scholarly journals Sensitive and Specific Enzyme-Linked Immunosorbent Assay for Detecting Serum Antibodies against Mycobacterium avium subsp. paratuberculosis in Fallow Deer

2014 ◽  
Vol 21 (8) ◽  
pp. 1077-1085 ◽  
Author(s):  
José M. Prieto ◽  
Ana Balseiro ◽  
Rosa Casais ◽  
Naiara Abendaño ◽  
Liam E. Fitzgerald ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Mycobacterium Avium ◽  
Cost Effective ◽  
Fallow Deer ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Igg Antibody ◽  
Tissue Samples ◽  
Content Type ◽  
Mycobacterium Avium Subsp Paratuberculosis

ABSTRACTThe enzyme-linked immunosorbent assay (ELISA) is the diagnostic test most commonly used in efforts to control paratuberculosis in domestic ruminants. However, commercial ELISAs have not been validated for detecting antibodies againstMycobacterium aviumsubsp.paratuberculosisin wild animals. In this study, we compared the sensitivities and specificities of five ELISAs using individual serum samples collected from 41 fallow deer with or without histopathological lesions consistent with paratuberculosis. Two target antigenic preparations were selected, an ethanol-treated protoplasmic preparation obtained from a fallow deerM. aviumsubsp.paratuberculosisisolate (ELISAs A and B) and a paratuberculosis protoplasmic antigen (PPA3) (ELISAs C and D). Fallow deer antibodies bound to the immobilized antigens were detected by using a horseradish peroxidase (HRP)-conjugated anti-fallow deer IgG antibody (ELISAs A and C) or HRP-conjugated protein G (ELISAs B and D). A commercially available assay, ELISA-E, which was designed to detectM. aviumsubsp.paratuberculosisantibodies in cattle, sheep, and goats, was also tested. Although ELISAs A, C, and E had the same sensitivity (72%), ELISAs A and C were more specific (100%) for detecting fallow deer with lesions consistent with paratuberculosis at necropsy than was the ELISA-E (87.5%). In addition, the ELISA-A was particularly sensitive for detecting fallow deer in the latent stages of infection (62.5%). The antibody responses detected with the ELISA-A correlated with both the severity of enteric lesions and the presence of acid-fast bacteria in gut tissue samples. In summary, our study shows that the ELISA-A can be a cost-effective diagnostic tool for preventing the spread of paratuberculosis among fallow deer populations.

scholarly journals Diagnosis of Bovine Paratuberculosis by a Novel Enzyme-Linked Immunosorbent Assay Based on Early Secreted Antigens of Mycobacterium avium subsp. paratuberculosis

2008 ◽  
Vol 15 (8) ◽  
pp. 1277-1281 ◽  
Author(s):  
Sung Jae Shin ◽  
Donghee Cho ◽  
Michael T. Collins
Keyword(s):  
Immunosorbent Assay ◽  
Mycobacterium Avium ◽  
Area Under The Curve ◽  
Cost Effective ◽  
Enzyme Linked Immunosorbent Assay ◽  
Protein Antigens ◽  
Characteristic Analysis ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
Cattle Herds ◽  
Low Passage

ABSTRACT We previously reported that protein antigens of serodiagnostic potential were more abundant in culture filtrates than cellular extracts from liquid cultures of Mycobacterium avium subsp. paratuberculosis (D. Cho and M. T. Collins, Clin. Vaccine Immunol. 13:1155-1161, 2006). Based on this observation, a novel enzyme-linked immunosorbent assay (ELISA) using antigens secreted by young (early- to mid-log-phase) cultures of M. avium subsp. paratuberculosis JTC303 (a low-passage isolate originating from the ileum of a Holstein bull) in mycobactin-supplemented Watson-Reid medium (pH 6.0) was developed and evaluated using a previously described panel of bovine sera (M. T. Collins et al., Clin. Diagn. Lab. Immunol. 12:685-692, 2005) that included 444 paratuberculosis cases and 412 controls. The new assay, called JTC-ELISA, had a significantly higher diagnostic sensitivity and an equivalent specificity compared to those of five commercial paratuberculosis ELISA kits. By receiver-operating characteristic analysis, the JTC-ELISA had the highest area under the curve of the six assays evaluated. The JTC-ELISA was particularly sensitive at detecting low-level fecal shedders of Mavium subsp. paratuberculosis (40%; the sensitivity of the commercial kits was 20%). The JTC-ELISA works effectively on both serum and milk samples for the detection of cattle with subclinical M. avium subsp. paratuberculosis infections, providing a cost-effective diagnostic tool to support paratuberculosis control programs in cattle herds.

