ORCID Profile
0000-0003-1924-1261
Current Organisations
Elsevier BV
,
University of Adelaide
Does something not look right? The information on this page has been harvested from data sources that may not be up to date. We continue to work with information providers to improve coverage and quality. To report an issue, use the Feedback Form.
Publisher: Cambridge University Press (CUP)
Date: 12-1988
DOI: 10.1017/S0950268800029563
Abstract: The efficiency of the direct detection of Mycoplasma pneumoniae in respiratory exudates by an antigen capture, indirect enzyme immunoassay (Ag-EIA), has been compared with its detection with a cDNA probe (‘Gen-Probe assay’) directed against the specific ribosomal RNA sequences of the organism (‘ Mycoplasma pneumoniae Rapid Diagnostic System’, Gen-Probe, San Diego, California). Both assays showed excellent specificity against a range of mycoplasma species suspended in negative nasopharyngeal aspirates only M. pneumoniae and M. genitalium reacted. In experiments with graded doses of viable M. pneumoniae cells suspended in negative nasopharyngeal aspirate, the Gen-Probe assay was more sensitive than Ag-EIA detection limits were respectively 2 × 103 c.f.u./ml (3·2 × 10 5 genomes) and 2·5 × 10 4 c.f.u./ml (4 × 10 6 genomes) detection levels 10–100 times less sensitive than culture. The two assays were also tested on nasopharyngeal aspirates or sputum specimens from 90 patients with respiratory infection 67 of these were culture-or seronegative for M. pneumoniae and 23 were culture-or seropositive. Ag-EIA detected 21 (91%) of the latter but the Gen-Probe assay detected only 5 (22%). Both assays were negative with the 67 culture-/sero-negatives there were no Gen-Probe assay positive/Ag-EIA negatives. Overall, it is concluded that although Ag-EIA and the Gen-Probe assay are effective substitutes for culture as a diagnostic procedure, there is a significant problem with s les which are culture-negative and from patients who have good serological evidence of current infection. Possible reasons for the disparity between the two assays are advanced.
Publisher: Elsevier BV
Date: 02-2008
DOI: 10.1016/J.BIOS.2007.10.011
Abstract: This report describes the fabrication and successful use of the ion channel switch biosensor (ICSB) for rapid point-of-care detection of influenza A in different types of respiratory specimens. Virus culture -- regarded as the "gold standard" -- and an immunochromatographic rapid point-of-care test for influenza A virus were compared with the biosensor. The ICSB rapid test provided an objective readout within 10 min of specimen inoculation into the ICSB chamber wells, without the need for chemical or other pretreatments. Construction of the ICSB with specific antibodies also enables rapid detection and identification of appropriate influenza A subtypes.
Publisher: Elsevier BV
Date: 05-2009
DOI: 10.1016/J.JVIROMET.2008.12.016
Abstract: Four IgG(1kappa) monoclonal antibodies (mAbs) against Influenza A/Chicken/Vietnam/8/2004 (H5N1) virus are described. Three of these showed neutralizing activities against H5N1 strains from clades 1, 2 and 3 using a retroviral pseudotype or live virus microneutralization assay. In the pseudotype assay, the IC(90) neutralizing titre range was >1600-51,200, and with the microneutralization was 80> or =10,240. MAb 1C1 showed strong neutralizing activities in both assays. All four mAbs reacted specifically to the immunogen by immunohistochemical staining and to A/Hong Kong/483/1997 (H5N1) and A/Thailand/1(KAN-1)/2004 (H5N1)-infected MDCK cells by immunofluorescence. ELISA titrations of the mAbs showed specificity for H5N1 haemagglutinin (HA) and no cross-reactivity to 15 other Influenza A subtypes. Only mAbs 1C1 and the non-neutralizing 1F7 reacted with HA(1), the cleaved subunit of HA, by Western blot. These results suggest that the mAbs recognize distinct or overlapping epitopes and will be useful reagents for construction of specific rapid point-of-care assays or for therapeutic use.
Publisher: Elsevier BV
Date: 03-2011
DOI: 10.1016/J.BIOS.2010.12.018
Abstract: Sensors based on surface plasmon resonance (SPR) allow rapid, label-free, highly sensitive detection, and indeed this phenomenon underpins the only label-free optical biosensing technology that is available commercially. In these sensors, the existence of surface plasmons is inferred indirectly from absorption features that correspond to the coupling of light into a thin metallic film. Although SPR is not intrinsically a radiative process, when the metallic coating which support the plasmonic wave exhibits a significant surface roughness, the surface plasmon can itself couple to the local photon states, and emit light. Here we show that using silver coated optical fibres, this novel SPR transducing mechanism offers significant advantages compare to traditional reflectance based measurements such as lower dependency on the metallic thickness and higher signal to noise ratio. Furthermore, we show that more complex sensor architectures with multiple sensing regions scattered along a single optical fibre enable multiplexed detection and dynamic self referencing of the sensing signal. Moreover, this alternative approach allows to combine two different sensing technologies, SPR and fluorescence sensing within the same device, which has never been demonstrated previously. As a preliminary proof of concept of potential application, this approach has been used to demonstrate the detection of the seasonal influenza A virus.
Publisher: Microbiology Society
Date: 06-2007
Abstract: Trichomonas vaginalis is the cause of one of the most common types of vaginitis, trichomoniasis. The incidence of trichomoniasis in developed countries has decreased substantially during the past decade, but high prevalence of this disease can still be found in rural and remote areas of Australia. Clinical manifestations of symptomatic women are generally non-specific, but include vaginal discharge, vaginitis and irritation. T. vaginalis infection has also been linked to the increased risk of human immunodeficiency virus transmission. Current diagnosis of T. vaginalis relies on the visualization of motile organisms in a wet-mount preparation. Culture is used mainly in reference laboratories. The latter two methods require viable organisms and would not be suitable for use where transportation of specimens can be delayed. Two real-time fluorescence resonance energy transfer (FRET) hybridization probe PCR assays were used in this study to test for T. vaginalis DNA, targeting the β -tubulin and 18S rRNA genes. We tested 500 randomly selected female patients, in an STD setting, for T. vaginalis DNA. The FRET PCRs targeting the β -tubulin gene and the 18S rRNA gene detected 96 % (85/89) and 100 % (89/89) , respectively, of the positive specimens (first-void urine s le or genital swabs). Wet-mount microscopy was performed on 76 of these PCR-positive specimens and showed a sensitivity of 38 % (29/76). The prevalence, by PCR, of trichomoniasis was 18 % in this study. The two real-time PCRs developed in this study, targeting different genetic regions of the organism, provide a rapid, sensitive and specific diagnosis of T. vaginalis infection.
