Search PubMed⌕ Search

Biomedical subjects

M J Alfa

Publications and source records attributed to M J Alfa.

At least 37 records · Page 2Linked to original sources
PubMed1996

Comparison of ion plasma, vaporized hydrogen peroxide, and 100% ethylene oxide sterilizers to the 12/88 ethylene oxide gas sterilizer.

OBJECTIVE: The performance of a standard gas sterilizer, which uses a mixture of 12% ethylene oxide (EtO) and 88% chlorofluorocarbon as the sterilizing gas (12/88), was compared to selected gas, ion plasma, and vaporized hydrogen peroxide (H2O2) sterilizers that do not use chlorofluorocarbons. The effect of serum and salt on sterilizer performance was evaluated. DESIGN: Test carriers (porcelain and stainless steel penicylinders, or 125-cm lengths of plastic tubing [internal diameter of 3.2 mm]) were inoculated with Escherichia coli, Enterococcus faecalis, Pseudomonas aeruginosa, Mycobacterium chelonei, Bacillus stearothermophilus spores, Bacillus subtilis spores, and Bacillus circulans spores and then subjected to sterilization using 12/88, 100% EtO, ion plasma, or vaporized H2O2. The bacterial inoculum was prepared with and without 10% serum and 0.65% salt, and the residual bacterial load after sterilization as determined using viable counts. RESULTS: All of the sterilizers tested effected a six-log10 reduction of the bacterial inoculum on penicylinders, unless 10% serum and 0.65% salt were present, in which case the 100% EtO, vaporized H2O2, and ion plasma sterilizers were not as effective as the 12/88 sterilizer. None of the sterilizers could eradicate 10(6) CFU of all of the bacteria in 10% serum and 0.65% salt when inoculated inside a narrow lumen. CONCLUSIONS: The margin of safety for the 100% EtO, vaporized H2O2, and ion plasma sterilizers is less than that of the 12/88 sterilizer. The inability of all sterilizers, including the 12/88, to kill organisms in narrow lumens reliably when serum and salt were present raises concern about the current practice of gas sterilization of flexible endoscopes.

Analysis of Variance↗
PubMed1995

Mycobacterium abscessus infection of a Norplant contraceptive implant site.

The authors report a case of Mycobacterium abscessus infection of a subdermal levonogestrel implant (Norplant) site. The infection lasted 12 weeks and was indolent, skin manifestations were low grade and difficult to detect. Culture of exudate samples showed that M. abscessus was the only causative agent. After the implant was removed the patient's arm healed uneventfully without antimycobacterial therapy. The authors recommend that if Gram staining of apparently infected material from an implant site does not reveal a causative organism, then cultures should be done for mycobacteria and fungi. Kinyoun staining for acid-fast bacteria and calcoflour-white staining for fungi should also be performed. The implant should be removed and the patient given antimicrobial therapy as indicated. The authors emphasize the need to be aware of the potential for M. abscessus infection of implant sites and stress that appropriate microbiologic culture procedures are essential for accurate diagnosis.

Adult↗
PubMed1995

Transmission of Ureaplasma urealyticum from mothers to full and preterm infants.

This study assessed maternal genital colonization and subsequent neonatal transmission rate of Ureaplasma urealyticum in pregnant women in an average socioeconomic population. In addition very low birth weight infants were assessed to determine whether the presence of U. urealyticum correlated with increased risk of developing respiratory problems. The study group consisted of 108 sequential full term mothers and 104 preterm mothers delivering in a tertiary care hospital in central Canada. The genital carriage rates (assessed using placental sampling) of ureaplasmas in term and preterm mothers were 25.9 and 19.2%, respectively (P = 0.3185). Acquisition of ureaplasmas in the neonatal respiratory tract of neonates occurred significantly (P = 0.0182) more often in preterm neonates (11 of 130; 8.5%) than in term neonates (2 of 110; 0.9%). Very low birth weight (VLBW) infants (< or = 1500 g) were at greater risk (P = 0.042) of acquiring ureaplasmas in their respiratory tracts (5 of 26; 19%) than larger preterm neonates (6 of 104; 5.8%). All VLBW infants with respiratory colonization by ureaplasmas (5 of 5) developed bronchopulmonary dysplasia compared with 33% (7 of 21) of VLBW neonates without ureaplasmas (P = 0.028). This difference in bronchopulmonary dysplasia development among VLBW infants was independent of further stratification by birth weight. These VLBW neonates with ureaplasmas also stayed significantly (P = 0.037) longer in the neonatal intensive care unit (43.6 +/- 10.4 days) than did other preterm neonates (22.1 +/- 20.8 days). Our results demonstrate that VLBW preterm neonates have increased risk of acquiring U. urealyticum.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1995

