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PubMed1991

Characterization of an immunoreactive 93-kDa core protein of Borrelia burgdorferi with a human IgG monoclonal antibody.

Lyme borreliosis is an infectious disease caused by the tick-borne spirochete Borrelia burgdorferi, which carries the potential for chronic infection. Ag on the etiologic Borrelia are currently being defined structurally and their ability to elicit immune responses delineated. EBV can be used to immortalize human B. burgdorferi-specific B cells from infected donors and generate antibodies against antigenic epitopes encountered in natural infection. A human mAb secreting EBV-transformed B cell line, D7, has been developed that is specific for a 93-kDa B. burgdorferi protein and has been used to characterize this potentially important Ag. D7 produces an IgG3 antibody that detects the 93-kDa Ag as well as smaller fragments at 46 kDa and lower molecular mass. The antibody detects similar epitopes on all B. burgdorferi isolates tested and on a Borrelia hermsii protein with molecular mass greater than 100 kDa but binds poorly to Treponema species. In contrast, polyclonal sera from Lyme disease patients show little binding to the homologous Ag in B. hermsii. Structurally, the 93-kDa protein is associated with the flagellum and may be firmly anchored in the protoplasmic cylinder. It is not solubilized by nonionic detergent treatment of the whole Borrelia. Antibodies against a comparable m.w. protein are present in sera from patients with both early and late infection. Thus, antibodies against this Ag are a sensitive and specific marker of Borrelia infection. This Ag is likely of structural importance and may represent a target of host defenses.

Antibodies, Bacterial↗
PubMed2026

CRISPR-Cas-based diagnostics for point-of-care detection of sexually transmitted infections: a laboratory development and evaluation study.

BACKGROUND: Timely, point-of-care diagnosis of sexually transmitted infections (STIs) is crucial for enabling prompt treatment and reducing transmission. We aimed to develop a portable, multiplexed, CRISPR-based assay panel for the detection of Neisseria gonorrhoeae (including the ciprofloxacin resistance marker gyrA S91F), Chlamydia trachomatis, Treponema pallidum, and herpes simplex virus (HSV). METHODS: In this laboratory development and evaluation study, we developed and optimised four multiplexed, CRISPR-based, diagnostic STI assays for point-of-care use. The complete assay panel comprised a CRISPR TP-HSV (cTP-HSV) panel for the detection of T pallidum and pan-HSV, with reflex testing to distinguish HSV-1 from HSV-2, and a CRISPR NG-CT (cNG-CT) panel for the detection of N gonorrhoeae and C trachomatis, with reflex testing to detect N gonorrhoeae using two additional genome regions and to identify the gyrA S91F mutation. Each pathogen was targeted at two independent genomic regions by isothermal amplification and CRISPR-Cas reaction using Cas12a and Cas13a, each with distinct fluorescent reporters. Analytical specificity and limits of detection (LODs) were determined, and a retrospective, masked concordance study was conducted on genomic DNA from 900 clinical samples (400 for cTP-HSV and reflex testing and 500 for cNG-CT and reflex testing), using quantitative PCR as the reference standard. The diagnostic accuracy of the test was assessed by analysis of receiver operating characteristic curves. FINDINGS: The overall sensitivity of the TP-HSV CRISPR assay was 82·5% (95% CI 74·0-88·7) for T pallidum and 94·4% (90·2-97·0) for pan-HSV; LODs were 6·2 copies per μL for T pallidum and 7·8 copies per μL for HSV. Reflex testing gave sensitivities of 97·0% (91·1-99·3) for HSV-1 and 96·0% (89·7-98·7) for HSV-2. The NG-CT CRISPR assay had an overall sensitivity of 80·0% (74·0-84·9) for N gonorrhoeae and 73·0% (65·5-79·3) for C trachomatis, with a LOD of 3·9 copies per μL for both pathogens. Reflex testing for the detection of the gyrA S91F mutation in N gonorrhoeae showed an overall sensitivity of 63·1% (55·1-70·4); however, this was dependent on sample type, with a sensitivity of 85·7% (46·7-99·5) in genital samples and 61·2% (52·8-68·9) in extragenital samples. For all pathogens, assay sensitivity was positively correlated with pathogen load. Area under the curve (AUC) values were 0·90 for T pallidum and 0·99 for pan-HSV in the TP-HSV assay, with values of 0·99 for HSV-1 and 0·97 for HSV-2 obtained in the reflex HSV-1-HSV-2 assay. For the cNG-CT assay, AUC values were 0·90 for N gonorrhoeae and 0·85 for C trachomatis, with a value of 0·72 obtained for gyrA S91F in the reflex cNG-gyrA assay. INTERPRETATION: Our multiplexed, CRISPR-based, point-of-care platform achieved performance consistent with WHO target product profiles for N gonorrhoeae and T pallidum. Proof-of-concept detection of the gyrA S91F resistance marker highlights its potential for resistance-guided therapy. Although optimisation is required before large-scale deployment, this suite offers a promising approach for rapid, decentralised, and resistance-informed STI diagnosis, particularly in resource-limited settings. FUNDING: Victorian Government Department of Health, Australian Government Department of Health, Disability and Ageing and Aged Care, and Australian Research Council.

