Papers by François Thiaucourt
Proteomic characterization of pleural effusion, a specific host niche of Mycoplasma mycoides subsp. mycoides from cattle with contagious bovine pleuropneumonia (CBPP)
Journal of Proteomics, 2015
Free exopolysaccharide from Mycoplasma mycoides subsp. mycoides possesses anti-inflammatory properties
Veterinary Research, 2015
In this study we explored the immunomodulatory properties of highly purified free galactan, the s... more In this study we explored the immunomodulatory properties of highly purified free galactan, the soluble exopolysaccharide secreted by Mycoplasma mycoides subsp. mycoides (Mmm). Galactan was shown to bind to TLR2 but not TLR4 using HEK293 reporter cells and to induce the production of the anti-inflammatory cytokine IL-10 in bovine macrophages, whereas low IL-12p40 and no TNF-α, both pro-inflammatory cytokines, were induced in these cells. In addition, pre-treatment of macrophages with galactan substantially reduced lipopolysaccharide (LPS)-induced production of pro-inflammatory cytokines TNF- and IL-12p40 while increasing LPS-induced secretion of immunosuppressive IL-10. Also, galactan did not activate naïve lymphocytes and induced only low production of the Th1 cytokine IFN-γ in Mmm-experienced lymphocytes. Finally, galactan triggered weak recall proliferation of CD4+ T lymphocytes from contagious bovine pleuropneumonia-infected animals despite having a positive effect on the expression of co-stimulatory molecules on macrophages. All together, these results suggest that galactan possesses anti-inflammatory properties and potentially provides Mmm with a mechanism to evade host innate and adaptive cell-mediated immune responses.
Whole Blood Transcriptome Analysis of Mycoplasma mycoides Subsp. mycoides-Infected Cattle Confirms Immunosuppression but Does Not Reflect Local Inflammation
PLOS ONE, 2015
Contagious bovine pleuropneumonia (CBPP), caused by Mycoplasma mycoides subsp. mycoides (Mmm), is... more Contagious bovine pleuropneumonia (CBPP), caused by Mycoplasma mycoides subsp. mycoides (Mmm), is a severe respiratory disease of cattle responsible for major economic losses in sub-Saharan Africa. Disease control relies mainly on the use of empirically attenuated vaccines that provide limited protection. Thus, understanding the virulence mechanisms used by Mmm as well as the role of the host immune system in disease development, persistence, and control is a prerequisite for the development of new, rationally designed control strategies. The aim of this study was to assess the use of whole blood transcriptome analysis to study cattle-Mmm interactions, starting by the characterization of the bovine response to Mmm infection during the acute form of the disease. For that purpose, we compared the transcriptome profile of whole blood from six cattle, before challenge by contact with Mmm-infected animals and at the appearance of first clinical signs, using a bovine microarray. Functional analysis revealed that 680 annotated genes were differentially expressed, with an overwhelming majority of down-regulated genes characterizing an immunosuppression. The main bio-functions affected were "organismal survival", "cellular development, morphology and functions" and "cell-to cell signaling and interactions". These affected functions were consistent with the results of previous in vitro immunological studies. However, microarray and qPCR validation results did not highlight pro-inflammatory molecules (such as TNFα, TLR2, IL-12B and IL-6), whereas inflammation is one of the most characteristic traits of acute CBPP. This global gene expression pattern may be considered as the result, in blood, of the local pulmonary response and the systemic events occurring during acute CBPP. Nevertheless, to understand the immune events occurring during disease, detailed analyses on the different immune cell subpopulations, either in vivo, at the local site, or in vitro, will be required. Whole blood transcriptome analysis remains an interesting approach for the identification of bio-signatures correlating to recovery and protection, which should facilitate the evaluation and validation of novel vaccine formulations.
