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Mardassi BB Béjaoui Khiari A Oussaief L Landoulsi A Brik C Mlik B Amouna F 《Veterinary microbiology》2007,119(1):31-41
A recombinant phage library harbouring Mycoplasma meleagridis (MM) genomic DNA fragments was generated in the bacteriophage lambda gt11 expression vector. The library was screened for expression of MM specific antigens with a polyclonal antiserum that had been preadsorbed with antigens of the most common unrelated avian mycoplasma species. A 49-amino acid antigenic domain unique to MM was isolated, expressed in Escherichia coli, and its serodiagnostic potential was demonstrated. An antiserum raised against this MM-specific antigenic domain recognized a cluster of seven membrane-associated MM proteins with molecular masses ranging from 34 to 75 kDa. Overall, this study resulted in the identification of a potent serodiagnostic tool and revealed the complex antigenic nature of MM. 相似文献
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Béjaoui Khiari A Landoulsi A Aissa H Mlik B Amouna F Ejlassi A Ben Abdelmoumen Mardassi B 《Avian diseases》2011,55(1):8-12
Mycoplasma meleagridis (MM) is a major cause of disease and economic loss in turkeys. Formerly it was thought that this species was very host specific and only restricted to turkey. In this study, we report on the recovery of MM from breeding flocks of chickens located near a turkey breeding unit. Ten MM field strains were isolated (by culture on Frey broth medium) from tracheal swabs of chickens displaying clinical signs of mycoplasmosis-essentially respiratory symptoms and poor performance. Assignment of the isolated field strains to MM was confirmed by a growth inhibition assay using MM-specific polyclonal antiserum and by PCR amplification targeting the 16S rRNA sequence as well as the Mm14 sequence, a MM-species-specific DNA fragment previously identified and characterized in our laboratory. The nucleotide sequence of Mm14 proved to be highly conserved among the 10 MM field strains, indicating a common source of infection. However, on the basis of slight differences in sodium dodecyl sulfate-polyacrylamide gel electrophoresis whole-cell proteins and western blot profiles, two groups of the isolated MM field strains could be distinguished. Evidence of MM infection of chickens was further provided by serology, since 13.77% (35/254) of sera proved positive to MM by either rapid serum agglutination or recombinant antigen-based enzyme-linked immunosorbent assay. In addition, sera of all chickens from which MM was isolated were positive for antibodies to MM. Collectively, the data unambiguously show that MM could infect chickens; thus, MM warrants further exploration to determine its pathogenicity in this unusual host. 相似文献
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Lyes Beninal Zouaoui Bouznad Roselyne Corbière Sihem Belkhiter Romain Mabon Abdelmoumen Taoutaou Abdelaziz Keddad Eve Runno-Paurson Didier Andrivon 《Plant pathology》2022,71(2):458-469
Potato is one of the most important agricultural crops in Algeria and worldwide. Each year, potato late blight, caused by Phytophthora infestans, is responsible for significant damage that leads to large production losses, and is thus a direct threat to food security in Algeria. In this study, 131 isolates of P. infestans and 92 DNA fingerprints captured on FTA cards were sampled from commercial and seed production fields in three major potato production regions (western, eastern, and central) during the main-season and late-season in Algeria over six cropping seasons (2010–2016). Genotypes of P. infestans and population genetic diversity were analysed using a 17-plex simple-sequence repeat (SSR) marker assay, and the mating type of all isolates was characterized. Both mating types (A1 and A2) were found, and often occurred in the same field. Differences in mating type proportion were observed between regions and between sampling periods. Analysis with SSR markers showed the prevalence of the EU_13_A2 lineage (70%) over EU_2_A1 (16%), EU_23_A1 (10%), and 4% of unknown multilocus lineage (MLL). The EU_13_A2 showed differentiation within the group. EU_23_A1 was found mainly in late-season crops. However, the cropping region did not influence the distribution of lineages due to the dispersal of the pathogen in Algeria by seeds. Genetic structure did not reveal a clear variation in distribution of the three lineages throughout the sampling regions. These data provide important new information on the composition and change over time of P. infestans populations in Algeria and open the way for a better understanding of the local epidemiology of this important pathogen. 相似文献
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B Ben Abdelmoumen R S Roy R Brousseau 《Journal of veterinary diagnostic investigation》1999,11(2):162-169