Humans↗
PubMed1999

A randomized, double-blind, placebo-controlled trial of single-dose ciprofloxacin versus erythromycin for the treatment of chancroid in Nairobi, Kenya.

A randomized, double-blind, placebo-controlled clinical trial was conducted in Nairobi, Kenya, to compare single-dose ciprofloxacin with a 7-day course of erythromycin for the treatment of chancroid. In all, 208 men and 37 women presenting with genital ulcers clinically compatible with chancroid were enrolled. Ulcer etiology was determined using culture techniques for chancroid, serology for syphilis, and a multiplex polymerase chain reaction for chancroid, syphilis, and herpes simplex virus (HSV). Ulcer etiology was 31% unmixed chancroid, 23% unmixed syphilis, 16% unmixed HSV, 15% mixed etiology, and 15% unknown. For 111 participants with chancroid, cure rates were 92% with ciprofloxacin and 91% with erythromycin. For all study participants, the treatment failure rate was 15%, mostly related to ulcer etiologies of HSV infection or syphilis, and treatment failure was 3 times more frequent in human immunodeficiency virus-infected subjects than in others, mostly owing to HSV infection. Ciprofloxacin is an effective single-dose treatment for chancroid, but current recommendations for empiric therapy of genital ulcers may result in high treatment failure due to HSV infection.

Adolescent↗
PubMed1992

Polymerase chain reaction for detection of Leptospira spp. in clinical samples.

A sensitive assay for Leptospira spp., the causative agent of leptospirosis, was developed on the basis of the polymerase chain reaction (PCR). A 331-bp sequence from the Leptospira interrogans serovar canicola rrs (16S) gene was amplified, and the PCR products were analyzed by DNA-DNA hybridization by using a 289-bp fragment internal to the amplified DNA. Specific PCR products also were obtained with DNA from the closely related nonpathogenic Leptospira biflexa but not with DNA from other spirochetes, such as Borrelia burgdorferi, Borrelia hermsii, Treponema denticola, Treponema pallidum, Spirochaeta aurantia, or more distant organisms such as Escherichia coli, Staphylococcus aureus, Mycobacterium tuberculosis, and Proteus mirabilis. The assay was able to detect as few as 10 bacteria. Leptospira DNA was detected in urine from experimentally infected mice. In addition, the test was found to be suitable for diagnosing leptospirosis in humans. Cerebrospinal fluid and urine from patients with leptospirosis were positive, whereas samples from control uninfected patients were negative.

Adolescent↗
PubMed2004

Alternative cerebrospinal fluid tests to diagnose neurosyphilis in HIV-infected individuals.

OBJECTIVE: To identify alternatives to the CSF-Venereal Disease Research Laboratory (VDRL) test for the diagnosis of neurosyphilis in HIV-infected individuals. METHODS: CSF fluorescent treponemal antibody (FTA) reactivity and % CSF lymphocytes that were B cells in fresh and frozen samples were determined for 47 HIV-infected cases with syphilis and 26 HIV-infected controls. As for serum, CSF fluorescent treponemal antibody reactivity > or =2+ was considered positive. Based on the results in controls and cases with normal CSF measures, cut-offs for elevated CSF B cells were proposed to be > or =9% in fresh and > or =20% in frozen samples. Neurosyphilis was defined as a reactive CSF-VDRL. RESULTS: CSF-FTA-ABS (absorbed) and CSF-FTA (unabsorbed and undiluted) were 100% sensitive for the diagnosis of neurosyphilis. Elevated % CSF B cells in fresh and cryopreserved samples was specific (100%) but not sensitive (40 and 43%) in post hoc analyses. The results of CSF-FTA and assessment of % CSF B cells together allowed 16% of cases with pleocytosis but nonreactive CSF-VDRL to be diagnosed with neurosyphilis and 26% to be diagnosed as not having neurosyphilis. CONCLUSION: When the CSF-VDRL is nonreactive, CSF-FTA and % CSF B cells may help exclude or establish the diagnosis of neurosyphilis.