Publisher: American Society for Microbiology
Date: 02-1989
DOI: 10.1128/JCM.27.2.364-366.1989
Abstract: Five different enzyme immunoassays, electron microscopy, and latex agglutination (Slidex bioMerieux) were compared for the rapid detection of human rotavirus in fecal specimens. The enzyme immunoassay using rotavirus polyclonal antiserum (Dakopatts) with simple in-house modifications was shown by the use of confirmatory tests to be the most sensitive and specific procedure.
Publisher: BMJ
Date: 02-1993
DOI: 10.1136/STI.69.1.51
Abstract: To determine whether the use of urine s les from male patients can replace urethral swabs for the rapid detection of Chlamydia trachomatis by the Pharmacia EIA. The STD clinic, Adelaide, South Australia. There were two separate groups of male patients. Group A (398) patients provided urethral specimens for the EIA and culture tests. The patients in Group B (356) provided an urethral swab and a urine s le for the EIA test. The urine s les and urethral swabs were tested for the presence of C trachomatis by the Pharmacia Chlamydia EIA. In addition, the urethral swabs from Group A patients were cultured for the organism by standard cell cultures. The infected cell cultures were identified by an immunofluorescence test using a FITC-monoclonal antibody to C trachomatis (Kallestad). When the EIA was validated against culture, it showed a sensitivity of 100% and a specificity of 95% with the urethral swabs from Group A patients. The urine specimens were positive in 24% of those patients who yielded a positive EIA result in the urethral swabs. Although the EIA test on urethral swabs showed high sensitivity and specificity when validated against culture, our results showed that the use of urine s les cannot replace urethral swabs for the laboratory diagnosis of this sexually transmitted disease.
Publisher: Springer Science and Business Media LLC
Date: 05-1997
Abstract: HIV infection causes dysregulation of cytokine gene expression in CD4+ T cells of the infected host. Azidothymidine (AZT) inhibits HIV replication by blocking reverse transcription. Using a one-stop cell-to-cell HIV infection model, we have investigated the expression of several key cytokines in HIV infected T cells in the absence or presence of AZT treatment. Acute HIV infection of T cells resulted in dramatic down regulation of the expression of IL-2 and INF-gamma mRNA. While beta-actin mRNA levels remained constant in both AZT-free and AZT treated cultures after HIV infection, it was found that AZT blocked the down regulation of IL-2 mRNA and INF-gamma mRNA in CD4+ T cells acutely infected with HIV.
Publisher: Elsevier BV
Date: 04-2021
Publisher: Cambridge University Press (CUP)
Date: 06-1994
DOI: 10.1017/S095026880005130X
Abstract: The distribution of adenovirus types in faecal s les of patients with suspected viral gastroenteritis from South Australia was determined during the 12-month period, July 1991–June 1992. There were 3299 s les tested and 226 (6·9%) were positive for adenovirus by enzyme immunoassay. Of these 226 s les. 154 (68%) were typed directly using virus DNA extracted from the faecal s les according to the Sma I, Hind III and Bst EII restriction patterns and Southern hybridization analysis with pooled viral genomic DNA probes. In this group, 86% of the s les were from patients who were 3 years of age. Enteric adenovirus types 40 and 41 accounted for 20 and 40% respectively, of these s les, and types 1, 2, 3, 5, 6, 7 and 31 comprised the remainder. Type 40 was detected mainly in the winter and spring periods, and type 41 predominated in the autumn period. The majority of the non-enteric types were found during the late winter and spring periods.
Publisher: American Society for Microbiology
Date: 12-1997
DOI: 10.1128/JCM.35.12.3355-3357.1997
Abstract: Two commercially available nucleic acid-based tests, ligase chain reaction (LCR Abbott Laboratories) and PCR (Roche Diagnostics), for the detection of Chlamydia trachomatis in male and female urine s les were compared with culture and enzyme immunoassay (EIA) (Microtrak Syva) for C. trachomatis detection in genital s les. The s les were collected from 1,005 patients who attended a sexually transmitted disease clinic. In this study population, the prevalence of the infection was 4%. Specimens which were reactive in any of the tests were retested with a different PCR test using primers directed against the major outer membrane protein gene. With a "gold standard" of a positive culture, or any other positive test result if it was confirmed by an independent test, the Roche PCR (95% sensitive, 99.9% specific) was more sensitive than the LCR (75% sensitive, 100% specific) (chi2, P 0.0001) while both tests were more sensitive than culture (58% sensitive, 100% specific) or EIA (45% sensitive, 100% specific) (chi2, P 0.001). The Roche PCR and Abbott LCR tests of urine identified 65% and 30% more positive patients, respectively, than did testing by culture of urethral or cervical specimens. Nucleic acid testing of urine specimens for C. trachomatis is a more sensitive and convenient method for the detection of genital infection.
Publisher: Wiley
Date: 12-1993
Abstract: An outbreak of adenovirus type 4 conjunctivitis occurred in South Australia between April and November 1992. Eye swabs were submitted by general practitioners and ophthalmologists who had seen patients with clinical conjunctivitis or keratitis. Apart from interfamilial spread, there were no other common epidemiological factors. Adenovirus was isolated from the eye swabs of 38 patients. Isolates were typed by neutralisation tests and restriction endonuclease cleavage patterns and found to be adenovirus type 4. This report serves to illustrate an infrequent cause of epidemic conjunctivitis, namely adenovirus type 4. There was no demonstrable focus of the outbreak.
