Detection of the hepatitis C virus in tear fluid of patients with chronic hepatitis C

The highest prevalence rate of hepatitis C virus [HCV] infection in the world has been reported among Egyptians and up to 20 to 40% of patients chronically infected with HCV, the mode of transmission is still unknown. So the aim of this study was to detect the presence of HCV genome in the tear fluid of the patients with chronic HCV infection which may be a source of infection and to investigate the tear film function and stability in these patients. This study was done on 50 chronic active hepatitis C patients and 20 healthy subjects as a control group. The studied groups were subjected to full ophthalmological examination, liver function tests, qualitative detection of HCV-RNA in the serum and tear fluid by PCR and quantitative measurement of HCV-RNA in serum. Schirmer's test and Break Up Time [BUT] show statistically significant decreased values in patients with +ve HCV RNA in their sera, while Rose Bengal scoring test show significant increased values. These results become more pronounced in patients with +ve HCV RNA in their tears. The viral RNA was detected by RT-PCR in the tear samples in 26 patients out of the 50 studied patients [52%] and most of cases with +ve HCV RNA in tears were associated with severe viraemia. We concluded that HCV-RNA was detected in the tear fluid quite, frequently. This fluid therefore may be potentially responsible for non-blood transmission of the hepatitis C virus. This study also indicated that HCV-RNA play a role in development of the dry eye in those patients. So we recommended screening for HCV-RNA in patients with unexplained dry eye and appropriate disinfection of equipment in ophthalmological practice to avoid viral transmission

Genetic variation at transforming growth factor-beta1 Promoter [-509 C/T] among atopic asthmatic patients

Bronchial asthma is characterized by chronic lung inflammation and airway remodeling associated with wheezing, shortness of breath, coughing and acute bronchial hypersensitivity to a variety of innocuous stimuli. During development of asthma TGF- beta1 is a negative regulator of inflammation, but in chronic asthma, it has a pro-inflammatory role and consequently exacerbates lung deterioration. The aim of this study was to screen for -509C/T single nucleotide polymorphism [SNP] in tgf-beta 1 gene promoter in asthmatic patients and to determine effect of TGF- beta1 on mechanism of asthma by correlating its level to serum total IgE and PBEC. The study enrolled 70 asthmatics of various severities, diagnosed clinically, by intradermal skin tests to various environmental allergens, radiologically and by determining FEV1. Also, the study enrolled 20 healthy controls. -509 C/TSNP were detected by PCR-RFLP and TGF- beta1. Total IgE was estimated by ELISA. The study showed that TT genotype and T-allele of tgf- beta1, gene prevails more frequently in asthmatic patients than controls [T allele: 44.3%, 22.5%, ccP. <<0.05>>], also severe asthmatics carried TT gene 9.5 times versus controls, whereas mild asthmatics carry TT genotypes 3.3 times as controls. TGF- beta1 plasma level was higher in patients than controls [16.73+/-2.34 ng/mf in mild asthmatics, 40.24+/-5.55ng/ml in severe asthmatics and 7.75 +/- 1.79 ng/ml in controls]. C/T genotypes and alleles have their effects on total IgE level and PBEC which are correlated to asthma severity, TGF- beta1 plasma level, serum total IgE and PBEC showed strong positive highly significant correlation [P<0.001]

On conclusion: -509/C/T SNP of tgf- beta1 gene promoter is a key locus for asthma susceptibility and severity through increasing basal level of TGF-beta1 which in turn is associated with increased serum IgE and PBEC

Detection of aminoglycoside resistance genes by multiplex PCR among coagulase negative staphylococci isolated from Ventriculoperitoneal shunt infections