Absorption↗
PubMed2002

Validation of syndromic algorithm for the management of genital ulcer diseases in China.

OBJECTIVES: 1. To determine the aetiologies of genital ulcers in China. 2. To evaluate a modified WHO syndromic management algorithm for genital ulcer disease (GUD). METHODS: Patients with genital ulcers were enrolled at their first visit to STD clinics in the cities of Shanghai and Chengdu. They were managed according to a modified WHO algorithm for GUD, in which no treatment was given for chancroid. A multiplex polymerase chain reaction was used to detect Treponema pallidum, Herpes simplex and Haemophilus ducreyi. Dark field examination and serology (rapid plasma reagin and Treponema pallidum particle agglutination assay (TPPA) were also used to diagnose syphilis. RESULTS: A total of 227 male and female patients were enrolled. Syphilis alone was diagnosed in 78 (35%), genital herpes alone in 43 (19%), and both infections were present in 28 (12%). No diagnosis was made in 76 (34%). No case of chancroid was identified. The sensitivity of the algorithm for syphilis and herpes was 88.7% and 69.0% respectively, the specificity 95.0% and 50.0%. 12/106 cases of syphilis were incorrectly classified as herpes (11%), and did not receive treatment. More than 97% of patients followed up responded clinically to treatment. CONCLUSION: Further validation and revision of the WHO flowchart for GUD are needed.

Adolescent↗
PubMed1985

[Pathogenesis, clinical aspects and diagnosis of the most important enteric bacterial diseases in animals].

There are adhesive and invasive species among enteropathogenic bacteria for animals. Most frequently isolated adhesive bacteria are enterotoxigenic E. coli (ETEC). Salmonella, T. hyodysenteriae, M. paratuberculosis, Y. enterocolitica, Cl. perfringens and C. jejuni bacteria represent the most important invasive germs. The main clinical finding connected with bacterial enteritis is diarrhoea resp. dysentery. Evaluation of feces consistency, time of appearance and accompanying symptoms (e.g. fever, colic) allow a clinical suspective diagnosis, however a definitive diagnosis must be based on the demonstration of the causative agent. In addition to known cultural, microscopic, biochemical and serological diagnostic methods, the demonstration of virulence factors (enterotoxin formation, adhesive pili) plays an important role in diagnosis of enteropathogenic bacteria.

Animals↗
PubMed1985

Monoclonal antibodies for the diagnosis of sexually transmitted diseases.

Monoclonal antibodies are already being used for the diagnosis of human sexually transmitted diseases. These antibodies can be used to detect a wide range of microorganisms, including bacteria, parasites, and viruses. For both culture and direct tests, monoclonal antibodies showed patterns of specificity and reproducibility that exceeded those available with conventionally prepared antisera. The direct tests for these organisms required less than an hour to perform, representing a major advancement in a diagnosis that previously required 2 to 6 days of culture followed by confirmatory testing. Furthermore, rapid differential diagnosis of infection will now be possible. Because some sexually transmitted diseases may be transmitted simultaneously and share similar clinical manifestations (that is, gonorrhea and chlamydia in cervicitis or urethritis, syphilis or herpes in genital ulcers), it will be possible to differentiate a single from a multiple infection by simultaneous testing of direct samples with the appropriate monoclonal antibody reagents.

Antibodies, Bacterial↗
PubMed1983

Pathophysiologic features of swine dysentery: cyclic nucleotide-independent production of diarrhea.

