Hepatic vein thrombosis (Budd-Chiari syndrome) in an adolescent with ulcerative colitis.
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Adolescent↗
Biomedical subjects
J McFadden
Publications and source records attributed to J McFadden.
PCR-single-stranded confirmational polymorphism analysis for non-culture-based subtyping of meningococcal strains in clinical specimens.
Subspecific typing of clinical meningococcal strains is important in the investigation of outbreaks and for disease surveillance. Serogrouping, typing, and subtyping of strains currently require isolation of a meningococcus from one or more clinical specimens. However, the increasing widespread practice of preadmission administration of parenteral antibiotics has resulted in a decrease in the frequency of positive cultures obtained from blood and cerebrospinal fluid. Confirmation of meningococcal disease can be obtained by meningococcus-specific PCR from both cerebrospinal fluid (H. Ni et al., Lancet 340:1432-1434, 1992) and peripheral blood (J. Newcombe et al., J. Clin. Microbiol. 34:1637-1640, 1996) specimens. However, current PCR protocols do not yield epidemiologically useful typing information. We report here the use of PCR-single-stranded confirmational polymorphism (PCR-SSCP) analysis to amplify and type meningococcal DNA present in clinical specimens. PCR-SSCP analysis with the VR1 region of the Neisseria meningitidis porA gene as the target produced unique banding patterns for each serosubtype. Direct PCR-SSCP of clinical specimens can therefore provide typing data that can be used to investigate the epidemiology of clusters of cases and outbreaks and for disease surveillance in situations in which culture of patient specimens proves negative.
Bacterial Typing Techniques↗
Recombination in mycobacteria.
Mycobacterium tuberculosis, the causative agent of tuberculosis (TB) is thought to infect a quarter of the world's population and accounts for 3 million deaths each year. Leprosy, caused by Mycobacterium leprae continues to afflict millions. In many countries, the incidence of TB is increasing due to its association with acquired immune deficiency syndrome (AIDS) and the emergence of multidrug resistance strains of tubercle bacilli. Genes that encode major antigens, enzymes, potential virulence determinants and drug resistance in mycobacteria have been isolated and characterized; however, further genetic analysis of pathogenic mycobacteria has been severely hampered by the difficulty in precisely defining the phenotype of both wild-type and mutant genes by utilizing homologous recombination to perform allele exchange. Recombination mechanisms have been intensely studied in Escherichia coli but it is unclear how far mechanistic pathways elucidated in this species are applicable to other organisms, such as mycobacteria. The aim of this review is to examine what is currently known about homologous recombination in mycobacteria. A model is proposed to account for both low levels of homologous recombination and high levels of illegitimate recombination found in the tubercle bacillus.
Alleles↗
The magnetic immuno-polymerase chain reaction assay for the detection of Campylobacter in milk and poultry.
A rapid and sensitive technique, based on the magnetic immuno-polymerase chain reaction assay (MIPA), was developed for the detection of Campylobacter jejuni in milk and chicken products. Target bacteria are captured from the food sample by magnetic particles coated with a specific antibody and the bound bacteria then lysed and subjected to PCR. The MIPA could detect 420 cfu g-1 of chicken after 18 h, 42 cfu g-1 after 24 h, and 4.2 cfu g-1 after 36 h enrichment. For artificially contaminated milk 63 cfu ml-1 could be detected after 18 and 24 h and 6.3 cfu ml-1 after 36 h enrichment.
Animals↗
Individual, ethnic and seasonal variability in irritant susceptibility of skin: the implications for a predictive human patch test.
Since irritants are the major cause of contact dermatitis, it is important to identify those chemicals that possess significant ability to cause skin irritation. This process must then be followed by risk assessment and risk management. Historically, animal tests have played a major rôle in this process, but human volunteer studies are of increasing importance in this field. Where the appropriate safety and ethical controls are in place, human testing can give data that identifies skin irritation hazard. To be of widest value, these human studies must not be flawed due to inter-individual, inter-ethnic or seasonal variation. We conducted a large dose-response study and studied the impact of summer and winter weather on a predictive human assay. Sodium dodecyl sulphate (SDS) was tested at 0.1%-20% in 3 national groups of approximately 100 volunteers, using 25 mm Hill Top chambers loaded with 0.2 ml, solution and applied to the upper outer arm for 4 h. Reactions were scored at 24, 48 and/or 72 h after patch application. The German and Chinese studies were completed in a few weeks under similar winter conditions, whereas the UK work was spread fairly evenly over about a 15-month period. Some relatively minor differences were observed in the dose-response curves obtained, probably due to weather conditions. The effect of the weather on the intensity, but not the pattern, of irritant reactivity was also evident in the smaller specific study that assessed reactions to SDS in summer and in winter. Whereas 45% of the panel reacted to 20% SDS in summer, 91% reacted in the winter. However, in both studies, substantial inter-individual variations in response to SDS dominated the pattern of response. When designing a human patch test to discriminate skin irritant substances from those that are of minimal effect, it is this inter-individual variability, rather than any small inter-ethnic or seasonal variation, which must be taken into account. This can be achieved by the routine inclusion of a suitable positive irritant control, which then calibrates each human volunteer panel.