Use of tissue culture and animal models to identify virulence-associated traits of Haemophilus ducreyi.

To identify virulence-associated properties of Haemophilus ducreyi, 34 strains of this sexually transmitted pathogen were evaluated for in vitro phenotypic characteristics of potential relevance to chancroid pathogenesis and for their ability to produce lesions in the temperature-dependent animal model for chancroid. Of the 34 strains tested, all but three produced a cytopathic effect on human foreskin fibroblasts (HFF) and all but six strains formed large microcolonies on HFF monolayers. A subset of 12 selected strains underwent more extensive analyses and, when evaluated for both their cytadherence kinetics and growth in the presence of HFF monolayers, it was found that several of these strains had a very limited ability to attach to HFF cells. When the same 12 strains were tested in the temperature-dependent rabbit model, only the seven strains which were positive in all of these in vitro-based tests readily produced lesions. In contrast, the five strains that were noted to be deficient in one or more of the phenotypic characteristics scored in the in vitro systems did not produce lesions. This association between the traits measured in vitro and the ability to produce dermal lesions was significant (P = 0.0012). These results suggest that in vitro behavior may be used to predict the virulence potential of H. ducreyi strains. Moreover, the phenotypic characteristics described in this study are appropriate focal points for efforts to determine the molecular basis of the virulence of this pathogen.

Animals↗
PubMed1994

A quantitative chemiluminescent ribosomal probe method for monitoring adherence of Haemophilus ducreyi to eukaryotic cells.

This study evaluated a commercially available chemiluminescent-labelled, ribosomal RNA-directed DNA probe (CRP) as a method to quantitate attachment of H. ducreyi to human foreskin fibroblast (HFF) cells. Evaluation of four strains of H. ducreyi demonstrated that the CRP assay was unaffected by eukaryotic cells and its advantages were: (i) quantitation was done after attachment so it did not interfere with the attachment process, and (ii) it was a rapid, reliable method for quantitating bound bacteria, despite bacterial clumping. Gentamicin-killed H. ducreyi attached to both HFF cells and viable controls, suggesting that the adhesins are components constitutively present on the surface of H. ducreyi. This method may be widely applicable, since the probe recognizes most prokaryotic rRNA sequences.

Bacterial Adhesion↗
PubMed1994

In-hospital evaluation of orthophthalaldehyde as a high level disinfectant for flexible endoscopes.

One hundred endoscopes used for bronchoscopy (30), gastroscopy (35) or colonoscopy (35) were studied to determine the efficiency of a new high level disinfectant, orthophthalaldehyde (OPA). Manual cleaning was the method studied since this would be the least effective and thereby provide the greatest challenge to the 0.5% (w/v) OPA solution. The OPA was convenient and easy to use since it did not have irritating vapours and as it is used directly, does not require dilution. Our study demonstrated that the OPA was stable for up to 14 days despite repeated re-use. The cleaning/disinfection procedure could achieve a > or = 5 LOG10 reduction in bacterial load. This in-hospital evaluation supports the conclusion that OPA is an effective choice as a high level disinfectant for flexible endoscopes.

Bronchoscopes↗
PubMed1994

Methicillin-resistant Staphylococcus aureus. Optimization of the agar screen plate method.

