Development and Application of a Polymerase Chain Reaction Assay for Mycoplasma synoviae

Lauerman, L. H.; Hoerr, Frederic J.; Sharpton, Alan R.; Shah, Syed Mohmad; Santen, Vicky L. van

doi:10.2307/1592037

Cited by 91 publications

(61 citation statements)

References 12 publications

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“…However, the latter is generally expensive and time consuming (requiring 1-2 weeks to complete). Recently, PCR has been employed for identification of suspected cultures or for rapid detection of M. synoviae directly from clinical samples (Garcia et al, 1995;Kiss et al, 1997;Lauerman et al, 1993;Silveira et al, 1996;Wang et al, 1997). Further identification of the M. synoviae isolates (i.e.…”

Section: Introductionmentioning

confidence: 99%

Classification of Mycoplasma synoviae strains using single-strand conformation polymorphism and high-resolution melting-curve analysis of the vlhA gene single-copy region

et al. 2007

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Mycoplasma synoviae is an economically important pathogen of poultry worldwide, causing respiratory infection and synovitis in chickens and turkeys. Identification of M. synoviae isolates is of critical importance, particularly in countries in which poultry flocks are vaccinated with the live attenuated M. synoviae strain MS-H. Using oligonucleotide primers complementary to the single-copy conserved 59 end of the variable lipoprotein and haemagglutinin gene (vlhA), amplicons of~400 bp were generated from 35 different M. synoviae strains/isolates from chickens and subjected to mutation scanning analysis. Analysis of the amplicons by single-strand conformation polymorphism (SSCP) revealed 10 distinct profiles (A-J). Sequencing of the amplicons representing these profiles revealed that each profile related to a unique sequence, some differing from each other by only one base-pair substitution. Comparative high-resolution melting (HRM) curve analysis of the amplicons using SYTO 9 green fluorescent dye also displayed profiles which were concordant with the same 10 SSCP profiles (A-J) and their sequences. For both mutation detection methods, the Australian M. synoviae strains represented one of the A, B, C or D profiles, while the USA strains represented one of the E, F, G, H, I or J profiles. The results presented in this study show that the PCR-based SSCP or HRM curve analyses of vlhA provide high-resolution mutation detection tools for the detection and identification of M. synoviae strains. In particular, the HRM curve analysis is a rapid and effective technique which can be performed in a single test tube in less than 2 h.

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Section: Introductionmentioning

confidence: 99%

Classification of Mycoplasma synoviae strains using single-strand conformation polymorphism and high-resolution melting-curve analysis of the vlhA gene single-copy region

et al. 2007

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“…(Table 3). There are earlier reports on PCR assays targeting the 16S rRNA gene regions [1,9,10] , whereas more recent assays attempted to target more species-specific gene regions [4,5,8,13] . rRNA genes are present in all prokaryotes and include regions that are highly conserved among bacteria.…”

Section: Mg and Ms Rpcrsmentioning

confidence: 99%

“…There are different PCR procedures such as conventional and real time PCRs (rPCR) for MG or MS detection and their advantages and disadvantages have previously been discussed [1,4,5,[7][8][9][10] .…”

Section: Introductionmentioning

confidence: 99%

Damızlık Tavuk Kümeslerinde Mycoplasma gallisepticum ve Mycoplasma synoviae’nin Gerçek Zamanlı PCR’lar ile ve Mycoplasma gallisepticum Antikorlarının ELISA ile Tespiti