Publisher: Elsevier BV
Date: 05-1993
DOI: 10.1016/0166-0934(93)90036-Q
Abstract: Primary Monkey Kidney (PMK) epithelial cells or egg inoculation have been traditionally used for the culture of influenza and parainfluenza viruses. The high cost and variability of obtaining high quality PMK cells prompted us to investigate the use of other cell strains for the growth of these viruses. For this study we investigated three cell lines viz. MDCK, MEK and LLC-MK2 for the culture of influenza A and B and parainfluenza 1, 2 and 3 viruses. Clinical specimens were spun onto cell monolayers in microtitre wells. The growth of these viruses was then identified by specific antibodies in an enzyme immunoassay (EIA). The LLC-MK2 and MDCK cell lines were found to provide optimal growth of parainfluenza and influenza viruses respectively. During the period from November, 1990 to July, 1992, 6501 respiratory specimens were tested. There were 100 influenza A, 36 influenza B and 261 parainfluenza virus isolates. The influenza isolates were further subtyped by the WHO Influenza Reference Centre. The use of these cell lines and the EIA provided an effective method for the routine culture of these viruses.
Publisher: Elsevier BV
Date: 05-2004
Publisher: Elsevier BV
Date: 07-1998
DOI: 10.1016/S0928-0197(98)00026-9
Abstract: Traditionally, primary monkey kidney (PMK) epithelial cells have been one of the more widely used cell types for the isolation of enteroviruses from clinical s les. For the isolation of coxsackie A viruses, intraperitoneal inoculation of newborn mice has been used in some laboratories. With the discontinued availability of PMK epithelial cells and the reported growth of coxsackie A viruses in rhabdomyosarcoma cells (RD), we compared the use of the latter cell line with our routinely used microwell cell cultures. Microwell cell cultures of buffalo green monkey epithelial cell line (BGM), human lung carcinoma epithelial cell line (A549) and human embryonic lung (HEL) fibroblasts were compared with RD cell line for the isolation of enteroviruses from clinical s les. A total of 39 enteroviruses was isolated from 3517 specimens. The HEL fibroblasts yielded 28 (72%) enteroviruses, followed by A549 (25 isolates, 64%), BGM (23 isolates, 59%) and RD cells (18 isolates, 46%). All isolates which grew in RD cells showed specific cytopathic effects in one or more of the other inoculated cell cultures. Quantitative determinations (TCID50) with five different enteroviruses showed that the HEL fibroblasts and RD cell line to be the most sensitive cell types, followed by BGM and A549 cell lines. However, integrity of the inoculated cell monolayers was best with HEL fibroblasts and A549 cells but morphology was not optimal with RD cells during the incubation period of 14 days.
Publisher: Elsevier BV
Date: 02-2000
DOI: 10.1016/S1386-6532(99)00066-9
Abstract: Extraction of viral nucleic acids from serum s les is widely used in diagnostic pathology tests. However, the heterogeneous nature of non-serum s les may contribute to variations in the yields of viral nucleic acids with different extraction methods and specimen types. Six different methods were compared for optimal extraction of viral DNA or RNA from four types of non-serum specimens. The DNA viruses used were herpes simplex virus and cytomegalovirus. The RNA viruses were poliovirus, rotavirus and small round structured virus. The specimens used were from respiratory, genital, faecal and peripheral blood mononuclear cell s les. The extracted nucleic acids were lified by PCR and detected in an enzyme immunoassay using digoxygenin-labelled licons. For extraction of viral DNA, the phenol-chloroform method yielded the highest amount of DNA as judged by endpoint titration. The three methods compared for extraction of viral RNA used guanidine isothiocyanate and the QiaRNA kit was shown to yield the highest amount of viral RNA.
Publisher: Oxford University Press (OUP)
Date: 10-1994
Publisher: SPIE
Date: 13-05-2011
DOI: 10.1117/12.883825
Publisher: Elsevier BV
Date: 05-1998
DOI: 10.1016/S0928-0197(98)00003-8
Abstract: The laboratory diagnosis of herpes simplex infection may require rapid (direct) tests, as well as cell cultures, for detection of the virus in clinical s les. The quantity of virus present in clinical s les is variable and this may depend on the period from onset of rash. In addition, not all patients may show obvious symptoms with this infection. The successful culture of herpes simplex virus requires prompt transportation after collection of the specimen as the virus is easily inactivated. Hence, rapid and culture tests would enable detection of non-viable and viable viruses. We describe the rapid detection of HSV by EIA directly in various clinical s les using commercially available polyclonal sera. In addition specimens were inoculated in microwell cell cultures and 4 days post inoculation the culture fluids were tested for HSV and subtyped by a similar EIA (culture lified EIA). The direct EIA showed an endpoint detection of 100 TCID50/ml, sensitivity of 92% (all specimen types) and specificity of 100%. The direct EIA sensitivity was 97% in non-genital specimens and 88% in genital specimens. The culture lified EIA showed a sensitivity of 95% compared to all confirmed HSV positive s les. The results of the HSV rapid tests were available within 24 h from receipt of specimens. Specimens which were culture negative/direct EIA positive were confirmed by blocking antisera. Culture positive specimens which were direct EIA negative were confirmed by subtyping of the virus.
Publisher: Oxford University Press (OUP)
Date: 08-1993
DOI: 10.1093/CLINIDS/17.SUPPLEMENT_1.S90
Abstract: Efforts to improve laboratory diagnostic methods for infection due to Mycoplasma pneumoniae have involved the use of a cell-sheet culture method and a modified indirect hemagglutination method for IgM antibody, while direct detection of mycoplasma has employed antigen capture-enzyme immunoassay (Ag-EIA) and polymerase chain reaction (PCR) lification of sequences within the P1 and 16S ribosomal RNA genes and quantification of the lified DNA by dot blot hybridization (DBH). Cell-sheet culture was slightly more sensitive and more rapid than culture with cell-free diphasic medium. Indirect hemagglutination detection of IgM antibody to M. pneumoniae was more sensitive than CF and EIA for detection of IgM antibody to mycoplasma. Ag-EIA gave a rapid and reasonably sensitive indication of infection and correlated well with a serological response of patients indicating a current infection. PCR-DBH was a highly sensitive substitute for culture of mycoplasma. Both Ag-EIA and PCR-DBH require confirmation by assessment of serological response to verify that the infection is current and that positive results of PCR-DBH, in particular, are not the result of continuing carriage of the organism from a previous infection, unrelated to the current episode under investigation.