A Ventriculoperitoneal [VP] shunt infection is a cause of significant morbidity causing shunt malfunction and chronic ill health. This study was carried out to detect the causative pathogens associated with VP shunt infections, to assess the frequency of Coagulase negative Staphylococci [CoNS] as well as study their phenotypic resistance pattern to methicillin and different aminoglycosides [AGs], and genotypic pattern for gene encoding aminoglycosides modifying enzymes [AMEs] by multiplex PCR. This study was carried out in Medical Microbiology and Immunology and Neurosurgery Departments, Faculty of Medicine, Zagazig University during the period from June 2008 to June 2010. During this period 240 shunt procedures were carried on 175 infants and children with hydrocephalus. CoNS were identified using standard microbiological laboratory techniques. Antimicrobial susceptibility testing was performed using disc diffusion method with 4 aminoglycosides and methicillin. Multiplex PCR assay was used to identify AMEs encoding genes. 58 out of 240 shunt procedures cases showed shunt infections, [64%] caused by CoNS, [28%] by Gram negative rods [5%] by Staph aureus and [3%] by candida. 26 [70%] of CoNS isolates were methicillin resistant [MRCoNS]. 25 [67.5%] isolates of CoNS were resistant to at least one of the tested AGs and the highest resistance was to gentamicin and tobramycin [67.5%] for both followed by amikacin [27%] and streptomycin [19%]. As regard results of multiplex PCR aac [6'] le+aph [2' '] gene encoding for the bifunctional enzyme was the most common [54%] followed by ant[4]la gene encoding for the ANT[4'] - la enzyme [46%] and the aph [3']-llla gene encoding APH[3'] - Illa enzyme [40.5%]. In conclusion this study increased our knowledge about the causative pathogens of VP shunt infections, the phenotypic pattern of CoNS susceptibility to different AGs and the distribution of AMEs encoding genes in relation to methicillin susceptibility. The usage of appropriate antibiotic according to antimicrobial susceptibility testing at the probable time and the removal of the shunt apparatus are essential for successful treatment of VP shunt infection. Continued surveillance at both phenotypic and genotypic levels are recommended for monitoring the presence of other variant of the genes or new AGs resistance genes that may be produced within CoNS population

Phenotypic and genotypic patterns of aminoglycosides-resistant staphylococcus aureus clinical isolates

Aminoglycosides still play an important role in anti-staphylococcal therapies, although emerging resistance amongst Staphylococcus aureus is widespread. The aim of this work is to study the different phenotypic patterns of aminoglycosides-resistant S. aureus in relation to the results of a multiplex PCR assay for the genes encoding aminoglycoside-modifying enzymes [AMEs]. This study was carried out in Microbiology and Immunology and Surgical Departments, Faculty of Medicine, Zagazig University during the period from August, 2008 to June, 2009. One hundred and seventy clinical samples were collected and cultured. S. aureus isolates were identified using the standard microbiology laboratory techniques. Antimicrobial susceptibility testing was performed using disc diffusion method with 5 aminoglycoside antibiotics. A multiplex PCR assay was used to identify AMEs-encoding genes. A total of 81 [47.6%] S. aureus isolates were collected during this work and 43 [53.1%] isolates of which were methicillin resistant [MRSA]. In this study, 31 [38.3%] isolates were resistant to at least one of the tested aminoglycosides, and the highest resistance was to kanamycin [38.3%], followed by tobramycin [30.9%], gentamicin [29.6%], amikacin [9.9%] and netilmicin [8.6%]. As regard the results of the multiplex PCR assay, the aac[6']-Ie+ aph[2"] gene encoding the bifunctional enzyme was the most common, followed by the ant[4']-Ia gene encoding the ANT[4']-Ia enzyme and the aph[3']-IIIa gene encoding the APH[3']-IIIa enzyme. The results of this study showed statistically significant agreement between phenotypic and genotypic aminoglycoside resistance and methicillin resistance. In conclusion, this study has increased knowledge of the distribution of AMEs in S. aureus isolated in our hospitals. Continued surveillance at both the phenotypic and genotypic levels is recommended for monitoring the presence of other variants of the genes or new aminoglycoside resistant genes that may be produced within the S. aureus population and detecting early emergence of resistant organisms to establish effective antibiotic therapies and prevent nosocomial as well as environmental spread of resistant isolates

Antibiotic susceptibility of escherichia coli forming biofilm in urinary tract infection

E. coli is implicated in many biomaterial related infections. Typically, these infections are associated with biofilm formation. Cells in biofilm display phenotypic traits that are dramatically different from those of their free-floating planktonic counterparts and are more resistant to antimicrobial agents. Consequently, biofilm - related infections are inherently challenging to treat and difficult to fully eradicate with normal treatment regimens. The aim of this work is to study E. coli biofilm formation and antimicrobial susceptibility of biofilm forming strains in the planktonic and biofilm states by broth and agar dilution methods according to NCCLs. Out of 102 catheterized patient's urine samples and 58 mid-stream urine samples, 91 [56.9%] E. coli strains were isolated, 46 of them [50.5%] were capable to form biofilm by Congo-red method and 43 [47.3%] were positive biofilm producers according to the quantitative spectrophotometric assay. Most biofilm producing strains were isolated from catheterized patients [67.6%]. No significant agreement was found between E. coli planktonic and biofilm positive isolates pertaining to their antimicrobial susceptibility. There was also a clear difference in antibiotic susceptibility seen between planktonic and biofilm populations of the same strain in both MIC50 and MIC90 which were in planktonic isolates much lower than those required to inhibit positive biofilm producers. In conclusion, this study indicated that there was a significant percentage of E. coli able to grow within biofilm, especially strains isolated from catheterized patients. E. coli biofilm positive strains were highly resistant to the antimicrobials in comparison to their planktonic counterparts. Accordingly, specimens collected from catheters should be tested for antibiotic susceptibility by special methods. Also, modification of the biomaterial surface to reduce biofilm formation or irrigation with antiseptic and antiadhesive materials is highly recommended

