Recombinant 46-kilodalton surface antigen (P46) of Mycoplasma hyopneumoniae expressed in Escherichia coli can be used for early specific diagnosis of mycoplasmal pneumonia of swine by enzyme-linked immunosorbent assay

Futo, Satoshi; Seto, Yasuhiro; Okada, Mizue; Sato, Shizuo; Suzuki, Tomohiko; Kawai, Kazuhiro; Imada, Yumiko; Mori, Yoshio

doi:10.1128/jcm.33.3.680-683.1995

Cited by 26 publications

(6 citation statements)

References 19 publications

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“…Both fluorescent antibody and enzyme-linked immunosorbent assay (ELISA) are commonly used assays. 2,9,10,12,14,16,20 Diagnosis by serology relies upon the response of the animal to the organism and thus can take 3-8 weeks to develop. Fluorescent techniques for the detection of antigen are compromised by crossreactivity with Mycoplasma hyorhinis and the nonpathogen Mycoplasma flocculare.…”

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Use of a Mycoplasma Hyopneumoniae Nested Polymerase Chain Reaction Test to Determine the Optimal Sampling Sites in Swine

et al. 2002

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A number of polymerase chain reaction (PCR)-based diagnostic tests have been developed for Mycoplasma hyopneumoniae, including one from this research group. This report presents further development, optimization, and standardization of a nested PCR test. Detection sensitivity was 1 fg of M. hyopneumoniae chromosomal DNA (approximately 1 organism). This exceeded the sensitivity of or compared favorably with other published PCR tests. Polymerase chain reaction primers to porcine 2-microglobulin were included as internal controls for amplifiable chromosomal DNA from porcine samples. To standardize the test, a number of samples from experimentally infected pigs, including nasal, tonsil, tracheobronchial swabs, lung tissue, bronchial alveolar lavage (BAL) fluid, and tracheobronchial brush samples, were examined by PCR. Samples obtained from BAL fluid and tracheobronchial sites were most predictive of infection, whereas nasal swabs and lung tissue were not reliable indicators of experimentally induced infection. In conclusion, the nested PCR developed for this study was found to be a highly sensitive and specific diagnostic tool for M. hyopneumoniae, but the enhanced sensitivity may be unnecessary if the proper sites are sampled. Porcine enzootic pneumonia caused by Mycoplasma hyopneumoniae continues to produce significant economic losses to swine producers. Mycoplasma hyop-neumoniae has recently gained new importance as an integral component of the porcine respiratory disease complex (PRDC). 24 Recent studies indicate that the incidence and severity of enzootic pneumonia have increased with the emergence of porcine reproductive and respiratory syndrome virus and the reemergence of swine influenza virus. Bacterial pathogens such as Pasteurella multocida also contribute to respiratory disease outbreaks. 17 Because of economic consideration , there is increased interest in the swine industry to improve diagnostic methods to gain a better understanding of PRDC in general and to provide farm-specific information for implementing effective prevention or control programs (or both) for M. hyopneu-moniae. Reliable diagnostic tests for M. hyopneumoniae are essential for developing cost-effective prevention and control programs. Visual analysis of lungs at the time of slaughter is commonly used to monitor the disease status of a herd. But it is not predictive of lifetime pneumonia. 21 The gold standard for detection of M. hyopneumoniae has been culture of the organism. But culture methods are rarely used because of several reasons: 1) prolonged time before a definitive diagnosis is obtained, 2) overgrowth with normal flora bacteria and non-M. hyopneumoniae mycoplasmas, and 3) lack of experience and capabilities to perform culture diagnosis of swine tissue samples because of the fastidious nature of M. hyopneumoniae. 18 Detection of serum antibodies is the most common diagnostic tool used to monitor M. hyopneumoniae infection. Both fluorescent antibody and enzyme-linked immunosor-bent assay (ELISA) are commonly used assays. 2,9,...

