Elevated anticardiolipin antibodies in acute liver failure.

Antibodies to cardiolipin (aCLA), a phospholipid primarily localized in inner mitochondrial membranes, were transiently elevated (P<0.01) when mice were exposed to an industrial surfactant and then infected with influenza B virus, a model of acute liver failure (ALF). Children with ALF also had elevated levels of aCLA.

Effects of antipyretics on mortality due to influenza B virus in a mouse model of Reye's syndrome.

Objectives: To determine the effects of acetylsalicylic acid (ASA) and acetaminophen on mortality due to influenza B infection in neonatal and weanling mice, as well as any synergistic, antagonistic or indifferent effects of the combined antipyretic and virus on mortality in mice pretreated with low doses of an industrial surfactant, Toximul MP8, which has been shown to reproduce many of the features of Reye's syndrome. In vitro studies were done to determine whether ASA or acetaminophen altered the normal, interferon-mediated antiviral responses of mammalian cells. The involvement of ASA or other commonly used xenobiotics in the induction of Reye's syndrome following virus illness has not been resolved; to do so, and to elucidate the underlying metabolic mechanism, requires these studies in an animal model.

Identification of IS1356, a new insertion sequence, and its association with IS402 in epidemic strains of Burkholderia cepacia infecting cystic fibrosis patients.

Burkholderia cepacia is now recognized as an important opportunistic pathogen in cystic fibrosis (CF) and other compromised patients. Epidemicity among CF patients has been attributed to at least one particularly infectious strain (strain ET12), and both genetic evidence and anecdotal evidence suggest that this strain, currently endemic in Ontario, and those causing an epidemic in the United Kingdom, are indeed the same. Our study was conducted to determine whether there was any association between the presence of various insertion sequence (IS) elements, the cable pilin subunit gene (cblA), electrophoretic type (ET), and ribotype (RT) in a collection of 97 clinical and 2 environmental isolates of B.

Differentiation of Listeria isolates by PCR amplicon profiling and sequence analysis of 16S-23S rRNA internal transcribed spacer loci.

The 16S-23S rRNA internal transcribed spacer region (ITS) in genomic DNA from Listeria species was amplified and sequenced so as to find sequence differences that would allow rapid species and strain differentiation. Agarose gel profiles of amplicons generated with primers designed to amplify ITS loci indicated that Listeria DNA can contain at least two distinct ITS regions. The direct sequencing of the smaller of these ITS amplicons (330 bp) was found useful for the rapid and accurate differentiation of various Listeria species.

Oligonucleotide primers designed to differentiate pathogenic pseudomonads on the basis of the sequencing of genes coding for 16S-23S rRNA internal transcribed spacers.

Universal primers targeting conserved sequences flanking the 3' end of the 16S and the 5' end of the 23S rRNA genes (rDNAs) were used to amplify the 16S-23S rDNA internal transcribed spacers (ITS) from eight species of pseudomonads which have been associated with human infections. Amplicons from reference strains of Pseudomonas aeruginosa, Pseudomonas cepacia, Pseudomonas gladioli, Pseudomonas mallei, Pseudomonas mendocina, Pseudomonas pickettii, Pseudomonas pseudomallei, and Xanthomonas maltophilia were cloned from each species, and sequence analysis revealed a total of 19 distinct ITS regions, each defining a unique sequevar with ITS sizes ranging from 394 (P. cepacia) to 641 (P.

Comparison by extended ribotyping of Pseudomonas cepacia isolated from cystic fibrosis patients with acute and chronic infections.

Multiple isolates of Pseudomonas cepacia, from two cystic fibrosis (CF) patients who were chronically infected and two others who suffered acute fatal lung infections, were examined by multilocus enzyme electrophoresis and four-enzyme ribotyping. The strains isolated from the fatalities belonged to a clone, electropherotype 12 (ET12) that is endemic in the Ontario patients' province of origin. ET12 strains have also been isolated from outbreaks in CF patients in the United Kingdom, where they are considered to be strains of high virulence and transmissibility and epidemiologically related to Ontario strains.

Linkage analysis of geographic and clinical clusters in Pseudomonas cepacia infections by multilocus enzyme electrophoresis and ribotyping.