Veterinary Research, 2015
Contagious caprine pleuropneumonia (CCPP), caused by Mycoplasma capricolum subsp. capripneumoniae... more Contagious caprine pleuropneumonia (CCPP), caused by Mycoplasma capricolum subsp. capripneumoniae (Mccp), is a devastating disease of domestic goats and of some wild ungulate species. The disease is currently spreading in Africa and Asia and poses a serious threat to disease-free areas. A comprehensive view of the evolutionary history and dynamics of Mccp is essential to understand the epidemiology of CCPP. Yet, analysing the diversity of genetically monomorphic pathogens, such as Mccp, is complicated due to their low variability. In this study, the molecular epidemiology and evolution of CCPP was investigated using a large-scale genomic approach based on next-generation sequencing technologies, applied to a sample of strains representing the global distribution of this disease. A highly discriminatory multigene typing system was developed, allowing the differentiation of 24 haplotypes among 25 Mccp strains distributed in six genotyping groups, which showed some correlation with geographic origin. A Bayesian approach was used to infer the first robust phylogeny of the species and to date the principal events of its evolutionary history. The emergence of Mccp was estimated only at about 270 years ago, which explains the low genetic diversity of this species despite its high mutation rate, evaluated at 1.3 × 10 −6 substitutions per site per year. Finally, plausible scenarios were proposed to elucidate the evolution and dynamics of CCPP in Asia and Africa, though limited by the paucity of Mccp strains, particularly in Asia. This study shows how combining large-scale genomic data with spatial and temporal data makes it possible to obtain a comprehensive view of the epidemiology of CCPP, a precondition for the development of improved disease surveillance and control measures.
Acta veterinaria Scandinavica, 2004
Contagious caprine pleuropneumonia (CCPP) is a major threat to goat farming in parts of Africa an... more Contagious caprine pleuropneumonia (CCPP) is a major threat to goat farming in parts of Africa and Asia. It classically causes acute high morbidity and mortality early in infection, but little is known of its long term epizootiology and course. In this study, 10 goats were inoculated with Mycoplasma capricolum subsp. capripneumoniae (M. capripneumoniae) and then mixed with 15 goats for contact transmission. The disease course was monitored in each goat for 56-105 days, whereafter the goats were killed and necropsied. Varying features signifying infection occurred in altogether 17 goats (7 inoculated, 10 in-contact). Clinical signs were severe in 8 goats but no fatalities occurred. Only 6 goats had serum antibody titres against M. capripneumoniae in ELISA. Fourteen goats (5 inoculated, 9 in-contact) had chronic pleuropulmonary lesions compatible with CCPP at necropsy and 7 of those showed M. capripneumoniae antigen in the lung by immunohistochemistry. Neither cultivation nor PCR test...
Clinical and diagnostic laboratory immunology, 1999
The genes encoding the 62-kDa lipoproteins from the Mycoplasma mycoides subsp. mycoides large-col... more The genes encoding the 62-kDa lipoproteins from the Mycoplasma mycoides subsp. mycoides large-colony type (LC) strain Y-goat and the M. mycoides subsp. capri strain PG3 were cloned and analyzed by sequencing. These two lipoproteins have been named LppA[MmymyLC] and LppA[Mmyca], and their corresponding genes have been named lppA[MmymyLC] and lppA[Mmyca], respectively. The nucleotide and deduced amino acid sequences of these two lipoproteins showed a very high degree of similarity between these two mycoplasmas. Given the sequence data, LppA seems to fulfill the same structural functions as the previously described major lipoproteins P72 of M. mycoides subsp. mycoides small-colony type and P67 of the Mycoplasma species bovine group 7. Based on lppA gene sequences of M. mycoides subsp. mycoides LC and M. mycoides subsp. capri type strains, a specific PCR assay was developed so that it amplified this gene in all field strains of the two species analyzed in this study but not in the other...