Swab inoculation of oxacillin agar screen plates was compared with drop inoculation for detection of methicillin-resistant Staphylococcus aureus. The poor sensitivity of the swab method (59%) was related to heteroresistance of the S. aureus isolates. We recommend the drop method (100% sensitivity) because interpretation of results was significantly easier, making it more reliable.

Methicillin Resistance↗
PubMed1993

Humoral immune response of humans to lipooligosaccharide and outer membrane proteins of Haemophilus ducreyi.

The humoral immune response to purified lipooligosaccharide (LOS) and outer membrane proteins (OMP) of Haemophilus ducreyi was evaluated. Sera from chancroid-endemic (Uganda, Kenya) and -nonendemic (Canada) countries were tested by an ELISA. The response to OMPs was cross-reactive with other Haemophilus species, and elevated levels of antibody were detected in patients that did not have chancroid. The LOS component stimulated an H. ducreyi-specific immune response that was detected only in patients with chancroid. The sensitivity of the LOS ELISA was 96% (95% confidence interval, 89.9%-100%) and the specificity was 97% (95% confidence interval, 95.8%-98.2%). Thus, the anti-H. ducreyi LOS immune response is a significant diagnostic and epidemiologic indicator.

Adolescent↗
PubMed1993

Haemophilus ducreyi adheres to but does not invade cultured human foreskin cells.

Haemophilus ducreyi is the etiologic agent of the localized genital ulcer disease known as chancroid. The pathogenesis of this organism is poorly understood. The role of attachment in the disease process has not been evaluated. In this study, 125I-H. ducreyi was used to quantitatively evaluate the interaction of virulent and avirulent H. ducreyi strains with human foreskin cells. Using this in vitro model system, we demonstrated that, at 22 and 35 degrees C, the attachment of virulent H. ducreyi 35000 to human foreskin cells was significantly more marked than that of avirulent H. ducreyi A77. Although H. ducreyi penetrated between human foreskin cells, internalization was not a major component. Our competition assay data suggest that the attachment mechanism of H. ducreyi may be similar to that of Neisseria gonorrhoeae. We speculate that the attachment and microcolony formation of virulent H. ducreyi may provide a mechanism for bacterial localization and evasion of host defenses.

Bacterial Adhesion↗
PubMed1993

Human interferon-gamma has three domains associated with its antiviral function: a neutralizing epitope typing scheme for human interferon-gamma.

An antiviral activity-neutralizing monoclonal antibody (mAb), MIF3037, was developed by the immunization of BALB/c mice with recombinant human interferon-gamma (rhuIFN-gamma). Its neutralizing activity suggests that its epitope may be at or adjacent to a functional domain on the huIFN-gamma. MIF3037 was compared with representative mAb of previously identified epitope-specific groups in a competitive binding assay. In an attempt to determine if there are other functional epitopes recognized by mAb developed with different preparations of huIFN-gamma or different hybridoma screening methods, 14 additional mAb contributed by five other laboratories were similarly analysed. Based on their ability to bind to huIFN-gamma, all the neutralizing mAb except MIF3037 may be classified into three previously defined groups: E1, E2 and E1/E2. Monoclonal antibodies of the E1 group do not compete with those of the E2 group for huIFN-gamma binding, indicating that the E1 and E2 epitopes are distinct domains on the huIFN-gamma important for the antiviral function. Monoclonal antibodies of the E1/E2 group compete with some of the mAb of E1 and/or E2 groups and may bind to regions of the huIFN-gamma that partially overlap the E1 and E2 epitopes. MIF3037 demonstrated no competitive binding inhibition with mAb of the previously identified epitope specificity groups and, therefore, must represent a distinct functional epitope, E3. The huIFN-gamma, therefore, must have at least three distinct functional domains; none of these appeared to be responsible for cell surface receptor binding. Based on this finding, the epitope typing scheme must be extended to include the E3 epitope. The epitope specificity relationships of 13 neutralizing mAb developed by five other laboratories were established which allows correlation of results obtained with these mAb by different laboratories. The location of the epitopes of four widely studied mAb, 69B, 73A, 113B and 220A12, have been deduced based on their competition with the E1 mAb which have recently been mapped.