Adult↗
PCR of peripheral blood for diagnosis of meningococcal disease.
Meningococcal disease is normally suspected on clinical grounds and is confirmed by isolation of Neisseria meningitidis bacteria from blood or cerebrospinal fluid or, more recently, by serology or PCR of cerebrospinal fluid. Achieving confirmation of a clinical diagnosis of meningococcal disease has become more difficult in the last few years. The pre-hospitalization administration of parenteral benzylpenicillin normally renders blood cultures sterile, and lumbar puncture is undertaken less frequently, especially in young children. We evaluated PCR for the detection of meningococcal DNA in 80 blood samples taken from patients with known or suspected meningococcal disease or from patients with other diagnoses (negative controls). Both the sensitivity and the specificity of the test were 100% for patients with confirmed cases of meningococcal disease when the blood buffy coat was used (83 to 100% sensitivity and 87 to 100% specificity with 95% confidence limits). Positive PCR results could be obtained from both blood buffy coat and serum samples. Sensitivity was unaffected by prior antibiotic treatment. PCR is a rapid, sensitive test that may be used to confirm a diagnosis of meningococcal disease by using peripheral blood samples. Introduction of this test into clinical laboratories may in some cases obviate the need for lumbar puncture to be performed on patients with suspected meningococcal disease. Our results demonstrate that a substantial number of cases of meningococcal disease are not confirmed by conventional techniques and remain undiagnosed. If the PCR test described here was widely applied, the number of cases of meningococcal disease ascertained might rise by as much as 60% greater than that recognized at present. It is likely that we are in a prevaccination era for meningococcal disease. Better case ascertainment is urgently required to assess the need for vaccines, to determine their costs and benefits, and to monitor their efficacies.
Bacteremia↗
Gene replacement by homologous recombination in Mycobacterium bovis BCG.
Gene replacement by homologous recombination is a powerful tool for fundamental studies of gene function, as well as allowing specific attenuation of pathogens, but has proved difficult to achieve for Mycobacterium tuberculosis. We have used a plasmid-based test system to demonstrate the occurrence of homologous recombination in the tuberculosis vaccine strain Mycobacterium bovis BCG, and we have successfully replaced a target gene in BCG by homologous recombination, using a shuttle plasmid. Specific inactivation of selected genes will facilitate study of virulence factors and drug resistance as well as allowing rational attenuation of M. tuberculosis for the production of new vaccines.
BCG Vaccine↗
Mycobacterial granulomatous inflammation in the ulcerated caecum of indomethacin-treated rats.
We report mycobacterial granulomatous inflammation in the ulcerated caecum of rats that received indomethacin. Two groups of male rats were treated with dietary indomethacin 3 mg/kg/day or untreated diet for 3 weeks. Six out of 8 indomethacin treated rats showed both ulceration and inflammation of the caecal mucosa. Two of the rats showing caecal ulceration also showed distinct granulomatous inflammation of the caecal mucosa and acid-fast bacilli were identified within granulomata. None of the other indomethacin treated or control rats contained acid-fast bacilli within caecal tissue sections but they were present, in the same sections, within the lumen of most rats in both groups. Polymerase chain reaction analysis identified the mycobacterial 65 kDa GroEL gene within control and granulomatous caecal tissue. In a repeat of indomethacin administration to a third group of rats, culture of both non-granulomatous caecal tissue (containing histologically identified luminal acid-fast bacilli) and faecal samples for mycobacteria was negative.
Animals↗
Varicella-associated acute supraglottitis.