Veterinary Microbiology, 2001
A monoclonal antibody (Mab 4.52) raised against Mycoplasma capricolum subsp. capripneumoniae (Mcc... more A monoclonal antibody (Mab 4.52) raised against Mycoplasma capricolum subsp. capripneumoniae (Mccp) cell lysate was used as a template to obtain substitute antigens recognised by its paratope. Two approaches were investigated: a 17-mer random peptide library displayed on the surface of a ®lamentous phage was screened by panning on the immobilised Mab 4.52 and antiidiotype antibodies were generated by immunising a chicken with the F(ab H ) 2 fragments of the antibody. Analysis of the peptide sequences displayed by the isolated phages identi®ed two peptides. Both contained two cysteine residues and had identical or similar amino acids in positions 5 (P), 8 (I/L) and 13 (L). The fusion phages were also recognised by Mab 4.52 in enzyme-linked immunosorbent assay (ELISA) and binding was shown by surface plasmon resonance. One of the peptides was a markedly better inhibitor (67%) of the binding of Mab 4.52 to its original antigen than the other (20%) at 1 mg/ml. After absorption, to remove isotypic and allotypic reactivities, the anti-idiotype IgY was speci®cally recognised by Mab 4.52 in ELISA and was able to inhibit its binding to the original antigen, whereas anti-idiotype antibodies raised against a bluetongue virus-speci®c antibody had no effect. In spite of unequivocal binding of the anti-idiotype antibodies and the fusion phages to the paratope of Mab 4.52, goat antisera appeared not to react with either of the surrogate antigens. In contrast, the test sera bound to the original antigen suggesting that Mab 4.52 Veterinary Microbiology 81 (2001) 165±179 : S 0 3 7 8 -1 1 3 5 ( 0 1 ) 0 0 3 3 8 -8 does not recognise exactly the same antigenic site as antibodies in the goat antisera. #
An adenoviral vector expressing lipoprotein A, a major antigen of Mycoplasma mycoides subspecies mycoides, elicits robust immune responses in mice
Vaccine, 2015
Contagious bovine pleuropneumonia (CBPP), caused by Mycoplasma mycoides subsp. mycoides small col... more Contagious bovine pleuropneumonia (CBPP), caused by Mycoplasma mycoides subsp. mycoides small colony type (MmmSC), is a devastating respiratory disease of cattle. In sub-Saharan Africa, where CBPP is enzootic, live attenuated vaccines are deployed but afford only short-lived protection. In cattle, recovery from experimental MmmSC infection has been associated with the presence of CD4(+) T lymphocytes that secrete interferon gamma in response to MmmSC, and in particular to the lipoprotein A (LppA) antigen. In an effort to develop a better vaccine against CBPP, a viral vector (Ad5-LppA) that expressed LppA was generated from human adenovirus type 5. The LppA-specific immune responses elicited by the Ad5-LppA vector were evaluated in mice, and compared to those elicited by recombinant LppA formulated with a potent adjuvant. Notably, a single administration of Ad5-LppA, but not recombinant protein, sufficed to elicit a robust LppA-specific humoral response. After a booster administratio...
Veterinary Research, 2008
A better understanding of protective immune memory against contagious bovine pleuropneumonia (CBP... more A better understanding of protective immune memory against contagious bovine pleuropneumonia (CBPP) is needed in order to facilitate the development of safer vaccines based on selected components of the pathogen. For this purpose, cells collected from lymph nodes draining the lungs of Mycoplasma mycoides subsp. mycoides small colony biotype (MmmSC)-infected cattle were stimulated with the pathogen in vitro and evaluated concurrently for proliferation (CFSE based method), expression of activation, memory markers and cytokine production. Direct evidence is presented for a major contribution of CD4 + T cells to the vigorous proliferative and T1 biased cytokine recall responses observed in cattle that have recovered from infection but not in animals developing the acute form of the disease. Two different phenotypes of MmmSC-specific memory CD4 were observed based on CD62L expression and proliferative capacities. Furthermore, recall proliferation of B cells also occurred but was strictly dependent on the presence of CD4. The information provided in this study will facilitate the search for MmmSC antigens that have potential for the development of subunit vaccines against CBPP. contagious bovine pleuropneumonia / Mycoplasma mycoides subsp. mycoides SC / vaccine / memory / CD4
Veterinary Microbiology, 2003
Contagious caprine pleuropneumonia (CCPP) is a major threat to goat farming in developing countri... more Contagious caprine pleuropneumonia (CCPP) is a major threat to goat farming in developing countries. Its exact distribution is not well known, despite the fact that new diagnostic tools such as PCR and competitive ELISA are now available. The authors developed a study of the molecular epidemiology of the disease, based on the ampli®cation of a 2400 bp long fragment containing two duplicated gene coding for a putative membrane protein. The sequence of this fragment, obtained on 19 Mycoplasma capricolum subsp. capripneumoniae (Mccp) strains from various geographical locations, gave 11 polymorphic positions. The three mutations found on gene H2prim were silent and did not appear to induce any amino acid modi®cations in the putative translated protein. The second gene may be a pseudogene not translated in vivo, as it bore a deletion of the ATG codon found in the other members of the``Mycoplasma mycoides cluster'' and as the six mutations evidenced in the Mccp strains would induce modi®cations