STOCK OF DRILL WATER FROM JEDDAH, SAUDI ARABIA

I-ISOLATION AND GROWTH CHARACTERISTICS OF SCENEDESMUS SP.

A.B. El-Sayed

Plant Nutrition Department, National Research Centre, Dokki, Cairo

The green alga Scenedesmus sp. was isolated from the drill water of Saudi Arabia among different microscopically observed species. Isolation was done based on dilution technique using N8 macronutrient solution. The isolated alga was scaled up and incubated with four different growth media under heterotrophic growth conditions. Dry weight (g.l-1), total chlorophyll (mg.l-1) and total carotene (mg.l-1) were daily measured. It was found that growth dry weight was more stimulated with BG-II enriched by 2.0% sodium chloride followed by Zarrouk medium and BG-II growth medium alone. Failure growth dry weight was observed with artificial seawater growth medium, while a dense turbid cultures were obtained due to cell enlargement as a result of inhibition of cell division and salt precipitation. Growth expressed as total chlorophyll accumulation exhibited a different manner, where BG-II surpasses the other examined media. As for total carotene, the maximum was obtained with Zarrouk medium which might be as a result of their higher content of nitrate and carbonate ions.

30/8 THE IMPACT OF SILVER SULFADIAZINE, BENZALKONIUM CHLORIDE AND CHLORHEXIDINE ON THE CELL SURFACE HYDROPHOBICITY OF CANDIDA ALBICANS AND CANDIDA TROPICALIS

A.A. Salama

Department of Microbiology and Immunology, Faculty of Pharmacy,

Cairo University

Candida albicans is the most common nosocomial fungal pathogen that causes both systemic and superficial infections. Non-albicans Candida species such as Candida tropicalis are now emerging as important agents of fungal infections. The adherence and cell surface hydrophobicity (CSH) of Candida are considered critical factors contributing to its colonization potential and virulence. A total of 70 clinical isolates of Candida species were recovered from hospitalized patients. These isolates included 43 isolates of C. albicans, 19 isolates of C. tropicalis, 3 isolates of C. krusei and 5 isolates of other non-albicans Candida species. The susceptibility of these isolates to silver sulfadiazine (SSD), benzalkonium chloride (BKC) and chlorhexidine (CHX), which are recently used for impregnating or coating central venous and urinary catheters to prevent catheter-related infections, was determined. The capability of adherence of all isolates to polystyrene was tested. All isolates were capable of adherence and biofilm formation but different capabilities were found among different species and even among isolates of the same species. The CSH of all isolates were assessed by a   biphasic   aqueous-hydrocarbon   assay.   Isolates of C. tropicalis and C. krusei were found to be significantly more hydrophobic than those of C. albicans. The effect of limited exposure of C. albicans and C. tropicalis isolates to concentrations of 25 and 50µg/ml of SSD, BKC and CHX on the CSH was investigated. The three antiseptics significantly reduced the CSH of all isolates of both C. albicans and C. tropicalis and this reduction was proportional to the antiseptic concentration. BKC induced a pronounced reduction that was significantly higher than that induced by SSD or CHX. Analysis of inter-species variation in the impact of antiseptics on the CSH of the two Candida species revealed a significant variation between C. albicans and C. tropicalis, where the former was more affected by all the tested antiseptics as compared to the latter. The present study revealed that exposure to silver sulfadiazine, benzalkonium chloride and chlorhexidine significantly reduced the CSH of both C.albicans and C.tropicalis isolates to varying degrees. This appears to be a salutary feature of these antiseptics potentiating their antimicrobial action.


31/8 Microbiological study on microorganisms causing respiratory tract infections in Upper Egypt

G.F. M. Gad*, M.A. Omar*, R.A.A. El-Domany*, M.S.E. Ashour**

and A.F. Abd El-Ghaffar***

Microbiology Departments, Faculty of pharmacy, El-Minia University*, El-Azhar University ** and Faculty of medicine, El-Minia University ***

Five hundreds and fifty nine clinical strains were isolated and identified from 322 patients suffering from respiratory tract infections. Patients were attending the clinics of El-Minia General, El-Minia University and El-Minia Chest hospitals. Patients represented different ages, sex, and types of infections. Out of the 322 patients, 204 were suffering from upper respiratory tract infections and 118 patients were suffering from lower respiratory tract infections. Patients of upper respiratory tract infections were suffering from chronic suppurative otitis media (63 patients), tonsillitis (50 patients), pharyngitis (48 patients), and sinusitis (43 patients). Out of the total isolates, Staphylococcus aureus was the most prevalent organism, followed by Streptococcus pyogenes and Klebsiella pneumoniae (17.71, 12.34, and 11.27% respectively). Pseudomonas aeruginosa represented 6.26%. Serratia marcescens and Morganella morganii were the least isolated organisms. The results revealed that 52.42% of the strains were isolated from males and 47.58% from females. Staphylococcus aureus was the most prevalent organism in males (21.16%) while in females Strept. pyogenes was the most prevalent organism (14.29%). Also, the study revealed that Staphylococcus aureus was the most frequent isolate in age groups between 1-20, 21-40 and 41-60 years old (20.85%, 17.02% and 16.67% respectively). However, both Staphylococcus aureus and Klebsiella pneumoniae were isolated with equal incidences, 12% each, in elder patients (more than 60 year). The susceptibility pattern of the isolated bacteria to different antimicrobial agents was studied. Both levofloxacin and gatifloxacin showed the highest activity (100%), followed by ofloxacin and ciprofloxacin (96.44% and 93.39% respectively). Those are followed by amikacin (91.86%), cefotaxime (89.31%), cefoperazone (86.26%), gentamicin (84.22%), ampicillin-sulbactam (70.48%), amoxycillin-clavulanic (62.34%), cefuroxime (62.09%), lincomycin (61.83%), vancomycin (61.07%), chloramphenicol (57%), cephalexin (48.35%), cephapirin (45.29%), erythromycin (44.78%), and trimethoprim-sulphamethoxazole (43%). Amoxycillin and tetracycline were the least active ((36.64% and 32.06% respectively). Staphylococcus aureus strains resistant to amoxycillin were tested for B-lactamase production. Out of the tested strains, 62.5% were B- lactamase producers and it may be responsible for the resistance to amoxycillin.