Acute bacterial supraglottitis is a well-recognized pediatric entity. There are reports in the literature of numerous supraglottic infections not limited to the usual organism, Haemophilus influenzae type b. We present a case of acute supraglottitis that is associated with varicella infection.
Airway Obstruction↗
Characterization of the virulence of Mycobacterium avium complex (MAC) isolates in mice.
The virulence of different isolates of MAC was studied in naturally susceptible BALB/c mice. In preliminary experiments, MAC bacteria forming smooth transparent colonies on solid media (SmT variants) were found to be virulent for BALB/c mice, causing progressive infection; smooth opaque (SmOp) were generally avirulent, being slowly eliminated from the infected organs; and rough (Rg) variants were either avirulent or as virulent as SmT variants. We chose to compare the virulence of different isolates of MAC of different origins, studying only the SmT morphotype. Strains of MAC isolated from naturally infected animals were those that most consistently caused progressive infections. AIDS patients-derived isolates were of intermediate virulence or devoid of virulence in mice. The environmental strains were eliminated from mice or did not proliferate. Strains of MAC isolated from individuals who were not infected by HIV varied in virulence from completely avirulent to highly virulent. There was no close correlation between virulence and restriction fragment length polymorphism (RFLP) type, although all highly virulent strains were of the A/I type. There was also no correlation between virulence analysed in vivo and the ability to grow in cultured macrophages.
Animals↗
Mycobacterium tuberculosis DNA in tissue affected by sarcoidosis.
OBJECTIVE: To investigate the prevalence of Mycobacterium tuberculosis DNA in granulomatous tissues from patients with sarcoidosis and from controls matched for age, sex, and tissue by using the polymerase chain reaction. DESIGN: Single blind control trial. SUBJECTS: 16 patients with sarcoidosis who had undergone diagnostic biopsy of lung, skin, or lymph node and 16 patients with squamous cell carcinoma or Hodgkin's disease to act as controls. In addition, four lung specimens infected with M tuberculosis were included as positive controls. RESULTS: M tuberculosis DNA was present in sarcoid tissues containing granulomas from seven of the 16 patients and one of the 16 matched controls. Two of the four specimens known to be infected with M tuberculosis were positive in the controlled experiment. CONCLUSION: These figures suggest that M tuberculosis DNA is detected as readily in patients with sarcoidosis as in patients with frankly tuberculous tissues and imply that M tuberculosis may be linked to the cause of sarcoidosis.
Base Sequence↗
Construction and use of integrative vectors to express foreign genes in mycobacteria.
We have constructed a mycobacterial integrative vector by placing two copies of the insertion sequence IS900 flanking a kanamycin-resistance gene into a 'suicide' vector unable to replicate in mycobacteria. The Mycobacterium leprae gene encoding the M. leprae 18 kDa protein was cloned between the two copies of IS900 to provide expression signals. Constructs were introduced into Mycobacterium species smegmatis, vaccae and bovis BCG by electroporation and selection for kanamycin resistance. The expression of the 18 kDa gene was analysed by Western blotting. Integration of the vector into the M. smegmatis chromosome was analysed by Southern blotting. One to five copies of the vector were detected in each transformant. The SIV gag p27 gene and the foot-and-mouth disease virus VP1 140-160 epitope were successfully cloned into the 18 kDa gene and expression in M. smegmatis was obtained.
Amino Acid Sequence↗
Polymerase chain reaction for detection of Mycobacterium leprae in nasal swab specimens.
The polymerase chain reaction based on the selective amplification of a 531-bp fragment of the gene encoding the proline-rich antigen of Mycobacterium leprae was applied to nasal swab specimens from leprosy patients, occupational contacts, and endemic and nonendemic controls. To prevent false-positive amplification, we used dUTP and uracil-DNA-glycosylase in all polymerase chain reactions. False-negative reactions were detected by using a 531-bp modified template as an internal control. Amplification products were found in 55% of untreated patients, in 19% of the occupational contacts, in 12% of endemic controls, and in none of the nonendemic controls. This study strongly suggests that not only leprosy patients but also healthy persons may carry M. leprae. We concluded that polymerase chain reaction is a reliable method to detect M. leprae in nasal specimens. The method holds promise for studying the spread and transmission of M. leprae within a population.
Contact Tracing↗
Polymerase chain reaction for diagnosis of meningococcal meningitis.