Animals↗
PubMed1992

Cytopathic effect of Haemophilus ducreyi for human foreskin cell culture.

An explant adult foreskin cell culture (FS2-3) was compared with human lung carcinoma cell culture (A549) with regard to the ability of Haemophilus ducreyi to produce a cytopathic effect. The survival of H. ducreyi for up to 26 days in FS2-3 cells was far greater than in any previously described in-vitro culture system. H. ducreyi survived for up to 7 days in A549 cells. The H. ducreyi cells grew and formed "fungal-like" microcolonies on the eukaryotic monolayer. Portions of the microcolonies remained attached despite extensive washing. Transmission electronmicroscopy indicated that, at 48 h after infection, the H. ducreyi cells did not penetrate the FS2-3 cells but they were closely associated with them; there was only a 2-5 nm gap between the H. ducreyi cell wall and the FS2-3 membrane. The virulent H. ducreyi strains RO18 and 35,000 produced a cytopathic effect on FS2-3 cells that did not appear to be due to a soluble toxin. These strains did not produce any CPE on A549 cells. H. influenzae and the avirulent H. ducreyi strain CIP542, inoculated in the same concentration and incubated for the same length of time, did not produce CPE on FS2-3 cells. This study demonstrated that the use of FS2-3 foreskin cell culture provided an in-vitro approach for evaluating the cytopathic effect of virulent H. ducreyi whereby, unlike in other in-vitro systems, viability of the micro-organism could be readily maintained.

Bacterial Adhesion↗
PubMed1992

Identification of highly conserved and species-specific polypeptides of Haemophilus ducreyi.

Chancroid is a sexually transmitted diseased caused by Haemophilus ducreyi. The pathological manifestations of chancroid are unique among Haemophilus species and the virulence factors of H. ducreyi that account for these features have not been identified. Some of these virulence factors may be unique components of H. ducreyi, but attempts to identify H. ducreyi-specific components have been unsuccessful. Four polypeptides--A, B, C and D of 83, 77, 56 and 28 kDa, respectively--were identified with a panel of nine H. ducreyi-specific monoclonal antibodies (MAbs). Polypeptide C was one of the five major proteins in H. ducreyi and demonstrated micro-heterogeneity in SDS-PAGE. Polypeptides A, B and D were present in only small amounts in whole-cell lysates of H. ducreyi. The relative amounts of A and B varied, suggesting that they may be precursor molecules. The unique polypeptides C and D were not exposed on the surface. Polypeptide C was highly soluble and did not appear to be membrane-bound, whereas polypeptide D appeared to partition with the cytoplasmic membrane and was soluble in Sarkosyl. All four polypeptides appeared to be unique to H. ducreyi since MAbs directed against them did not cross-react with H. influenzae, H. parainfluenzae or Neisseria gonorrhoeae. The mol. wts of all of these polypeptides were conserved throughout 35 clinical isolates collected from 15 cities in eight countries and one reference strain of H. ducreyi that were tested.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Monoclonal↗
PubMed1991

In-hospital evaluation of contamination of duodenoscopes: a quantitative assessment of the effect of drying.

A prospective, quantitative assessment was undertaken of the effect of drying on the bacterial load in duodenoscopes that had been used for endoscopic retrograde cholangiopancreatography procedures. The endoscopes were washed and disinfected using an automatic washer and samples were taken through the suction channel at 2, 24 and 48 h post-disinfection. Twenty-one of the 42 duodenoscopes tested were contaminated. The ratio of Gram-negative bacilli to Gram-positive cocci increased from 70:1 at 2 h up to 4000:1 at 48 h for those duodenoscopes that were contaminated. Pseudomonas species (6 of 12 contaminated endoscopes) and Acinetobacter species (7 of 21 contaminated endoscopes) were the most common isolates. There was visible moisture remaining in the suction channel despite the use of the complete recommended automatic washer cycle. Bacterial concentrations reached as high as 1 x 10(7) colony forming units (cfu) ml-1. An additional 10 min of drying using either an 'in house' air line or the manual machine dry prevented bacterial overgrowth of all 19 endoscopes tested 48 h post-disinfection. If the additional 10 min of drying was used, then no alcohol rinse was required. Although no infections related to use of contaminated endoscopes were reported, it was apparent that Gram-negative bacilli were multiplying to unacceptably high concentrations and that this could be prevented by an additional 10 min of drying. The additional drying was only required at the end of the endoscopy list and not between patients.