Prevalence of hepatitis G virus RNA and anti HGV-E2 among hemodialysis patients

Hepatitis G virus [HGV] is, like hepatitis C virus, a blood-borne virus and a member of the family of flaviviridae. HGV is distributed globally and is present in the volunteer blood donor population. Thus, for epidemiological reasons, HGV is of interest in hemodialysis [HD] patients, who are at risk of parenterally transmitted infections. To study HGV infection among hemodialysis patients in our locality, 50 HD patients and 25 apparently healthy volunteer blood donors were studied. All subjects' sera were tested for transaminases [ALT and AST], HBsAg, anti-HCV IgG, HCV-RNA, anti-HGV-E2 and HGV RNA. Anti-HGV-E2 was determined using an enzyme immunoassay; HGV RNA and HCV RNA were detected using reverse transcription polymerase chain reaction [RT-PCR]. HBsAg was detected in 10 [20%] of HD patients in comparison to none of the control. Anti-HGV-E2 and HGV- RNA were found in 18% and 42% of HD patients respectively, in comparison to 8% and 12% in the control group. The total prevalence of HGV infection in our HD patients was 60%. All the HGV-RNA positive sera were negative for anti-HGVE2 and vice versa. 15 of the HGV-RNA positive patients were followed up for 18 months, one of them showed anti-HGV-E2 seroconvertion with loss of viremia. HGV exposure did not correlate with age, gender, duration of HD therapy, or history of blood transfusions. HCV-RNA and HGV co-infection was found in 71.4% of HD patients. In addition, HGV infection was not found to cause significant elevation of ALT or AST enzymes levels in the group exposed to HGV. Although 18/21[85.7%] of HGV-RNA positive HD patients had history of blood transfusion, 3 [14.3%] HD patients who were HGV-RNA positive had neither history of transfusion nor other risk of parenteral exposure, supporting the hypothesis of nosocomial transmission. To conclude, patients on maintenance dialysis are at increased risk for HGV infection in our locality. In addition to transmission through blood transfusion, HGV may have been transmitted nosocomially, patient-to-patient, within the HD unit and the universal precautions should be strictly followed to prevent transmission of viruses among HD patients. The HGV RNA is considered a marker for current HGV viremia, while the anti-HGV-E2 is considered a marker for recovery from HGV infection. Detection of both antibody and viraemia is important to establish the real rate of HGV infection

Detection of metallo beta lactamase producing pseudomonas aeruginosa in clinical isolates

Background and objectives: Metallo beta lactamase [MBL] producing Pseudomonas aeruginosa have been reported to be important cause of nosocomial infections. The appearance of MBL genes and their spread among bacterial pathogens is a matter of concern with regard to the future of antimicrobial therapy. The present study aimed to assess the susceptibility and cross-resistance of the widely used antimicrobials against P. aeruginosa and to study the prevalence of MBL enzyme by phenotypic and genotypic methods

Methods: A total of 45 isolates of P. aeruginosa from 120 different clinical specimens [37.5%] between May 2009 and January 2010 were subjected to susceptibility testing against various antibiotics by disc diffusion method. Imipenem and l or ceftazidime resistant isolates by MIC method were selected for detection of MBL production by disc potentiation test. All P.aeruginosa isolates were subjected to duplex polymerase chain reaction to detect IMP and VIM genes of transferable MBLs

Results: Out of 45 P. aeruginosa isolates, 21 [46.7%] were resistant to imipenem [IPM] and 45[100%] were resistant to cefepime. 18[62%] of ceftazidime [CAZ] resistant P. aeruginosa and 15[21%] of imipenem resistant P. aeruginosa were MBL producers by disc potentiation test. Among 45 P. aeruginosa isolates, 15 [33.3%] were positive by duplex PCR. All MBL producer P. aeruginosa were resistant to all tested antimicrobial agents. In comparing the results of disc potentiation test with PCR results IPM / EDTA disc potentiation test was highly sensitive and specific while CAZ/EDTA was highly sensitive but less specific