A genomic library of Mycoplasma synoviae (MS) was generated by using bacteriophage lambda gt11 as a cloning and expression vector. Identification of recombinant clones highly specific to MS was achieved by screening the library for expression of MS proteins with polyclonal antiserum that had been preadsorbed with 6 heterologous avian mycoplasma species antigens. Expression of the recombinant clones in Escherichia coli followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the total cell lysates and immunoblot yielded a predominant reactive fusion protein of 165 kD. Two clones (MS2/28 and MS2/12) that yielded inserts of different size were selected. The 2 MS DNA inserts were subcloned in a plasmid vector, labeled with digoxigenin, and used as probes for the specific recognition of several MS strains. A high degree of conservation was demonstrated for the MS2/12 and MS2/28 genes in tested MS strains. In addition, neither DNA fragment recognized any other avian mycoplasma species (M. gallisepticum, M. meleagridis, M. gallinarum, M. iners, M. anatis, and M. iowae), thus indicating their high specificity to MS. The sensitivity of the slot blot hybridization method using digoxigenin-labeled MS2/12 and MS2/28 probes for direct detection of MS from broth cultures of field isolates was 10(5) colony-forming units/ml. These results demonstrate the effectiveness of adsorbed antisera for the isolation of species-specific mycoplasma DNA and the potential for its use as probes for the specific and direct detection of MS from broth cultures of field isolates. 相似文献
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We have previously identified species-specific DNA fragments, referred to as MS2/28 and Mm14, of Mycoplasma synoviae and Mycoplasma meleagridis, respectively. In the present study, we extended our analysis of the MS2/28 fragment that was found to encode a species-specific antigenic site, and we demonstrated the specificity of the Mycoplasma gallisepticum hemagglutinin protein encoded by pMGA1.2 (a member of the vlhA gene family). Then, we combined the Escherichia coli-expressed products of MS2/28, Mm14, and pMGA1.2, to develop a recombinant antigen-based enzyme-linked immunosorbent assay (recELISA), for the simultaneous and specific detection of antibodies to the three aforementioned major avian mycoplasma species. For comparative purposes, a novel in-house crude antigen capture ELISA (capELISA) was developed in parallel. In the latter protocol, the microtiter wells were enriched in species-specific antigens by capturing sonicated crude antigens on coated rabbit polyclonal antibodies that had been extensively adsorbed with the whole antigen of the heterologous species. With regard to rapid serum agglutination, both ELISA tests were highly specific, and they showed a significant correlation when field sera from naturally infected birds were tested. recELISA proved to be highly specific because absorbance values, with the heterologous species, were significantly lower (P < 0.001) than those obtained with capELISA. Given its cost-effectiveness and simplicity, the recombinant antigen-based ELISA seems to represent a valid tool for the specific screening of the three major avian mycoplasma species. recELISA will be particularly useful with regard to trade control because a large number of samples from various fields could be rapidly processed. 相似文献
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The Mycoplasma synoviae haemagglutinin gene, vlhA, encodes two major immunodominant and surface-exposed membrane proteins, MSPB and MSPA. Both products are antigenically variable but only MSPA mediates binding to erythrocytes. Previously we have shown that M. synoviae type strain WVU 1853 could express a variant vlhA gene, referred to as MS2/28.1, with a considerably shorter and divergent MSPA region. A finding that prompted detailed characterization of its antigenic and functional properties. Here we mutagenized each of the six opal codons of the variant MS2/28.1 vlhA member into tryptophan, thus allowing its expression in Escherichia coli as well as its cleavage products, MSPB and MSPA. In addition, we expressed 5 contiguous regions of MS2/28.1 extending from the last part of MSPB to the C-terminus of MSPA. Colony immunostaining with region-specific antisera mapped antigenic variation to the N-terminal half of MS2/28.1 MSPA. No haemagglutinating activity was observed for MSPB, but consistent haemadsorption inhibition was mapped to the region extending from amino acid 325 to 344. Inhibition of both haemagglutination and haemadsorption activities were obtained with sera directed against the C-terminal region of MSPA, with the highest titers (1/320 and 1/160, respectively) being recorded for its last 60 residues. Importantly, antibodies to this region also yielded the highest metabolic inhibition titer of 1/1280. Overall, aside from mapping the functional domains of a M. synoviae highly divergent haemagglutinin gene, this study shows that the C-terminal half of its MSPA region induced the highest titers of antibodies inhibiting haemagglutination, haemadsorption, and metabolism. 相似文献
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