Multilocus enzyme electrophoresis and ribotyping were used to characterize 83 strains of Pseudomonas cepacia, mostly isolated from cystic fibrosis (CF) patients, although a number of isolates from non-CF nosocomial infections and reference environmental strains were represented. Twenty enzyme electrophoretic types (ETs) were determined; of these, one clone (ET12) was associated with six of nine ribotypes (RTs) said to be geographically representative of the United Kingdom and all of the Ontario (Canada) isolates from CF patients. This clone was not associated with nosocomial infections or environmental strains and was never found in CF isolates from British Columbia or Nova Scotia, Canada, or a center in the eastern United States.

Streptococcal erythrogenic toxin genes: detection by polymerase chain reaction and association with disease in strains isolated in Canada from 1940 to 1991.

The presence of genes encoding pyrogenic exotoxins type A (speA), B (speB), and C (speC) and streptolysin O (slo) was determined by the polymerase chain reaction (PCR) to target specific sequences in 152 strains of group A streptococci. These included reference strains, representative M and T type strains, and strains associated with scarlet fever and pharyngitis collected between 1940 to 1991 and included strains from patients with severe invasive streptococcal infections. PCR amplicons were detected by agarose gel electrophoresis, and specificity was established by restriction fragment analysis.

Production of interferon by peripheral blood mononuclear cells from normal individuals and patients with chronic lymphocytic leukemia.

Peripheral blood mononuclear cells (PBMC) from normal individuals were studied to identify which cells produce alpha-interferon (IFN-alpha) in response to a virus stimulus. It was found that cells both adherent and nonadherent to plastic formed IFN-alpha after induction by any one of several viruses studied. When nonadherent cells were separated on discontinuous Percoll gradients, only the cells in the less dense Percoll fractions produced IFN, whatever the virus used.

Comparison of polymerase chain reaction and chlamydiazyme for the detection of Chlamydia trachomatis in clinical specimens.

An attempt was made to improve laboratory diagnosis of Chlamydia trachomatis and to validate the Abbott Chlamydiazyme confirmatory test used at present by comparing the polymerase chain reaction (PCR) procedure and the Abbott enzyme immunoassay. A total of 275 routine clinical specimens representing a range of positive and negative findings by Chlamydiazyme were retested by PCR. The procedures demonstrated 99% concordance for specimens with optical density (OD) readings above the Chlamydiazyme cut-off of 0.

A mouse model of coxsackievirus myocarditis.

Objective: To develop a mouse model of coxsackievirus B3 (CVB3) myocarditis.

Design: Preliminary studies have indicated that mice infected with CVB3 alone erratically responded with viral myocarditis. Prospective evaluation of the effect of cyclophosphamide at two time intervals resulted in consistency for the development of myocarditis.

Detection of genes coding for listeriolysin and Listeria monocytogenes antigen A (ImaA) in Listeria spp. by the polymerase chain reaction.

Two pairs of synthetic oligonucleotide primers were used in a polymerase chain reaction (PCR) protocol to detect targeted sequences in genes coding for listeriolysin O and Listeria monocytogenes antigen A (ImaA). Strains of Listeria spp. used in this study were isolated from clinical specimens, contaminated foods, and environmental sources.

Identification of verotoxin type 2 variant B subunit genes in Escherichia coli by the polymerase chain reaction and restriction fragment length polymorphism analysis.

A set of synthetic oligonucleotide primers was designed for use in a polymerase chain reaction protocol to specifically detect the B subunit genes in vtx2ha and vtx2hb, which code for the production of the VT2 (Shiga-like toxin II) variant cytotoxins VT2v-a and VT2v-b, respectively. An additional set of primers amplified a fragment common to the B subunits of the VT2 and the VT2 variant genes. Subsequent restriction endonuclease digestion of this amplicon permitted prediction of specific VT2 and variant genotypes on the basis of predetermined restriction fragment length polymorphisms.

Detection of genes for enterotoxins, exfoliative toxins, and toxic shock syndrome toxin 1 in Staphylococcus aureus by the polymerase chain reaction.

Eight pairs of synthetic oligonucleotide primers were used in a polymerase chain reaction (PCR) protocol to detect genes for staphylococcal enterotoxins A to E, exfoliative toxins A and B, and toxic shock syndrome toxin 1 in Staphylococcus aureus strains isolated from clinical specimens and contaminated foods. Primers were targeted to internal regions of the toxin genes, and amplification fragments were detected after the PCR by agarose gel electrophoresis. Unequivocal discrimination of toxin genes was obtained by the PCR by using nucleic acids extracted from 88 strains of S.

Surfactant-potentiated increases in intracranial pressure in a mouse model of Reye's syndrome.