Cholangiopancreatography, Endoscopic Retrograde↗
PubMed1989

Production of monoclonal antibodies specific for Haemophilus ducreyi: a screening method to discriminate specific and cross-reacting antibodies.

Haemophilus ducreyi is the etiological agent of chancroid. The organism shares extensive immunological cross-reactivity with other Haemophilus species. This presents substantial difficulties for the production of specific monoclonal antibodies (MAbs). A competition ELISA was devised for hybridoma screening which allowed the detection of H. ducreyi-specific antibody-producing hybridoma cultures during the initial screening process. With this screening method, seven MAbs specific for H. ducreyi were obtained in a single cell fusion exercise. The specificities of the 7 MAbs were demonstrated by direct ELISA and dot immunobinding assays against several strains each of H. influenzae, H. parainfluenzae and Neisseria gonorrhoeae. Five of the MAbs reacted against all ten strains of H. ducreyi. These MAbs may permit the development of rapid and efficient immunodiagnostics for chancroid. The principle of the competition ELISA for hybridoma screening should be widely applicable to the development of specific MAbs to other organisms in which immunological cross-reactivity is an impediment to hybridoma screening by conventional methods.

Animals↗
PubMed1988

Distinct domains of recombinant human IFN-gamma responsible for anti-viral effector function.

A panel of 21 independently isolated neutralizing mAb directed against the human rIFN-gamma (rHuIFN-gamma) was used to characterize those epitopes that are involved in the antiviral function of the rHuIFN-gamma. A sandwich competition assay was developed to determine the cross-reactivities between the mAb. The 125I-labeled mAb were allowed to compete with varying amounts of unlabeled mAb for binding to rHuIFN-gamma under Ag-limiting conditions, and the 50% inhibition endpoints were determined for each of the 21 mAb. The competition of each heterologous mAb relative to the competition of the homologous mAb was determined. By grouping the competition patterns of the 21 mAb, it was apparent that at least two epitopes (E1 and E2) were important to the antiviral function of rHuIFN-gamma. The possibility of the separation of the receptor binding site and signal transduction effector site(s) is discussed.

Antibodies, Monoclonal↗
PubMed1987

Production of monoclonal antibodies against recombinant human interferon-gamma: screening of hybridomas without purified antigen.

A panel of mouse monoclonal antibodies (MAbs) against the recombinant human gamma interferon (HuIFN-gamma) has been produced for the study of the structure-function relationships of this important lymphokine. Enzyme linked immunosorbent assay (ELISA) is the current method of choice to screen hybridomas for specific MAb production. The purity of the antigen used for screening dictates the specificity of the ELISA. As often is the case in many systems, adequately purified biologically active HuIFN-gamma was not readily available for this purpose. A sandwich ELISA which allowed the use of unpurified HuIFN-gamma for hybridoma screening was developed. A rabbit antiserum against the denatured HuIFN-gamma purified by SDS-PAGE was prepared and the nonspecific binding activity was removed by adsorption to control cell proteins immobilized on Sepharose. The adsorbed immunoglobulin fraction was bound to the ELISA plate: (i) to trap HuIFN-gamma specifically from the whole cell lysate, thus providing specificity for MAb detection, and (ii) to avoid direct adsorption of the HuIFN-gamma to the ELISA plate because others have found that this prevented detection of neutralizing MAb. The sandwich ELISA detected both neutralizing and non-neutralizing MAbs with relatively low false positive reactions. This approach to the development of an ELISA method to screen hybridomas without purified antigen should be applicable to the production of MAbs to other proteins.

Animals↗