Net electrolyte and water transport and unidirectional Na+ fluxes were examined in ligated colonic loops of clinically normal pigs and in pigs with swine dysentery (etiologic agent Treponema hyodysenteriae) in the presence or absence of theophylline. In normal pigs, theophylline abolished net Na+ absorption via a reduction in the lumen-to-blood flux, decreased Cl- absorption, and increased HCO3- accumulation in the lumen. In infected pigs, all net ion transport was abolished, with the addition of theophylline producing little effect. The absence of net Na+ absorption in infected pigs was also the result of a decreased lumen-to-blood flux. Seemingly, colonic malabsorption may be the primary transport alteration in swine dysentery. Concentrations of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) were measured in samples of colonic mucosa from normal and infected pigs after in vitro exposure to a Ringer's solution containing 0 or 20 mM theophylline. Basal values of cAMP or cGMP did not increase in infected colonic mucosa. There was a diminished capacity of the infected mucosa to respond to theophylline. Alterations in ion transport in conjunction with measurements of cAMP and cGMP indicated that the pathogenic mechanism(s) in swine dysentery were not similar to those of Salmonella, Shigella, Vibrio cholerae, or Escherichia coli diarrhea.

Animals↗
PubMed1997

Studies on the lysozyme independence of immune immobilisation of Treponema pallidum and the frequency of lysozyme autoantibodies in syphilitic sera.

The role of lysozyme in the immune immobilisation of Treponema pallidum is not yet fully understood. The T. pallidum immobilisation assay was used to demonstrate that the immobilisation and lysis of T. pallidum in vitro by antibodies (serum, IgG fraction or IgM fraction) and complement proceed in a lysozyme-independent mode. In the presence of lysozyme the rate of immobilisation increased. In contrast with its effect on Escherichia coli, the effect of lysozyme on T. pallidum was governed exclusively by its enzymic activity rather than by the cationic protein nature of the molecule. Lysozyme, released from stimulated phagocytes, induced formation of lysozyme antibodies in 59.6% of syphilis patients as determined by lysozyme antibody ELISA. The highest frequency was found in patients with untreated secondary syphilis, whereas untreated primary syphilis was only rarely accompanied by the presence of lysozyme antibodies. Cross-reactivities between lysozyme and treponemal antigens were excluded by immunoblotting. The autoantibodies did not influence the lysozyme activity. It was concluded that the formation of lysozyme antibodies is only an epiphenomenon in the host defence against treponemal infection.

Animals↗
PubMed1994

Detection of Treponema pallidum by polymerase chain reaction in the cerebrospinal fluid of syphilis patients.

To find out if polymerase chain reaction could be used as a diagnostic tool for detecting neurosyphilis, we have applied the PCR for the detection of Treponema pallidum DNA in the cerebrospinal fluid (CSF) of syphilis patients. The results of the PCR of the CSF in 26 patients with at various stages of illness were compared with the results of other conventional tests used in the WHO criteria. T. pallidum was detected in the CSF of patients at all stages of syphilis, which indicates that they invade the central nervous system from the early stages of infection. However, the presence of T. pallidum in the CSF was not correlated with the results of other tests used in the WHO criteria, and its significance in the diagnosis of neurosyphilis should further be evaluated.

Cerebrospinal Fluid↗
PubMed2003

Common transfusion-transmitted infectious agents among thalassaemic children in Bangladesh.

Transfusion-dependent children are more prone to acquiring various transfusion-transmitted infections (TTIs), such as hepatitis B (HBV), hepatitis C (HCV), HIV, and others. Since the magnitude of these infections among thalassaemic children in Bangladesh is not well-known, this study was conducted to assess the prevalence of TTIs among them (who received more than three blood transfusions) compared to their age- and sex-matched controls (non-thalassaemics and those who had never had a transfusion). Seromarkers for HBV, HCV, HDV, Treponema pallidum, and HIV were tested, and the results were analyzed using SPSS/Windows 10.5. Of 259 children studied, 152 (58.69%) were thalassaemic (mean age 6.8 +/- 3.6 years), and 107 were controls (mean age 6.7 +/- 3.53 years). The HBV and HCV-markers were found significantly more often among multi-transfused thalassaemic children than among the controls in terms of HBsAg (13.8% vs 6.5%, p < 0.04), anti-HBc total (39.5% vs 9.4%, p < 0.0001), and anti-HCV (12.5% vs 0.9%, p < 0.0001). HBeAg did not differ (p = 0.82) between the thalassaemics (9.52%) and the controls (14.28%), whereas anti-HBe differed (0% vs 57.14%, p < 0.003). Neither the thalassaemics nor the controls were positive for HDV, HIV, or T. pallidum. Since more thalassaemic children acquired hepatitis B and C infections through multiple blood transfusions, it is recommended that the safe blood-transfusion programme be strengthened and mass vaccination against HBV (even who suffer from HCV) in Bangladesh be undertaken.