scholarly journals Relationship between Presence of Cows with Milk Positive for Mycobacterium avium subsp. paratuberculosis-Specific Antibody by Enzyme-Linked Immunosorbent Assay and Viable M. avium subsp. paratuberculosis in Dust in Cattle Barns

2013 ◽  
Vol 79 (18) ◽  
pp. 5458-5464 ◽  
Author(s):  
Susanne W. F. Eisenberg ◽  
Ruj Chuchaisangrat ◽  
Mirjam Nielen ◽  
Ad P. Koets
Keyword(s):  
Dairy Cattle ◽  
Immunosorbent Assay ◽  
Mycobacterium Avium ◽  
Specific Antibody ◽  
Enzyme Linked Immunosorbent Assay ◽  
Young Stock ◽  
Content Type ◽  
Lactating Cows ◽  
Settled Dust ◽  
Mycobacterium Avium Subsp Paratuberculosis

ABSTRACTParatuberculosis, or Johne's disease, in cattle is caused byMycobacterium aviumsubsp.paratuberculosis, which has recently been suspected to be transmitted through dust. This longitudinal study on eight commercialM. aviumsubsp.paratuberculosis-positive dairy farms studied the relationship between the number of cows withM. aviumsubsp.paratuberculosisantibody-positive milk and the presence of viableM. aviumsubsp.paratuberculosisin settled-dust samples, including their temporal relationship. Milk and dust samples were collected in parallel monthly for 2 years.M. aviumsubsp.paratuberculosisantibodies in milk were measured by enzyme-linked immunosorbent assay (ELISA) and used as a proxy forM. aviumsubsp.paratuberculosisshedding. Settled-dust samples were collected by using electrostatic dust collectors (EDCs) at six locations in housing for dairy cattle and young stock. The presence of viableM. aviumsubsp.paratuberculosiswas identified by liquid culture and PCR. The results showed a positive relationship (odds ratio [OR], 1.2) between the number of cows with ELISA-positive milk and the odds of having positive EDCs in the same airspace as the adult dairy cattle. Moreover, the total number of lactating cows also showed an OR slightly above 1. This relationship remained the same for settled-dust samples collected up to 2 months before or after the time of milk sampling. The results suggest that removal of adult cows with milk positive forM. aviumsubsp.paratuberculosis-specific antibody by ELISA might result in a decrease in the presence of viableM. aviumsubsp.paratuberculosisin dust and therefore in the environment. However, this decrease is likely delayed by several weeks at least. In addition, the data support the notion thatM. aviumsubsp.paratuberculosisexposure of young stock is reduced by separate housing.

scholarly journals Flow Cytometric Detection of Mycobacterium avium subsp. paratuberculosis-Specific Antibodies in Experimentally Infected and Naturally Exposed Calves

2013 ◽  
Vol 20 (9) ◽  
pp. 1457-1465 ◽  
Author(s):  
S. Schillinger ◽  
P. S. Bridger ◽  
H. Bulun ◽  
M. Fischer ◽  
Ö. Akineden ◽  
...  
Keyword(s):  
Mycobacterium Avium ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Adult Cattle ◽  
Content Type ◽  
Specific Antibodies ◽  
Bacterial Surface ◽  
Flow Cytometric ◽  
Mycobacterium Phlei ◽  
Mycobacterium Avium Subsp Paratuberculosis