 

 

32/8 DEVELOPMENT OF A RAPID AND QUANTITATIVE METHOD FOR THE DETECTION OF HAV USING A COMBINED IMS-MOLECULAR BEACON-RT-PCR ASSAY

K.H. Abd El Galil, M.A. El Sokkary, S.M. Kheira and A. Mulchandani*

Department of Microbiology, Faculty of pharmacy, Mansoura University, Egypt *Department of Chemical and Environmental Engineering, University of California, Riverside, CA 92521

Molecular beacons (MBs) are oligonucleotide probes that fluoresce upon hybridization. In this study, the development of a real-time RT-PCR assay to detect the presence of Hepatitis A virus (HAV) using these fluorogenic reporter molecules is described. A set of primers was designed to amplify a 125 bp of a highly conserved 5'-noncoding region (5'-NCR) in the HAV genome. A molecular beacon was designed to specifically recognize a 20 bp sequence within the amplicon. The sensitivity of the real-time RT-PCR assay was tested with serial 10-fold dilutions of either viral RNA or a synthetic HAV RNA control. Detection limits of 1 PFU (plaque forming unit) and 50 copies of synthetic RNA were obtained. The specificity of the molecular beacon was demonstratedby obtaining negative signals with a variety of environmental pathogens and indicator microorganisms. When combined with immunomagnetic separation, the real-time RT-PCR assay successfully detected HAV recovered from seeded groundwater samples. Due to its simplicity and specificity, this assay has broad applications for the quick detection of HAV in contaminated foods or water.

33/8 EFFECT OF HEAVY METAL ACCUMULATION ON THE GROWTH, DNA, AND ULTRASTRUCTURE OF BACILLUS CIRCULANS AND KLEBSIELLA PNEUMONIAE

BY

W.A. Hassanein; E.Y. Tohamy; N.M. Awny and S.M. Meselhy,

FROM

Botany Departments, Faculty of Science, Zagazig University, Egypt

Three industrial waste water samples from different sites were used for isolation of some bacterial strains which have the ability to accumulate cadmium and nickel ions. Chemical analysis of water samples revealed the presence of the highest contents of cadmium and nickel in samples A and B respectively. The microbiological analysis indicated the presence of high bacterial counts, considerably coliforms and low counts of actinomycetes in all tested water samples. Two resistant bacterial strains were isolated at the sublethal concentration of 92.8 mg/L Cd+2 and 172.8 mg/L Ni+2 added to the culture media from sample A and B, and identified as Bacillus circulans and Klebsiella pneumoniae, respectively. Increasing concentrations of Cd+2 or Ni+2 on the growth medium decreased the bacterial growth, dry weight of bacterial cells, amount of metal absorbed and metal accumulation for both bacterial isolates. Change in culture conditions, such as incubation temperature, pH value and incubation period strongly affected the growth and metal accumulation for both bacterial isolates. The optimum environmental conditions of growth and accumulation of Cd+2 by B. circulans was obtained in the presence of 37.1 mg/L Cd+2 added to the growth medium, after incubation for 2 days at 35oC and pH 7. The optimum environmental conditions of growth and Ni+2 accumulation by K. pneumoniae was obtained in the presence of 24.6 mg/L Ni+2 added to the growth medium, after incubation for 2 days at 45oC and pH 7. The biosorption of tested metal ions (Cd+2 and Ni+2) was obtained by both living or dead cells of the tested bacterial isolates. The maximum Cd+2 biosorption of living cells (13.8 mg/g dry wt) and dead cells (12.9 mg/g dry wt) by B. circulans were achieved after 40 and 60 min incubation respectively. The maximum Ni+2 biosorption of living cells (11.8 mg/g dry wt) and dead cells (9.7 mg/g dry wt) of K. pneumoniae were achieved after 80 and 40 min respectively. Molecular studies of the total DNA (chromosomal and plasmid) for both resistant bacterial isolates B. circulans and K. pneumoniae in the absence and presence of 92.9 mg/L Cd+2 or 172.8 mg/L Ni+2 in the growth medium respectively, revealed complete inhibition of the total DNA for both bacterial isolates as a result of metal treatment. Ultrastructure studies using transmission electron microscope TEM was carried out for both B. circulans and K. pneumoniae cells in the absence or presence of 92.9 mg/L Cd+2 and 172.8 mg/L Ni+2 in the growth medium respectively. There were significant morphological and ultrastructure changes of both bacterial isolates treated with metals as well as dark of metal precipitates on the cell wall and inside the cells was observed.

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