In conclusion: MBL- mediated IPM and /or CAZ resistance in P. aeruginosa is a cause for concern in therapy of critically ill patients, so there is a need to do surveillance to detect MBL producers

Fas- mediated peripheral T-cell apoptosis in chronic hepatitis-C virus infection

Hepatitis-C virus [HCV] infection is considered to be the leading cause of chronic liver disease with a prevalence rate between 0.5% and 10% in different countries. HCV is remarkably efficient in establishing persistent infection, possibly mediated by an impaired immune response to HCV infection. Regulation of T-cell apoptosis, which is mediated by interaction of a death receptors Fas [CD[95]] and its ligand [Fas ligand] is critical in generation of effective immune response against viral infection. Increase in peripheral T-cell apoptosis contributes to the down regulation of the immune system in chronic HCV infection

Aim of the work: Measuring the amount of apoptosis and the expression of Fas receptor in peripheral T-lymphocytes and their relationship to laboratory criteria in chronic HCV patients

Materials and methods: The study included two groups, 30 patients with chronic HCV infection [16 males, 14 females], aged 38 +/- 13.6 years and 10 healthy subjects as a control group [6 males, 4 females] aged 35.5 +/- 12.5 years. Both groups were subjected to peripheral blood mononuclear cell [PBMC] separation by Ficoll-Hipaque density gradient centrifugation, culture of peripheral blood T-lymphocytes in RBMI 1640 with 10% fetal calf serum for 48 hours in 37[degree]C with 5% CO[2]. Then, apoptosis of cultured T-lymphocytes was determined on flow cytometry using Annexin-V FITC [fluorescein isothiocyanate isomer-1] technique. The amount of Fas expression on separated Tlymphocytes was determined using FITC anti-Fas monoclonal antibodies. Liver function tests and viral load were taken from patient's reports

Results: Fas-expression in patients with chronic HCV was 32.19% +/- 12.42% which was significantly higher than that of healthy control group 6.4% +/- 2.65% [P < 0.001]. Amount of apoptosis of cultured T-lymphocytes from patient group [48.52% +/- 14.18%] was significantly higher than that of the control group [12.48% +/- 6.35%] [P < 0.001]. There was a significant positive correlation between Fas expression and the amount of apoptosis in the patient group [P < 0.001]. Also there were significant positive correlations between viral load and both of Fas expression and lymphocyte apoptosis [P < 0.01], while there was no significant correlation between liver enzymes levels and Fas expression or apoptosis in chronic HCV patients

Conclusion: There was increased Fas expression and apoptosis in peripheral T-lymphocytes in patients with chronic HCV infection which may contribute to down regulation of the immune response and persistent infection

Assesment of soluble ICAM-1 and soluble E-selectin in juvenile idiopathic arthritis: clinical and laboratory correlation

To determine serum levels of soluble adhesion molecules [sICAM-1 and sE-selectin] in juvenile idiopathic arthritis patients. Also, to assess any changes in these levels during remission and exacerbation and to correlate them with clinical and laboratory variables. The serum and synovial levels of soluble intercellular adhesion molecule 1 [sICAM-1] and soluble E-selectin were determined using enzyme linked immunosorbent assay [Elisa] in 44 patients with JIA, 14 systemic [JIA-sys], 15 polyarticular [JIA-poly] and 15 oligoarticular [JIA-oligo], who had active disease or were in remission and 20 healthy controls. The erythrocyte sedimentation rate [ESR], C-reactive protein [CRP] and complete blood count [CBC] were recorded. [sICAM-1] and [E-selectin] were significantly higher in both [JIA-sys] and [JIA-poly] than in [JIA-oligo] or healthy controls. There was a significant negative correlation [sICAM-1] and [s-E-selectin] and age. There was no correlation with some sensitive markers of inflammation [ESR] or [CRP]. [sE-selectin] correlated with white blood cells [WBC] and platelet count in the remission stage .There was a significant correlation between synovial adhesion molecules concentration and total synovial leukocytic count. Increased levels of soluble adhesion molecules in both [JIA-sys] and [JIA-pol] may be due to endothelial cell activation and its persistence in spite of clinical remission could be used as markers of aggressive disease