ABSTRACTA desirable test to diagnose infections withMycobacterium aviumsubsp.paratuberculosisfacilitates identification of infected cattle prior to the state ofM. aviumsubsp.paratuberculosisshedding. This study aimed at adjusting a flow cytometry (FC)-based assay, using intactM. aviumsubsp.paratuberculosisbacteria as the antigen, for diagnosis ofM. aviumsubsp.paratuberculosisinfections in calves. Serum samples were collected from experimentally infected (n= 12) and naturally exposed (n= 32) calves. Samples from five calves from positive dams were analyzed to determine the dynamics of maternal antibodies. Samples from adult cattle with defined infection status served as the standard (18M. aviumsubsp.paratuberculosisshedders, 22M. aviumsubsp.paratuberculosisfree). After preadsorption withMycobacterium phlei, sera were incubated withM. aviumsubsp.paratuberculosisandM. aviumsubsp.aviumbacterial suspensions, respectively, followed by the separate detection of bovine IgG, IgG1, IgG2, and IgM attached to the bacterial surface.M. aviumsubsp.paratuberculosis-specific sample/positive (S/P) ratios were compared to enzyme-linked immunosorbent assay (ELISA) S/P ratios. In adult cattle, the FC assay for IgG1 had a sensitivity of 78% at a specificity of 100%. Maternally acquired antibodies could be detected in calves up to 121 days of life. While all but two sera taken at day 100 ± 10 postnatum from naturally exposed calves tested negative, elevated S/P ratios (IgG and IgG1) became detectable from 44 and 46 weeks postinoculation onwards in two calves infected experimentally. Even with the optimized FC assay,M. aviumsubsp.paratuberculosis-specific antibodies can only occasionally be detected in infected calves less than 12 months of age. The failure to detect such antibodies apparently reflects the distinct immunobiology ofM. aviumsubsp.paratuberculosisinfections rather than methodological constraints.

scholarly journals Seroprevalence of Immunoglobulin G Antibodies Against Mycobacterium avium subsp. paratuberculosis in Dogs Bred in Japan

2020 ◽  
Vol 7 (3) ◽  
pp. 93
Author(s):  
Takashi Kuribayashi ◽  
Davide Cossu ◽  
Eiichi Momotani
Keyword(s):  
Mycobacterium Avium ◽  
Immunoglobulin G ◽  
Mycobacterium Avium Subsp ◽  
Clinical Samples ◽  
Enzyme Linked Immunosorbent Assay ◽  
Igg Antibodies ◽  
Igg Antibody ◽  
The Difference ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
Veterinary Hospital

In this study, the seroprevalence of immunoglobulin G (IgG) antibodies against Mycobacterium avium subsp. paratuberculosis (MAP) in dogs bred in Japan was evaluated. Ninety-two non-clinical samples were obtained from three institutes and fifty-seven clinical samples were obtained from a veterinary hospital in Japan. Serum titers of total IgG, IgG1 and IgG2 isotype antibodies against MAP were measured using an indirect enzyme-linked immunosorbent assay (ELISA). The IgG antibodies against MAP in non-clinical serum obtained from three institutes was observed to be 2.4%, 20% and 9.0%. Similarly, the IgG1 antibodies titers against MAP were observed to be 7%, 20% and 0%. Lastly, the IgG2 antibodies against MAP were observed to be 7%, 20% and 4.4%. No significance differences in these titers were observed among the three institutes. The IgG, IgG1 and IgG2 antibodies in serum obtained from a veterinary hospital were observed to be 55.3%, 42% and 42%, respectively. Significant differences were found between the non-clinical and clinical samples. The titers in the clinical samples showed a high degree of variance, whereas low variance was found in the non-clinical samples. The IgG antibody levels were thought to be induced following exposure to MAP-contaminated feed. The difference in titers between the clinical and non-clinical samples is likely to be related to the amount of MAP antigen contamination in dog foods.

scholarly journals Development of a Novel Cocktail Enzyme-Linked Immunosorbent Assay and a Field-Applicable Lateral-Flow Rapid Test for Diagnosis of Contagious Bovine Pleuropneumonia

2016 ◽  
Vol 54 (6) ◽  
pp. 1557-1565 ◽  
Author(s):  
Martin Heller ◽  
Nimmo Gicheru ◽  
Georgina Tjipura-Zaire ◽  
Cecilia Muriuki ◽  
Mingyan Yu ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Rapid Test ◽  
Lateral Flow ◽  
Sub Saharan Africa ◽  
Enzyme Linked Immunosorbent Assay ◽  
Contagious Bovine Pleuropneumonia ◽  
Serum Samples ◽  
Content Type ◽  
Mycoplasma Mycoides ◽  
Sub Saharan

Contagious bovine pleuropneumonia (CBPP) is a severe respiratory disease that is widespread in sub-Saharan Africa. It is caused byMycoplasma mycoidessubsp.mycoides, a bacterium belonging to theMycoplasma mycoidescluster. In the absence of an efficient CBPP vaccine, improved and easy-to-use diagnostic assays for recurrent testing combined with isolation and treatment of positive animals represent an option for CBPP control in Africa. Here we describe the comprehensive screening of 17 immunogenicMycoplasma mycoidessubsp.mycoidesproteins using well-characterized bovine sera for the development of a novel cocktail enzyme-linked immunosorbent assay (ELISA) for laboratory use. Two recombinantMycoplasmaimmunogens, MSC_0136 and MSC_0636, were used to set up a standardized cocktail ELISA protocol. According to the results from more than 100 serum samples tested, the sensitivity and specificity of the novel cocktail ELISA were 85.6% and 96.4%, respectively, with an overall diagnostic accuracy comparable to that of the Office International des Epizooties (OIE)-prescribed serological assays. In addition, we provide a proof of principle for a field-applicable, easy-to-use commercially produced prototype lateral-flow test for rapid (<30-min) diagnosis of CBPP.