Publisher: BMJ
Date: 11-1984
Abstract: One hundred and eight children presenting with Mycoplasma pneumoniae infection were assessed during the acute illness and followed for three years. The incidence of wheezing with the acute infection (40%) was greater than expected in a normal childhood population. The initial illness precipitated wheezing for the first time in some subjects but others wheezed only with the acute illness. In non-asthmatic subjects significant bronchodilator responsiveness was present one month after infection. Children given erythromycin during the first seven days of their illness had a significantly shorter fever duration compared with those treated inappropriately. No significant effects of treatment were noted on pulmonary function three years later but non-asthmatic children had abnormal mean forced expiratory volume in one second and forced expiratory flow after 50% of the expired vital capacity compared with 64 healthy controls. These findings indicate impaired function three years after initial infection.
Publisher: Elsevier BV
Date: 05-1995
DOI: 10.1016/S0140-6736(95)92579-1
Abstract: Eucalyptus camaldulensis EcDQD/SDH2 and 3 combine gallate formation, dehydroquinate dehydratase, and shikimate dehydrogenase activities. They are candidates for providing the essential gallate for the biosynthesis of the aluminum-detoxifying metabolite oenothein B. The tree species Eucalyptus camaldulensis shows exceptionally high tolerance against aluminum, a widespread toxic metal in acidic soils. In the roots of E. camaldulensis, aluminum is detoxified via the complexation with oenothein B, a hydrolyzable tannin. In our approach to elucidate the biosynthesis of oenothein B, we here report on the identification of E. camaldulensis enzymes that catalyze the formation of gallate, which is the phenolic constituent of hydrolyzable tannins. By systematical screening of E. camaldulensis dehydroquinate dehydratase/shikimate dehydrogenases (EcDQD/SDHs), we found two enzymes, EcDQD/SDH2 and 3, catalyzing the NADP
Publisher: Cambridge University Press (CUP)
Date: 12-1988
DOI: 10.1017/S0950268800029551
Abstract: Direct and indirect antigen capture enzyme immunoassays (Ag-EIA) have been developed for the detection of Mycoplasma pneumoniae in nasopharyngeal aspirates or sputum from respiratory infection. The sensitivity of the two Ag-EIA were similar, but the indirect method using polyclonal rabbit and guinea-pig antisera was more convenient. The Ag-EIA had a detection limit of 10 4–4·5 colony-forming units/ml of s le. It was specific for M. pneumoniae and gave a low level response with M. genitalium. There were no cross-reactions with 10 other species of mycoplasmas. Tests with a wide range of bacteria and chlamydia group antigen, representing agents sometimes found in the respiratory tract, were also negative. At the current level of development, the Ag-EIA detected about 90% of specimens that were also positive for culture 43% of specimens from culture-negative-seropositive patients gave a positive result. The overall pattern of results indicated that while antigen detection is a quick and effective substitute for the slow culture method, serological examination for specific IgM antibody is also necessary to give a complete diagnostic coverage.
Publisher: Microbiology Society
Date: 09-2018
DOI: 10.1099/JMM.0.000813
Abstract: Respiratory tract infections are a major cause of global morbidity and mortality. Pneumonia is the ninth leading cause of mortality in Sri Lanka. Atypical pathogens cause about one-fifth of community-acquired pneumonia, while Mycoplasma pneumoniae accounts for about 50 %. This study aimed to determine the seroprevalence of M. pneumoniae respiratory tract infections in Sri Lanka while attempting to understand the relationships between the serology and PCR. Paired sera from 418 adult patients (pneumonia, n=97 bronchitis, n=183 pharyngitis, n=138) and 87 healthy controls were studied. IgM, IgG and IgA antibodies were tested by M. pneumoniae enzyme-linked immunosorbent assay (ELISA). Positive IgM and or IgG seroconversion was considered to be seropositive. M. pneumoniae DNA were tested by PCR in age and gender-matched seropositives and seronegatives. M. pneumoniae IgG was in 14.4 % (14/97), 6.0 % (11/183) and 1.5 % (2/138) of pneumonia, bronchitis and pharyngitis patients, respectively, whilst IgM was in 6.2 % (6/97), 1.1 % (2/183) and 0 % (0/138), respectively. Amongst the pneumonia seropositives, 64.7 % (11/17) showed IgG alone, 17.5 % (3/17) showed IgM alone and 17.5 % (3/17) showed IgM and IgG. Amongst the bronchitis seropositives, 84.6 % (11/13) had IgG alone and 15.4 % (2/13) had IgM alone. In the pharyngitis seropositives, only IgG was detected 100 % (2/2). M. pneumoniae DNA was in 52.2 % (12/23) of seropositives and 15.4 % (4/26) of seronegatives. In pneumonia or bronchitis patients, specific DNA was in 77.8 % (7/10) and 50 % (6/12) of patients, respectively. M. pneumoniae DNA was not found in pharyngitis patients. Of the seropositive PCR-negative pneumonia patients, 66.7 % (2/3) showed IgG alone and 33.3 % (1/3)showed IgM alone. In bronchitis patients, 83.3 % (5/6) showed IgG alone and 16.7 % (1/6) showed IgM alone. Of the seronegative PCR-positive patients, 16.7 % (2/12) had pneumonia and 18.2 % (2/11) had bronchitis. The serological evidence for M. pneumoniae infection in Sri Lanka comprised the following prevalences: 17.5 % (17/97), 7.1 % (13/183) and 1.4 % (2/138) in adults with pneumonia, bronchitis or pharyngitis, respectively. M. pneumoniae DNA was in 52.2 % (12/23) of seropositives and 15.4 % (4/26) of seronegatives. IgG was predominant in PCR positives and negatives.