Meningococcal disease is normally suspected on clinical grounds but confirmed by isolation of Neisseria meningitidis from blood or cerebrospinal fluid (CSF), or by detection of gram-negative diplococci in CSF. After parenteral antibiotics are started the isolation rate of meningococci from blood cultures drops from 50% to less than 5% and the chances of CSF being positive by culture or microscopy are also reduced. We used the polymerase chain reaction (PCR) in a blinded study to detect meningococcal DNA in 54 CSF samples from patients with meningococcal disease or from controls. The PCR primers were specific for the meningococcal insertion sequence IS1106. The sensitivity and specificity of this PCR for diagnosis of meningococcal meningitis were both 91%. Sensitivity was not affected by prior antibiotic treatment. The IS1106 PCR is a rapid and sensitive test for confirmation of the diagnosis of meningococcal meningitis.
DNA, Bacterial↗
Detection of mycobacterial DNA in sarcoidosis and tuberculosis with polymerase chain reaction.
The cause of sarcoidosis is unknown. However, the histological similarity between the disorder and tuberculosis suggests that mycobacteria might contribute to the pathogenesis of sarcoidosis. We have used the polymerase chain reaction (PCR) to detect mycobacterial DNA in clinical samples from patients with sarcoidosis. 104 patients were included in the study (62 referred for possible tuberculosis and 20 for possible sarcoidosis, and 22 control patients who had undergone bronchoscopy for other reasons). Bronchoalveolar lavage samples, bronchial washings, and tissue specimens (1 from each patient) underwent assay by PCR as well as bacteriological, histological, and cytological examination. We used two PCR reactions: in the first the complex-specific insertion sequence IS986/IS6110 was used to specifically detect DNA from Mycobacterium tuberculosis complex bacteria; in the second, conserved sequences of the mycobacterial groEL gene were used to detect DNA from mycobacteria other than M tuberculosis. The PCR was more sensitive than culture for diagnosis of tuberculosis. However, the false-positive PCR rate for M tuberculosis was 9%. M tuberculosis DNA was found in half the sarcoidosis patients, and non-tuberculosis mycobacterial DNA in a further 20%. The findings that a significant proportion of the sarcoidosis patients in this study have mycobacteria in their lungs and that most of these mycobacteria belong to M tuberculosis complex suggest an aetiological role for mycobacteria in sarcoidosis.
Base Sequence↗
First International Conference on the Pathogenesis of Mycobacterial Infections: a summary.
Two hundred scientists representing 25 countries assembled for the First International Conference on the Pathogenesis of Mycobacterial Infections, which was held in Stockholm on 27-29 June 1990. A total of 40 speeches and 80 poster presentations covered nearly all of the recent progress made in the field of mycobacterial pathogenicity. The present report is intended as a brief summary of the research results presented during the conference and focuses on findings of broad scientific interest. However, the omission of other presented findings from this report does not constitute any judgment of their scientific quality.
Antigens, Bacterial↗
Topically applied verapamil hydrochloride inhibits tuberculin-induced delayed-type hypersensitivity reactions in human skin.
Calcium channel antagonists have been reported to possess inhibitory effects on lymphocyte migration and activation in vitro, and on cell-mediated immune reactions in the skin of experimental animals. We have therefore studied the effects of topically applied 8% (w/v) verapamil hydrochloride in propylene glycol on tuberculin-induced delayed-type hypersensitivity reactions in the skin of normal human volunteers. There was significant inhibition of the tuberculin reactions by the verapamil preparation compared to vehicle controls, as determined by forearm skin fold thickness measurement and assessment of the density of mononuclear cell infiltrates in skin biopsies. The precise mechanism of action of verapamil hydrochloride remains unclear, but could include effects on T-cell migration and activation, on antigen-presenting cells, and/or on other cells. The potential for the use of topical calcium channel antagonist preparations in inflammatory skin diseases warrants further study.
Administration, Topical↗
Unilateral subungual hyperkeratosis following a cerebrovascular incident in a patient with psoriasis.
The prevalence of nail abnormalities in psoriasis is reported to be between 50 and 90% of affected adults. These include pitting, nail discoloration, onycholysis and subungual hyperkeratosis. Although nail changes may be associated with neurological disease, there are no previous reports of psoriatic nail changes following cerebrovascular events. We report the first case to our knowledge of unilateral subungual hyperkeratosis in a psoriatic patient which developed following a cerebrovascular event.
Cerebrovascular Disorders↗