scholarly journals Detection of Francisella tularensis-Specific Antibodies in Patients with Tularemia by a Novel Competitive Enzyme-Linked Immunosorbent Assay

2012 ◽  
Vol 20 (1) ◽  
pp. 9-16 ◽  
Author(s):  
Neekun Sharma ◽  
Akitoyo Hotta ◽  
Yoshie Yamamoto ◽  
Osamu Fujita ◽  
Akihiko Uda ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Francisella Tularensis ◽  
Immunosorbent Assay ◽  
Enzyme Linked Immunosorbent Assay ◽  
The Novel ◽  
Serum Samples ◽  
Content Type ◽  
Microagglutination Test ◽  
Highly Sensitive ◽  
High Degree

ABSTRACTA novel competitive enzyme-linked immunosorbent assay (cELISA) was developed and evaluated for detection of antibodies againstFrancisella tularensisin humans. The assay is based on the ability of serum antibodies to inhibit the binding of monoclonal antibodies (MAbs) directed againstF. tularensislipopolysaccharide antigens. The assay was evaluated using serum samples of tularemia patients, inactivatedF. tularensis-immunized rabbits, andF. tularensis-infected mice. Antibodies againstF. tularensiswere successfully detected in serum samples of tularemia patients as well as the immunized and infected animals. The cELISA method was compared to indirect ELISA (iELISA) and the commonly used microagglutination test (MA) using serum samples of 19 tularemia patients and 50 healthy individuals. The sensitivity and specificity of cELISA were 93.9 and 96.1%, respectively, in comparison to the iELISA. MA was less sensitive than cELISA with a sensitivity and specificity of only 81.8 and 98.0%, respectively. A high degree of correlation (R2= 0.8226) was observed between cELISA and iELISA results. The novel cELISA developed in this study appears to be highly sensitive and specific for serodiagnosis of human tularemia. The potential of the MAb-based cELISA to be used in both human and animal samples emphasizes its usefulness for serological survey of tularemia among multiple animal species.

scholarly journals Peptide-Based OspC Enzyme-Linked Immunosorbent Assay for Serodiagnosis of Lyme Borreliosis

1998 ◽  
Vol 36 (12) ◽  
pp. 3474-3479 ◽  
Author(s):  
Marianne J. Mathiesen ◽  
Michael Christiansen ◽  
Klaus Hansen ◽  
Arne Holm ◽  
Eva Åsbrink ◽  
...  
Keyword(s):  
Lyme Borreliosis ◽  
Immunosorbent Assay ◽  
Erythema Migrans ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Amino Acid Residues ◽  
Serum Samples ◽  
Igg Antibody ◽  
Acrodermatitis Chronica Atrophicans ◽  
Increased Sensitivity

Sera from 210 patients with Lyme borreliosis (LB) were studied by an enzyme-linked immunosorbent assay (ELISA) based on a synthetic peptide (pepC10) comprising the C-terminal 10-amino-acid residues of OspC of Borrelia burgdorferi. We found that 36.3 and 45.0% of the serum samples from patients with erythema migrans (EM) and neuroborreliosis (NB), respectively, displayed immunoglobulin M (IgM) anti-pepC10 reactivities, while these samples rarely (≤8%) displayed IgG antibody reactivities. Sera from patients with acrodermatitis chronica atrophicans did not contain anti-pepC10 antibodies. The diagnostic performance of this newly developed peptide ELISA was compared with those of an ELISA based on the full-length recombinant OspC protein (rOspC) and a commercially available ELISA based on theB. burgdorferi flagellum (Fla). The sensitivity of the IgM pepC10 ELISA was slightly lower (P < 0.04) than that of the rOspC ELISA for EM patients (36.3 versus 43.8%), while there was no difference for NB patients (45.0 versus 48.0%). However, the optical density values obtained by the pepC10 ELISA were generally higher than those obtained by the rOspC ELISA, leading to a significantly better quantitative discrimination between seropositive patients with NB and controls (P < 0.008). The specificity of the pepC10 ELISA was similar to those of the rOspC ELISA and the Fla ELISA for relevant controls including patients with syphilis and mononucleosis. Although the overall diagnostic sensitivity of the Fla ELISA was superior, 8.8 and 12.0% of the EM and NB patients, respectively, were antibody positive only by the pepC10 ELISA. Thus, use of a diagnostic test for LB based on the detection of IgM antibodies to pepC10 and Fla has increased sensitivity for the diagnosis of early LB.