Publisher: Microbiology Society
Date: 05-2011
Abstract: A universal influenza vaccine that does not require annual reformulation would have clear advantages over the currently approved seasonal vaccine. In this study, we combined the mucosal adjuvant alpha-galactosylceramide (αGalCer) and peptides designed across the highly conserved influenza precursor haemagglutinin (HA 0 ) cleavage loop as a vaccine. Peptides designed across the HA 0 of influenza A/H3N2 viruses, delivered to mice via the intranasal route with αGalCer as an adjuvant, provided 100 % protection following H3N2 virus challenge. Similarly, intranasal inoculation of peptides across the HA 0 of influenza A/H5N1 with αGalCer completely protected mice against heterotypic challenge with H3N2 virus. Our data suggest that these peptide vaccines effectively inhibited subsequent influenza A/H3N2 virus replication. In contrast, only 20 % of mice vaccinated with αGalCer-adjuvanted peptides spanning the HA 0 of H5N1 survived homologous viral challenge, possibly because the HA 0 of this virus subtype is cleaved by intracellular furin-like enzymes. Results of these studies demonstrated that HA 0 peptides adjuvanted with αGalCer have the potential to form the basis of a synthetic, intranasal influenza vaccine.
Publisher: Cambridge University Press (CUP)
Date: 12-1989
DOI: 10.1017/S0950268800031010
Abstract: The indirect haemagglutination (IHA) test was compared with the complement-fixation (CF) test for the measurement of antibodies to Mycoplasma pneumoniae . A modification of the IHA was used to measure M. pneumoniae IgM antibodies. Sera were obtained from various groups of patients who were either culture or antigen positive for M. pneumoniae in nasopharyngeal aspirates or who had fourfold or greater increase in CF antibody or a titre ≥ 320.
Publisher: Elsevier BV
Date: 1997
DOI: 10.1080/00313029700169135
Abstract: Sputum s les from adult patients are routinely used for bacteriological tests, but not for the diagnosis of viral/mycoplasmal infections. We examined 511 sputum s les submitted for bacterial tests from patients at the Royal Adelaide Hospital. Each specimen was tested directly (and after six days of cell culture lification) for antigens to influenza A and B, parainfluenza 1, 2 and 3, adenovirus, respiratory syncytial virus (RSV) and Mycoplasma pneumoniae. Respiratory viruses or M. pneumoniae were found in 11% of all specimens but were most common (14%) in sputa reported as containing only "oral flora". Respiratory virus or M. pneumoniae infection was significantly more common in medical patients (12%) than in surgical patients (5%), and was most common in oncology (hematology/radiotherapy) patients (25%). Influenza A and RSV were equally common in medical patients, while RSV was the most frequent isolate in oncology patients. Respiratory viral infection is an underdiagnosed condition in adults, particularly the immunocompromised, which can be successfully diagnosed by virological examination of sputum.
Publisher: Informa UK Limited
Date: 1995
DOI: 10.3109/00365549509047055
Abstract: A study was undertaken to determine the incidence of chlamydial respiratory infection in paediatric patients during a 12-month period by antigen detection. Nasopharyngeal aspirates (NPA) from 514 patients were screened for genus-specific chlamydia antigen using the Pharmacia Chlamydia enzyme immunoassay (EIA) and Kallestad immunofluorescence (IF) assays. EIA screen-positive s les were confirmed by specific blocking antibody. Specimens which were EIA positive or IF positive were cultured for chlamydia. The NPAs from 7 patients were positive in the EIA and IF assays. Four of these patients were culture positive for chlamydia. Our results showed that the incidence of chlamydia respiratory infection by antigen detection was 1.4% or 0.8% if confirmed by culture.
Publisher: Elsevier BV
Date: 12-1994
DOI: 10.1016/0166-0934(94)90166-X
Abstract: A composite EIA, using 8-well microstrips, was used for the rapid detection of seven respiratory viruses and M. pneumoniae. The viruses included influenza A and B, parainfluenza 1, 2 and 3, adenovirus and respiratory syncytial virus. During the 61 month period--June 1988 to June 1993--17326 respiratory specimens, submitted from three states, were tested by this EIA. The specimens were mainly from a paediatric population (hospitals and private physicians). RSV was the predominant virus detected, followed by adenovirus, parainfluenza 3, M. pneumoniae, influenza A, parainfluenza 2, influenza B and parainfluenza 1. The use of blocking antibodies confirmed the identification of the agents, in particular with s les showing absorbance values greater than the cutoff with more than one infectious agent. Different methods for processing specimens in order to obtain a uniform suspension, and interpretation of non-specific reactions, are discussed. The assays showed an average sensitivity of 85% and specificity of 99%, compared to virus culture. This EIA system provided an efficient method for the rapid diagnosis of viral and mycoplasmal infections in a busy diagnostic laboratory.
Publisher: Oxford University Press (OUP)
Date: 25-06-2009
DOI: 10.1093/QJMED/HCP077
Abstract: Our previous studies of persistence of Coxiella burnetii in humans after an initial acute Q fever infection revealed raised, maintained antibody levels and low levels of coxiella genomic DNA at the age of 5 years from onset in Australian patients and at 12 years in patients in the 1989 Birmingham UK Q fever outbreak. Attempts to isolate the coxiella in standard cell culture and susceptible mice by serial passage of PCR positive PBMC and bone marrow were negative. To retest PCR positive patient s les by more sensitive methods for viable coxiellas and for the coxiella cell components of antigen and specific lipopolysaccharide (LPS). To re-interpret the previous results in the light of the new information. To review the pertinent literature for a concept of an immuno-modulatory complex generated by the current studies. Laboratory case study. Stored patient s les were inoculated into SCID mice that were followed for 60 days. Mouse spleen and liver s les were then examined by PCR assay for targets in the COM1 and IS1111a sequences and for antigens by IFA with a polyclonal rabbit antiserum to C. burnetii Phase 1 and a monoclonal antiserum to Phase 1 LPS (details O. Sukocheva et al., unpublished data). All specimens, including a recently excised heart valve from a Birmingham patient with late developing endocarditis, were infection negative in SCID mice. Dilutions of SCID mouse spleen and liver homogenates titrated in PCR assays were negative at dilutions attained by control mice inoculated with an endpoint dilution of a viable prototype strain of C. burnetii. Sections of the spleens from all specimens showed a complex of coxiella antigen-LPS by IFA. DISCUSSION/REVIEW: We advance a concept of long-term persistence of a non-infective, non-biodegraded complex of coxiella cell components with its antigens and specific LPS [so called Immunomodulatory complex (IMC)] associated with traces of genomic DNA that signalled its presence in our earlier studies. The IMC's survival in patients for at least 12 years, and in one patient for 70 years implies a capacity for serial passage in macrophages with effective down-regulation of their biodegrading functions. The review assesses the compatibility of the IMC concept in relation to cogent literature on C. burnetii interactions with macrophage and cell-mediated immunity. Some remaining gaps in our knowledge of the organ sites and duration of carriage of viable coxiellas after initial infection are also identified.