in the translated amino acids. In addition, an Mccp strain isolated in the United Arab Emirates showed a deletion of the whole pseudogene, a further indication that this gene is not compulsory for mycoplasma growth. Four lineages were de®ned, based on the nucleotide sequence. These correlated relatively well with the geographical origin of the strains: North, Central or East Africa. The strain of Turkish origin had a Veterinary Microbiology 85 (2002) 111±123 : S 0 3 7 8 -1 1 3 5 ( 0 1 ) 0 0 5 0 9 -0 sequence similar to that found in North African strains, while strains isolated in Oman had sequences similar to those of North or East African strains. The latter is possibly due to the regular import of goats of various origins. Similar molecular epidemiology tools have been developed by sequencing the two operons of the 16S rRNA gene or by AFLP. All these various techniques give complementary results. One (16S rRNA) offers the likelihood of a ®ner identi®cation of strains circulating in a region, another (H2) of determining the geographical origin of the strains. These tools can make a very useful contribution to understanding the epidemiology of CCPP. #
Veterinary Microbiology, 1994
Veterinary Microbiology, 2000
The Mycoplasma mycoides cluster is made of six species that are closely related both genetically ... more The Mycoplasma mycoides cluster is made of six species that are closely related both genetically and phenotypically. Two are of particular importance, M. mycoides subsp. mycoides SC causing contagious bovine pleuropneumonia and M. capricolum subsp. capripneumoniae causing contagious caprine pleuropneumonia. The sequences of a putative membrane protein gene and partial¯anking open reading frames have been obtained from various strains in this cluster, including all reference strains. Sequence analysis showed this locus is present and fully conserved in all strains of M. mycoides subsp. mycoides SC isolated from geographically most distant places worldwide. In M. capricolum subsp. capripneumoniae polymorphism in this locus has been found at seven positions and revealed that they can be used as epidemiological markers. Conserved regions were used to de®ne a primer pair that enables the ampli®cation by PCR of two fragments 302 and 1298 bp long, respectively. The 302 bp long fragment contains an intergenic sequence that can be used for phylogenetic studies or for identi®cation purposes. Parsimony analysis on an alignment of 49 DNA sequences show a subdivision of the M. mycoides cluster into two subgroups that is in accordance with results obtained by phenotypic methods. Two lineages exist within the capricolum subgroup, one of them clustering strains identi®ed as M. capricolum subsp. capricolum, M. capricolum subsp. capricolum and M. sp Bovine Group 7. However M. capricolum subsp. capripneumoniae strains can readily be identi®ed by three speci®c nucleotide positions or by sequencing the 1298 bp long fragment. There is no clear subdivision within the mycoides subgroup, supporting the idea that M. mycoides subsp. mycoides LC and M. mycoides subsp. capri should not be separated into two subspecies. Mycoplasma mycoides subsp. mycoides SC strains can easily be Veterinary Microbiology 72 (2000) 251±268 : S 0 3 7 8 -1 1 3 5 ( 9 9 ) 0 0 2 0 4 -7 distinguished as they bear an insertion sequence 15 bp downstream from the stop codon of the membrane protein gene. #
Veterinary Microbiology, 1999
The lppA gene, encoding the lipoprotein named LppA [Mcaca] was characterised in Mycoplasma capric... more The lppA gene, encoding the lipoprotein named LppA [Mcaca] was characterised in Mycoplasma capricolum subsp. capricolum. It encodes a lipoprotein with an apparent molecular mass of 57 kDa as determined by SDS-PAGE. Using antibodies directed against recombinant LppA [Mcaca], we showed the expression of this lipoprotein in all M. capricolum subsp. capricolum by immunoblot analysis. The serum did not cross-react with other members of the Mycoplasma mycoides cluster, hence showing that LppA [Mcaca] was antigenically specific to M. capricolum subsp. capricolum. The lppA gene was conserved within the subspecies and was used for the development of a specific PCR assay for the identification of M. capricolum subsp. capricolum. The taxonomically related Mycoplasma capricolum subsp. capripneumoniae (F38) was found to contain an lppA-pseudo-gene. It showed high similarity to functional lppA genes of other mycoplasmas in the M. mycoides cluster. However, it contained interrupted open reading frames. Moreover, the nucleotide sequence of the lppA pseudo-genes in different strains of M. capricolum subsp. capripneumoniae were quite variable. Interestingly, the lppA pseudo-gene had a size similar to that of the functional lppA genes of other mycoplasmas of the M. mycoides cluster and occupied the same genomic location as the latter ones in the vicinity of the mtlD genes. This study showed that all members of the M. mycoides cluster contain each a species-, subspecies-respectively type-specific lppA gene analogue which encodes a lipoprotein that has structural and functional relationship to the surface lipoprotein LppA [MmymySC], previously named P72, of M. mycoides subsp mycoides SC, with the exception Veterinary Microbiology 69 (1999) 157±172 (J. Frey)
Veterinary Microbiology, 2010
Please cite this article in press as: Nwankpa, N.D., et al., Variable Number Tandem Repeat (VNTR)... more Please cite this article in press as: Nwankpa, N.D., et al., Variable Number Tandem Repeat (VNTR) analysis reveals genetic diversity within Mycoplasma mycoides mycoides Small Colony isolates from Nigeria.