scholarly journals Uptake and Persistence of Mycobacterium avium subsp. paratuberculosis in Human Monocytes

2012 ◽  
Vol 80 (11) ◽  
pp. 3768-3775 ◽  
Author(s):  
Dayle A. Keown ◽  
David A. Collings ◽  
Jacqueline I. Keenan
Keyword(s):  
Mycobacterium Avium ◽  
Human Cells ◽  
Monocytic Cell ◽  
Monocytic Cell Line ◽  
Tissue Samples ◽  
Content Type ◽  
Human Monocytic Cell Line ◽  
Human Monocytic Cell ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
Lysosomal Protein

ABSTRACTMycobacterium aviumsubsp.paratuberculosisis a bacterium sometimes found in human blood and tissue samples that may have a role in the etiology of Crohn's disease in humans. To date, however, there have been few studies examining the interactions of these bacteria with human cells. Using the THP-1 human monocytic cell line, this study shows that the uptake and trafficking ofM. aviumsubsp.paratuberculosisin human cells are cholesterol dependent and that these bacteria localize to cholesterol-rich compartments that are slow to acidify.M. aviumsubsp.paratuberculosisbacteria containing phagosomes stain for the late endosomal marker Rab7, but recruitment of the Rab7-interacting lysosomal protein that regulates the fusion of bacterium-containing phagosomes with lysosomal compartments and facilitates subsequent bacterial clearance is significantly reduced. Disruption of phagosome acidification via this mechanism may contribute toM. aviumsubsp.paratuberculosispersistence in human cells, but there was no evidence that internalizedM. aviumsubsp.paratuberculosisalso affects the survival of bacteria taken up during a secondary phagocytic event.

scholarly journals Development of an Enzyme-Linked Immunosorbent Assay for Serodiagnosis of Ringworm Infection in Cattle

2013 ◽  
Vol 20 (8) ◽  
pp. 1150-1154 ◽  
Author(s):  
Elena Tatiana Băguţ ◽  
Ludivine Cambier ◽  
Marie-Pierre Heinen ◽  
Vasile Cozma ◽  
Michel Monod ◽  
...  
Keyword(s):  
Positive Predictive Value ◽  
Negative Predictive Value ◽  
Immunosorbent Assay ◽  
Predictive Value ◽  
Skin Lesions ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Predictive Values ◽  
Content Type ◽  
Ringworm Infection

ABSTRACTThe aim of this study was to develop an in-house enzyme-linked immunosorbent assay (ELISA) for the serological diagnosis of ringworm infection in cattle. We used available recombinant forms ofTrichophyton rubrumdipeptidyl peptidase V (TruDppV) andT. rubrumleucin aminopeptidase 2 (TruLap2), which are 98% identical toTrichophyton verrucosumorthologues. Field serum samples from 135 cattle with ringworm infection, as confirmed by direct microscopy, fluorescence microscopy, and PCR, and from 55 cattle without any apparent skin lesions or history of ringworm infection that served as negative controls were used. Sensitivities, specificities, and positive and negative predictive values were determined to evaluate the diagnostic value of our ELISA. Overall, the ELISAs based on recombinant TruDppV and TruLap2 discriminated well between infected animals and healthy controls. Highly significant differences (P< 0.0001, Mann-Whitney U test) were noted between optical density values obtained when sera from infected versus control cattle were tested. The ELISA developed for the detection of specific antibodies against DppV gave 89.6% sensitivity, 92.7% specificity, a 96.8% positive predictive value, and a 78.4% negative predictive value. The recombinant TruLap2-based ELISA displayed 88.1% sensitivity, 90.9% specificity, a 95.9% positive predictive value, and a 75.7% negative predictive value. To the best of our knowledge, this is the first ELISA based on recombinant antigens for assessing immune responses to ringworm infection in cattle; it is particularly suitable for epidemiological studies and also for the evaluation of vaccines and/or vaccination procedures.

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