Publisher: Springer Science and Business Media LLC
Date: 18-04-2016
Publisher: Cambridge University Press (CUP)
Date: 12-1992
DOI: 10.1017/S0950268800050512
Abstract: Direct detection assays for Mycoplasma pneumoniae were established by PCR lification of short sequences within the foot protein/adhesin (P1) gene and the 16S ribosomal RNA gene. Specificity and sensitivity was excellent, no hybridization was observed with M. genitalium and other human Mycoplasma species. In nose and throat washings from subjects with respiratory infection a pattern of high counts (c.f.u./ml) of M. pneumoniae (deduced from the amount of lified PCR product), and a positive antigen capture assay, was found in 83% of subjects with serological evidence of current infection with M. pneumoniae . A small proportion of subjects with serological patterns suggesting infection in the more distant past had positive PCR assays. This was considered to represent either persistence of the organism from a previous infection or perhaps transient carriage during a reinfection, without substantial change in antibody response. PCR-based assay of M. pneumoniae offers a powerful, rapid, and sensitive substitute for culture of the mycoplasma. Antigen capture, while less sensitive than PCR, offers the advantage that it is more often positive with s les from current infection and requires less stringent laboratory organization to contain false positive results. We conclude however that the laboratory diagnosis of a chosen clinical episode should not rest on the PCR or Ag-EIA assays alone, but must also include antibody assays to confirm whether infection is current or represents persistence from past exposure.
Publisher: Elsevier BV
Date: 05-1998
Abstract: Analogous to transmission of human T-cell leukemia virus type 1 (HTLV-1) in vivo, an in vitro cell-to-cell infection model was established by coculturing MT-2 cells as virus donors and HUT78 cells as recipients. At a donor:recipient ratio of 1:2, cell fusion occurred and a new round of HTLV-1 genome replication was initiated in the cocultured cells. Newly synthesized unintegrated viral DNA was detected by Southern blot within 4-8 h and then increased between 8 and 48 h following cell mixing. The most dominant species of unintegrated viral DNA was 3.7 kb in size which hybridized to a full-length HTLV-1 DNA probe but not to a Kpnl viral DNA fragment that is absent from a defective proviral genome that has been previously identified in MT-2 cells. Northern blot analysis showed large amounts of viral RNA in the virus donor cells and in the cocultured cells, with a 3.4-kb species being the most abundant. This 3.4-kb RNA gave a pattern identical to that of the 3.7-kb unintegrated viral DNA in hybridization studies using the two probes. It seems likely that the unspliced RNA transcript from the defective proviral genome in MT-2 cells was effectively reverse transcribed upon initiation of cell-to-cell viral transmission to susceptible HUT78 cells. Despite active de novo reverse transcription, however, viral RNA levels remained unchanged following cell-to-cell transmission of HTLV-1 infection and no viral antigen production could be attributed to the newly initiated round of viral genome replication. As an abortive infection model this simple cell-to-cell infection system warrants more detailed study as it has the potential to provide reliable information regarding the early events in HTLV-1 transmission and infection.
Publisher: Springer Netherlands
Date: 2010
Publisher: Springer Science and Business Media LLC
Date: 10-1998
Abstract: Using a one-step model for cell-to-cell transmission of HIV infection we have identified two distinct phases of HIV RNA synthesis. The first phase (4 h-12 h p.i.) was marked by an increase in only the full-length 9 kb RNA, while the second phase (24 h p.i. onwards) comprised a significant increase in the levels of all three species of viral RNA. We now report that while the continual presence of actinomycin D at 50 micrograms/ml abolished all detectable viral nucleic acid synthesis when virus donor H3B cells were pre-treated with 50 micrograms/ml of actinomycin D (AmD), washed free of unbound drug (a procedure which inhibited > 99% of total cellular RNA transcription) then mixed with untreated recipient Hut 78 cells, normal amounts of full length linear unintegrated viral DNA were produced and the first phase of viral RNA transcription was unaffected. Similarly, when both the virus donor cells and recipient cells were arrested in the late G1 phase of the cell cycle by aphidicolin and then mixed, linear unintegrated viral DNA was the major viral DNA species produced. The appearance of circular viral DNA and progeny virus was inhibited, but the first phase of induced viral RNA synthesis was unaffected. When AZT was added at 2 h or 4 h after cell-cell mixing, the level of 9 kb RNA detected was significantly lower, corresponding to reduction in the level of viral DNA. These and previous results indicate that the template for the first phase of viral RNA synthesis was likely to be newly synthesized, linear unintegrated viral DNA and not pre-existing proviral DNA present in the donor cells. Taken together, our results suggest that there exists a yet to be fully characterized pathway of concurrent viral DNA and RNA synthesis early after cell to cell transmission of HIV infection.