Kahya¹,

Yılmaz²,

Eyigör³

et al. 2015

Kafkas Univ Vet Fak Derg

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This study aimed to determine the prevalence of Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS) in breeder flocks showing respiratory symptoms. A total of 77 flocks (2153 tracheal swabs and blood samples) were sampled and all were tested by MG real time PCR (MG-rPCR) and MG-ELISA, and 32 flocks were tested by MS real time PCR (MS-rPCR). In the first part of this study covering 28 flocks, all samples from chickens with marked clinical symptoms and high MG-antibody levels gave negative results with MG-rPCR1. Therefore, the MG-lipoprotein gene-specific primers (MG-rPCR1) of this PCR were replaced with MG-16S rRNA primers (MG-rPCR2), as were the MS-16S rRNA primers (MS-rPCR), thus the study was pursued accordingly. All of the first 28 flocks, which were 100% positive by MG-ELISA, were MG-rPCR1 negative, whereas in the second part of the study, other 49 flocks, which were 87.8% MG seropositive, were found 42.9% positive by MG-rPCR2. In addition, 5 selected flocks from the first 28 were negative, whereas 7.4% of the 27 selected flocks from the second 49 were positive by MS-rPCR. Overall, 81 out of 432 MG-rPCR1-2 (18.7%) performed from 77 flocks, and 13 out of 187 MS-rPCRs (6.9%) in 32 flocks were determined as positive. ELISA results indicated that there could be significantly high false-positives in serological tests, thus results should not be relied upon one test system. Also, this study revealed that, for the confirmation of Mycoplasma-infected flocks in laboratories, rPCR is a reliable method as long as suitable primers are selected, and that MG and MS prevalence is considerably high in winter season. Keywords

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“…The samples were used in a polymerase chain reaction (PCR) to detect MS, in the laboratory of Embrapa Suínos Aves, in Concórdia, SC, according to the technique described by Lauerman et al (1993). Swabs with plastic stem and cotton tip was introduced within the trachea, the cotton tip was immediately immersed in 0.5mL of phosphate buffered saline (PBS; 150mM NaCl, 2.6mM NaH 2 PO 4 , 7.4mM Na 2 HPO 4 , pH 7.5) and sent to the laboratory under refrigeration.…”

Section: Polymerase Chain Reaction (Pcr)mentioning

confidence: 99%

“…Serum plate agglutination (SPA), hemagglutination inhibition (HI) and ELISA are the most common serological techniques. Direct diagnosis requests isolation and identification of the agent in selective culture media (Kleven, 1997) or the demonstration of the DNA of the pathogen in the host using the polymerase chain reaction (PCR) (Lauerman et al, 1993).…”

Section: Introductionmentioning

confidence: 99%

Test profiles of broiler breeder flocks housed in farms with endemic Mycoplasma synoviae infection

Fiorentin

Trevisol

et al. 2003

Rev. Bras. Cienc. Avic.

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There is a need for a better understanding of the epidemiology of Mycoplasma synoviae (MS) infection in broiler breeders in Brazil. Many features of the infection remain unrecognizable, because there are no clinical signs of the disease. A detailed testing was performed at each 6 to 8 weeks in three MS-free flocks introduced in farms with endemic MS infection for a follow-up epidemiological study. Every flock was monitored by polymerase chain reaction (PCR), by serum plate agglutination (SPA) and hemagglutination inhibition (HI) for serology studies, and isolation of mycoplasmas from tracheal swabs. PCR was found to be the most sensitive test, detecting early MS infection. Serology was positive in less than 50% of the sera and MS was isolated only between 27 and 28 weeks of age and in a maximum of 60% positive hens. A similar profile was seen for MS infection in all three flocks. Infection started at brooding, whereas laboratory detection of the assymptomatic infection was more probable in the weeks of increasing egg production. This predictable profile during rearing may be very useful for the optimization of monitoring MS infection in broiler breeder flocks.

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Development and Application of a Polymerase Chain Reaction Assay for Mycoplasma synoviae

Cited by 91 publications

References 12 publications

Classification of Mycoplasma synoviae strains using single-strand conformation polymorphism and high-resolution melting-curve analysis of the vlhA gene single-copy region

Classification of Mycoplasma synoviae strains using single-strand conformation polymorphism and high-resolution melting-curve analysis of the vlhA gene single-copy region

Damızlık Tavuk Kümeslerinde Mycoplasma gallisepticum ve Mycoplasma synoviae’nin Gerçek Zamanlı PCR’lar ile ve Mycoplasma gallisepticum Antikorlarının ELISA ile Tespiti

Test profiles of broiler breeder flocks housed in farms with endemic Mycoplasma synoviae infection

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