Severe encephalopathy, the usual cause of death in Reye's syndrome (RS), is characterized by cerebral edema with associated increases in intracranial pressure (ICP). In previous studies, we have shown that exposure of neonatal mice to nontoxic doses of an industrial surfactant and subsequent infection with mouse-adapted influenza B (Lee) virus result in a significant increase in mortality rate and that this is associated with several of the characteristic features of human RS. In the present study we have measured ICP in the young mice undergoing their version of the disease, and we now report that the animals treated with surfactant plus virus experience increases in intracranial pressure that are significantly in excess of those in any of the three control groups.

Detection of the aerolysin gene in Aeromonas hydrophila by the polymerase chain reaction.

Synthetic oligonucleotide primers were used in a polymerase chain reaction (PCR) technique to detect the gene for aerolysin in strains of Aeromonas hydrophila and to screen for identical genes in A. caviae, A. sobria, and A.

Differentiation of Shiga toxin and Vero cytotoxin type 1 genes by polymerase chain reaction.

Two sets of synthetic oligonucleotide primers were used in a polymerase chain reaction technique to distinguish genes for Shiga toxin in Shigella dysenteriae 1 and type 1 Vero cytotoxin (VT1) in Escherichia coli. VT1a and VT1b primers directed at a common 130-base-pair (bp) fragment of the stx and sltI genes detected template nucleic acid in both Shiga toxin-positive S. dysenteriae 1 and VT1-producing E.

Differentiation of genes coding for Escherichia coli verotoxin 2 and the verotoxin associated with porcine edema disease (VTe) by the polymerase chain reaction.

Two sets of synthetic oligonucleotide primers were used in a polymerase chain reaction adaptation to distinguish the closely related genes for type 2 verotoxin (VT2 or Shiga-like toxin [SLT-II]) and the verotoxin associated with porcine edema disease (VTe or SLT-II variant [SLT-IIv]) in Escherichia coli.

Immunobiological study of interferon-gamma-producing cells after staphylococcal enterotoxin B stimulation.

Staphylococcal enterotoxin B (SEB) induced the production of human interferon-gamma (hIFN-gamma) in peripheral blood mononuclear cells (PBMC). Using specific mouse monoclonal antibodies (mAb) to hIFN-gamma, the patterns of cytoplasmic fluorescence in the PBMC from five individuals were studied. Discrete polar bodies in a ring-formation adjacent to the nuclear membrane was the most frequently observed fluorescent pattern throughout the 76-hr observation period.

Rapid and specific detection of verotoxin genes in Escherichia coli by the polymerase chain reaction.

A set of four synthetic oligonucleotide probes derived from sequences of the VT1 (Shiga-like toxin I [SLT-I]) and VT2 (SLT-II) genes were used in a polymerase chain reaction (PCR) amplification procedure to detect these genes in some enteric pathogens. A total of 40 verotoxin-producing Escherichia coli strains and 43 isolates of other recognized enteric pathogens were studied. PCR amplification products identifying the VT1 and VT2 gene sequences were observed only in nucleic acid extracted from strains found to be VT positive in traditional tissue culture assays.

A polymerase chain reaction (PCR) protocol for the specific detection of Chlamydia spp.

The polymerase chain reaction is an in vitro procedure for primer-directed enzymatic amplification of specific template nucleic acid sequences. This technique was used to detect and differentiate Chlamydia trachomatis and Chlamydia psittaci in laboratory samples of infected McCoy cells. The polymerase chain reaction was shown to be both sensitive, detecting in the order of one chlamydial DNA molecule in 10(5) cells, and specific.

Plaque assay for virulent Legionella pneumophila.

Methods of assessing virulence of Legionella pneumophila, the etiologic agent of Legionnaires disease, include the infection of guinea pigs, fertile chicken eggs, and mammalian and protozoan cell cultures. Guinea pig assays, in particular, are expensive, laborious, or unsuitable for routine screening of Legionella isolates. We have developed a virulence assay that requires the enumeration of viruslike plaques which are the result of virulent L.

Passive transfer of HIV antibody by hepatitis B immune globulin.

Two newborns of mothers carrying hepatitis B and at high risk for human immunodeficiency virus (HIV) infection developed HIV-positive test results by enzyme-linked immunosorbent assay and Western blot tests after birth. Both had been administered hepatitis B immune globulin within 48 hours of birth. Serological tests detected HIV antibody as long as 17 days after birth.