Publisher: Cambridge University Press (CUP)
Date: 04-1991
DOI: 10.1086/646330
Abstract: To define the extent of shedding of respiratory viruses and Mycoplasma pneumoniae among a population of pediatric patients admitted to the hospital during a winter epidemic period and to identify nosocomial infections within this population. An open, prospective survey of patients admitted to three wards (General Medical, Respiratory Infectious, and Infectious Diseases) of a pediatric hospital during a defined three-month period. All patients with medical, respiratory, and infectious conditions admitted to three wards of the Adelaide Children's Hospital had nasopharyngeal aspirations performed at the time of admission with the purpose of documenting viral and M pneumoniae shedding. Patients were monitored daily for the development of symptoms of respiratory infection or new symptoms of respiratory disease. Such patients underwent a further nasopharyngeal aspiration for the purpose of diagnosing hospital-acquired infection. Nasopharyngeal aspirations were obtained from 601 patients. Forty-seven percent of asymptomatic patients were positive for a respiratory virus or M pneumoniae, and 61% of patients with respiratory symptoms were also positive. Gastroenteritis patients shed viruses in 66% of cases. Respiratory symptoms were initially overlooked by admitting physicians but subsequently identified in 110 cases, and 46% of these were found to be positive for a respiratory virus or M pneumoniae. There were 18 possible hospital acquired infections among the 293 initially virus-negative patients. Multiple isolates were obtained from a substantial number of patients, especially those with respiratory symptoms. A substantial proportion of all patients admitted to a pediatric hospital during winter represent a potential source of infection, and strict infection control measures should be enacted to limit the spread of these infections.
Publisher: American Society for Microbiology
Date: 05-2010
DOI: 10.1128/JVI.02508-09
Abstract: We previously demonstrated that a single dose of nonadjuvanted intranasal γ-irradiated influenza A virus can provide robust protection in mice against both homologous and heterosubtypic challenges, including challenge with an H5N1 avian virus strain. We investigated the mechanism behind the observed cross-protection to define which arms of the adaptive immune response are involved in mediating this protection. Studies with gene knockout mice showed the cross-protective immunity to be mediated mainly by T cells and to be dependent on the cytolytic effector molecule perforin. Adoptive transfer of memory T cells from immunized mice, but not of memory B cells, protected naïve recipients against lethal heterosubtypic influenza virus challenge. Furthermore, γ-irradiated influenza viruses induced cross-reactive Tc-cell responses but not cross-neutralizing or cross-protective antibodies. In addition, histological analysis showed reduced lung inflammation in vaccinated mice compared to that in unvaccinated controls following heterosubtypic challenge. This reduced inflammation was associated with enhanced early recruitment of T cells, both CD4 + and CD8 + , and with early influenza virus-specific cytotoxic T-cell responses. Therefore, cross-protective immunity induced by vaccination with γ-irradiated influenza A virus is mediated mainly by Tc-cell responses.
Publisher: Public Library of Science (PLoS)
Date: 18-07-2019
Publisher: Springer Science and Business Media LLC
Date: 10-2001
Abstract: A one-step cell-to-cell transmission model of human immunodeficiency virus (HIV) infection was used to study viral DNA integration in the early phase of viral replication. Co-culturing H3B cells as virus donors with CD4+ Hut78 recipient cells in a ratio of 1:4 produced a synchronous, one-step viral infection with de novo synthesis of unintegrated HIV DNA within 4 h p.i., which subsequently integrates in the host genomic DNA to form provirus. To study the kinetics of viral DNA integration, cellular chromosomal DNA was isolated at different times after co-culturing and extensive electrophoresis was used to remove residual unintegrated viral DNA. Removal of contaminating, unintegrated viral DNA in the purified chromosomal DNA fraction was confirmed by various experiments. When purified chromosomal DNA (free of contaminating unintegrated viral DNA)--from the mix of acutely infected cells--was digested with restriction enzymes KpnI, BamHI or PstI and analysed by Southern blot hybridization, integration of viral DNA into chromosomal DNA was first observed at 8 h p.i. and was essentially complete by 72 h p.i. In addition, evidence was found for a relatively stable, partially integrated HIV DNA structure within the chromosomal DNA, that was first detectable at 8 h p.i. and did not become fully integrated until 72 hours post infection.
Publisher: American Society for Microbiology
Date: 06-1990
DOI: 10.1128/JCM.28.6.1458-1459.1990
Abstract: A premarket trial of the Becton Dickinson Directigen respiratory syncytial virus membrane-based enzyme immunoassay compared the test with virus isolation for the detection of respiratory syncytial virus in 583 nasopharyngeal aspirates. After modification, the Directigen test showed a sensitivity of 83% and a specificity of 90%. It offers the potential for an efficient bedside test--without the need for any equipment--for the diagnosis of respiratory syncytial virus infection and requires only a 0.25-ml s le volume. However, for optimum reliability, freezing-thawing of s les and access to a confirmatory test were shown to be necessary.
Publisher: Public Library of Science (PLoS)
Date: 20-03-2015
Publisher: Elsevier BV
Date: 02-2018
DOI: 10.1016/J.JVIROMET.2017.10.022
Abstract: Inhibition of viral replication by icIgA antibodies has only been observed with in vitro studies using epithelial cell lines in transwell cultures. This effect appears to involve an interaction between polymeric immunoglobulin A (pIgA) and viral particles within an intracellular compartment, since IgA is transported across polarized cells. Polyclonal guinea pig antisera against purified influenza A virus and mouse antisera prepared against Influenza A/H3N2 hemagglutinin (HA The above findings in clinical specimens would contribute strongly to our understanding of the mechanisms and kinetics of icIgA neutralization in relation to viral entry and early replication steps of mucosal viral infections. A rapid, objective and sensitive assay - by ex vivo enumeration of respiratory epithelial cells that have co-localized influenza virus and icIgA - would contribute to further mucosal immunity studies and inform the design of more effective vaccines against influenza and other viral infections transmitted via the mucosal route e.g. respiratory syncytial virus, rotavirus.