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Use of a Mycoplasma Hyopneumoniae Nested Polymerase Chain Reaction Test to Determine the Optimal Sampling Sites in Swine

et al. 2002

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“…Accurate diagnosis of EP is essential to prevent infection from spreading. The most commonly used diagnostic methods are serological analyses such as direct or blocking enzymelinked immunosorbent assay (6,9,16,18,23) and direct detection of the organism in clinical samples by immunofluorescence (10). In recent years, molecular genetic techniques which have improved sensitivity and specificity have become available (3,16).…”

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Detection of Mycoplasma hyopneumoniae by Air Sampling with a Nested PCR Assay

Stärk

Nicolet

Frey

1998

Appl Environ Microbiol

115

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This article describes the first successful detection of airborneMycoplasma hyopneumoniae under experimental and field conditions with a new nested PCR assay. Air was sampled with polyethersulfone membranes (pore size, 0.2 μm) mounted in filter holders. Filters were processed by dissolution and direct extraction of DNA for PCR analysis. For the PCR, two nested pairs of oligonucleotide primers were designed by using an M. hyopneumoniae-specific DNA sequence of a repeated gene segment. A nested PCR assay was developed and used to analyze samples collected in eight pig houses where respiratory problems had been common. Air was also sampled from a mycoplasma-free herd. The nested PCR was highly specific and 104 times as sensitive as a one-step PCR. Under field conditions, the sampling system was able to detect airborne M. hyopneumoniae on 80% of farms where acute respiratory disease was present. No airborne M. hyopneumoniae was detected on infected farms without acute cases. The chance of successful detection was increased if air was sampled at several locations within a room and at a lower air humidity.

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“…MHP168 392 and MHP168 393 were downexpressed and MHP168 418 were upexpressed in the virulent strain 168. Most lipoproteins are surface exposed and important components of mycoplasma membranes, and some in M. hyopneumoniae have been identified to play important roles in pathogenesis [43][44][45][46][47][48][49]. Mhp378, a homolog of MHP168 392 from M. hyopneumoniae 232, has been identified as a speciesspecific, highly immunogenic membrane-associated protein [50].…”

Section: Proteins Involved In Inositol Phosphate Metabolismmentioning

confidence: 99%

Quantitative Proteomic Analyses of a Pathogenic Strain and Its Highly Passaged Attenuated Strain ofMycoplasma hyopneumoniae

Fang

Liu

et al. 2019

BioMed Research International

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Mycoplasma hyopneumoniae is the causative agent of porcine enzootic pneumonia, a chronic respiratory disease in swine resulting in enormous economic losses. To identify the components that contribute to virulence and unveil those biological processes potentially related to attenuation, we used isobaric tags for relative and absolute quantification technology (iTRAQ) to compare the protein profiles of the virulent M. hyopneumoniae strain 168 and its attenuated highly passaged strain 168L. We identified 489 proteins in total, 70 of which showing significant differences in level of expression between the two strains. Remarkably, proteins participating in inositol phosphate metabolism were significantly downregulated in the virulent strain, while some proteins involved in nucleoside metabolism were upregulated. We also mined a series of novel promising virulence-associated factors in our study compared with those in previous reports, such as some moonlighting adhesins, transporters, lipoate-protein ligase, and ribonuclease and several hypothetical proteins with conserved functional domains, deserving further research. Our survey constitutes an iTRAQ-based comparative proteomic analysis of a virulent M. hyopneumoniae strain and its attenuated strain originating from a single parent with a well-characterized genetic background and lays the groundwork for future work to mine for potential virulence factors and identify candidate vaccine proteins.

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Recombinant 46-kilodalton surface antigen (P46) of Mycoplasma hyopneumoniae expressed in Escherichia coli can be used for early specific diagnosis of mycoplasmal pneumonia of swine by enzyme-linked immunosorbent assay

Cited by 26 publications

References 19 publications

Use of a Mycoplasma Hyopneumoniae Nested Polymerase Chain Reaction Test to Determine the Optimal Sampling Sites in Swine

Use of a Mycoplasma Hyopneumoniae Nested Polymerase Chain Reaction Test to Determine the Optimal Sampling Sites in Swine

Detection of Mycoplasma hyopneumoniae by Air Sampling with a Nested PCR Assay

Quantitative Proteomic Analyses of a Pathogenic Strain and Its Highly Passaged Attenuated Strain ofMycoplasma hyopneumoniae

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