Adolescent↗
PubMed1987

Identification of cells producing anti-treponemal lymphotoxin (ATL).

Identification of cells producing ATL was carried out on suspensions of cells enriched with lymphocytes T, B and macrophages. The cells were isolated from the spleen of syphilitic rabbits and from their testes when the rabbits were infected intratesticularly. The isolation of lymphocytes and macrophages was performed on different days after infection. It has been found that all examined cells are able to produce ATL but most active are T lymphocytes. These cells isolated from the testes were able to produce ATL already 6 days after infection, whereas lymphocytes B and macrophages acquired this capability not before day 10-13. The ability of cells to produce ATL was followed by marked reduction of treponemes from testes. The isolated treponemes were also immobilized and destroyed. Dependence between capability of cells to produce ATL and disappearance of treponemes from infected organ indicated that ATL kills treponemes also in vivo.

Animals↗
PubMed2001

[Notification according to the law for protection against infections].

Since January 1st 2001, the Law for Protection Against Infections decrees the duty of notification along two tracks, on the one hand by notification of infectious diseases and on the other hand by reporting the pathogens. However, the names of pathogens change due to scientific progress. Furthermore, these names include in some cases not only harmful agents of infectious diseases but also inoffensive organisms, i. e. Giardia lamblia. There are also problems in the notification of other pathogens such as Brucella sp., Campylobacter sp., Chlamydia psittaci,Clostridium botulinum, Cryptosporidium parvum, Leptospira interrogans, Listeria monocytogenes, Mycobacterium tuberculosis and similar agents, Norwalk-like viruses, Salmonella Paratyphi, Treponema pallidum, Trichinella spiralis and pathogenic serovars of Yersinia enterocolitica. Finally, the abandonment of notification for glanders, lymphogranuloma venereum, ulcus molle and variola appears to be risky, the reasons being partly unconvincing and contradictory.

Communicable Disease Control↗
PubMed2001

Clinical periodontal findings and microflora profiles in children with chronic neutropenia under supervised oral hygiene.

BACKGROUND: This is the first known case report that used a polymerase chain reaction (PCR)-based method to help identify the oral microflora in patients with chronic neutropenia. In this study, we report clinical periodontal findings and microflora profiles of 2 children, 1 with severe congenital neutropenia (SCN, Kostmann type) and 1 with cyclic neutropenia (CN). METHODS: The SCN patient had severe gingivitis, whereas the patient with CN had mild gingivitis in the gingival margins. Monthly oral cleaning instruction and review were performed without subsequent periodontal therapy. Oral hygiene conditions remained satisfactory and visible plaque was scarce, despite the persistence of mild gingivitis. Under supervised oral hygiene, we examined the presence of periodontal pathogens from patient plaque samples. RESULTS: By a PCR-based method, Prevotella nigrescens, Bacteroides forsythus, Campylobacter rectus, and Capnocytophaga gingivalis were detected in the SCN patient and P. intermedia, C. rectus, C. gingivalis, and C. sputigena in the CN patient, suggesting the existence of periodontal pathogens. Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Treponema denticola, and C. ochracea were not found in either patient. CONCLUSIONS: Use of 1% povidone iodine solution and local antibiotic application under supervised oral hygiene were helpful to improve gingival conditions in patients with chronic neutropenia.

Aggregatibacter actinomycetemcomitans↗
PubMed1976

Growth and subculture of pathogenic T. pallidum (Nichols strain) in BHK-21 cultured tissue cells.

The growth and nine subcultivations of the experimental Nichols strain of pathogenic Treponema pallidum were successfully accomplished in cultured baby hamster kidney tissue cells (BHK-21) using serum-free media. The number (motile and non-motile cells occurring extracellulary) of cell generations generally increased with each subcultivation, the largest increases (greater than 3-0) occurring in subcultures 4, 5, 7, 8, and 9. Also, the number of motile cells decreased with each subcultivation. Virulent organisms were demonstrated in subcultures 1, 4, 5, 7, and 8, and their numbers estimated with the use of standards established by the animal inoculation titration of counted numbers of organisms freshly harvested from experimental infections; the estimated number of virulent organisms was higher than or equal to the counted cultivated treponemes injected, which may be attributable to uncounted virulent spirochaetes occurring intracellularly.

Animals↗