Publisher: Springer Science and Business Media LLC
Date: 2002
Abstract: Prompt laboratory diagnosis of Herpes simplex virus (HSV) infection facilitates patient management and possible initiation of antiviral therapy. In our laboratory, which receives various specimen types for detection of HSV, we use enzyme immunoassay (EIA) for rapid detection and culture of this virus. The culture of HSV has traditionally been accepted as the diagnostic 'gold standard'. In this study, we compared the use of real time PCR (LightCycler) for lification, detection and subtyping of specific DNA with our in-house developed rapid and culture tests for HSV. The LightCycler PCR (LC-PCR) detected and subtyped HSV in 99% (66/67) of HSV positive specimens, compared to 81% (54/67) by rapid antigen EIA or 57% (36/63) by culture. A specimen was considered positive when two or more tests yielded HSV identifications or was culture positive. Discordant results were confirmed with an in-house developed PCR-ELISA or DNA sequence analysis. The typing results obtained with the LC-PCR and by culture lified test were completely concordant. This study showed that the LC-PCR provided a highly sensitive test for simultaneous detection and subtyping of HSV in a single reaction tube. In addition to increased sensitivity, the LightCycler PCR provided reduced turn-around-times (2 hours) when compared to enzyme immunoassay (4 hours) or culture (4 days).
Publisher: Wiley
Date: 10-01-2023
DOI: 10.1111/SJI.13253
Abstract: Virus neutralization at respiratory mucosal surfaces is important in the prevention of infection. Mucosal immunity is mediated mainly by extracellular secretory immunoglobulin A (sIgA) and its role has been well studied. However, the protective role of intracellular specific IgA (icIgA) is less well defined. Initially, in vitro studies using epithelial cell lines with surface expressed polymeric immunoglobulin receptor (pIgR) in transwell culture chambers have shown that icIgA can neutralize influenza, parainfluenza, HIV, rotavirus and measles viruses. This effect appears to involve an interaction between polymeric immunoglobulin A (pIgA) and viral particles within an intracellular compartment, since IgA is transported across the polarized cell. Co‐localization of specific icIgA with influenza virus in patients' (virus culture positive) respiratory epithelial cells using well‐characterized antisera was initially reported in 2018. This review provides a summary of in vitro studies with icIgA on colocalization and neutralization of the above five viruses. Two other highly significant respiratory infectious agents with severe global impacts viz. SARS‐2 virus (CoViD pandemic) and the intracellular bacterium— Mycobacterium tuberculosis —are discussed. Further studies will provide more detailed understanding of the mechanisms and kinetics of icIgA neutralization in relation to viral entry and early replication steps with a specific focus on mucosal infections. This will inform the design of more effective vaccines against infectious agents transmitted via the mucosal route.
Publisher: SAGE Publications
Date: 2013
Abstract: When establishing animal models of viral respiratory infection, the optimal dose and route of delivery are critical to ensure reproducible outcomes. The mouse model for influenza infection is widely used due to the small animal size and simplicity of viral inoculation. During establishment of a mouse model of influenza A infection we observed a marked shift in morbidity when identical influenza A inoculum doses were delivered in less than 35 μL. We show for the first time that mice challenged with a 25 μL inoculum volume readily recovered following infection with an infectious dose of influenza A virus that was fatal when inoculated in 35 or 50 μL volumes.
Publisher: Microbiology Society
Date: 08-1994
DOI: 10.1099/0022-1317-75-8-1917
Abstract: H3B cells, a laboratory clone of H9 cells persistently infected with the HTLV-IIIB strain of human immunodeficiency virus (HIV), contained significant levels of cell-associated reverse transcriptase (RT) activity measured by in vitro assays using either exogenous or endogenous templates. The cell-associated RT activity detected using exogenous template was almost wholly in a soluble (non-sedimentable) form whereas endogenous activity sedimented as a particulate structure associated with viral RNA. Despite this, H3B cells did not contain episomal HIV DNA detectable by Southern blot, indicating that in vivo reverse transcription was not occurring to any significant extent in these cells. However, when susceptible HUT 78 cells were infected by co-cultivation with H3B cells, dramatic synthesis of episomal HIV DNA occurred. Concurrently with this de novo initiation of reverse transcription, however, we found no detectable change in intracellular levels or cleavage profiles of immunoprecipitable RT polypeptides. Finally, actinomycin D pre-treatment of H3B cells to prevent de novo transcription from donor cell proviral DNA after co-cultivation did not affect the initiation of in vivo reverse transcription following cell-to-cell HIV infection. These results demonstrated that cells persistently infected with HIV contained significant fully cleaved cell-associated RT in a form that was active in vitro but not in vivo and that following cell-to-cell transmission of HIV infection to susceptible cells, de novo reverse transcription was initiated without detectable evidence of further synthesis or proteolytic processing of HIV RT. The nature of this initiation process requires further study.
Publisher: Microbiology Society
Date: 1993
DOI: 10.1099/0022-1317-74-1-33
Abstract: A cell clone persistently infected with human T cell-lymphotrophic virus type IIIB (H3B cells) contained mainly the multiply spliced (2 kb) and singly spliced (4.3 kb) species of human immunodeficiency virus (HIV) RNA. When H3B cells were co-cultured with susceptible HUT78 cells, cell fusion occurred within 4 h of cell mixing and was accompanied by a marked increase of the unspliced full-length (9.2 kb) HIV RNA. This first phase of viral RNA induction (4 to 12 h post-infection) was followed by a second phase of viral RNA synthesis from 24 h p.i. in which there were significant increases in all three species of HIV RNA. Reverse transcriptase (RT) inhibitors such as azidothymidine (AZT) at concentrations that abolished de novo HIV DNA synthesis, abolished the first phase but not the second phase of viral RNA synthesis in our model system. A comparable one-step cell-free virus infection showed a pattern of viral RNA synthesis similar to that of the cell-to-cell transmission of infection. However, viral RNA synthesis following cell-free virus infection was totally inhibited by RT inhibitors. The early phase (4 to 12 h) expression of 9.2 kb HIV RNA is likely to use newly synthesized HIV DNA as template during this phase, HIV RNA and DNA syntheses occur simultaneously, with each process being dependent on the other for maximal yield. During the later (24 to 48 h) phase, all three HIV RNA species may be transcribed at least in part from proviral DNA from the original donor cells. This later phase may provide one of the mechanisms for natural spread of virus to new cells and for enhanced viral gene expression in vivo, despite the presence of AZT.
Location: Australia
Location: Australia
No related grants have been discovered for Tuck-Weng Kok.