Veterinary Immunology and Immunopathology, 1997
The Veterinary Journal, 2005
Diagnostic differentiation between the ruminant pathogens Mycoplasma agalactiae and Mycoplasma bo... more Diagnostic differentiation between the ruminant pathogens Mycoplasma agalactiae and Mycoplasma bovis is known to be problematic when only conventional serological and biochemical tests are used. The main reason for this is that both agents share a considerable number of related proteins and common epitopes. DNA-based detection methods offer advantages in terms of specificity and sensitivity. However, there is an urgent need to compare currently used PCR assays because they target different genomic regions and, therefore, may perform differently. In the present work, five laboratories, which use PCR routinely, evaluated the specificity of four different PCR systems for M. agalactiae and three systems for M. bovis on a total of 41 strains of the two Mycoplasma species including six previously unidentified strains. As the vast majority of PCR examinations (97.1% of all tests) correctly identified the strains the specificity of all seven detection systems appears to be high. In four cases, incorrect identification by conventional diagnostic methods was rectified by PCR. Isolates from non-typical hosts, i.e. three M. bovis strains from small ruminants and two M. agalactiae strains from cattle, were characterised by sequencing the 16S and part of the 23S ribosomal RNA genes.
Preventive Veterinary Medicine, 2011
Contagious bovine pleuropneumonia (CBPP) is a severe respiratory disease of cattle and buffalo ca... more Contagious bovine pleuropneumonia (CBPP) is a severe respiratory disease of cattle and buffalo caused by Mycoplasma mycoides subsp. mycoides "Small Colony" (MmmSC). The agent of CBPP has been isolated from goats in different countries including CBPP-free areas. Goats can therefore be regarded as a putative MmmSC reservoir. No diagnostic test for CBPP surveillance in goats has been proposed as yet. Furthermore, serological tests could be seriously hampered by a widespread caprine infection due to the subspecies M. mycoides subsp. capri (Mmc), which is antigenically very close to MmmSC and displays high levels of genetic variability. A competition ELISA (cELISA) is currently used to screen for CBPP in cattle at the herd level in infected areas. The aim of this study was to see if the same cELISA would be specific enough to be used to screen goats despite the potential concomitant infection with Mmc.
Preventive Veterinary Medicine, 2004
Contagious bovine pleuropneumonia (CBPP) is a respiratory disease of cattle; CBPP is caused by My... more Contagious bovine pleuropneumonia (CBPP) is a respiratory disease of cattle; CBPP is caused by Mycoplasma mycoides subsp. mycoides small colony. CBPP is a major cause for concern for African countries (because of mortality, animal-production losses and cost of control). The clinical form of the disease is the more infectious (contagion occurs essentially through coughing). However, chronic lung lesions with viable mycoplasmas can persist in recovering cattle. Animals presenting these lesions might have a time-delimitated infectious phase. Such carriers are suspected to generate field outbreaks (although this hypothesis remains debated). We investigated the potential quantitative effects of these chronic carriers on the within-herd CBPP spread. Data were collected during a longitudinal field herd survey in a mixed crop-livestock system in the Ethiopian highlands. Two stochastic Markov-chain models' outputs (seroconversion dynamics, basic reproduction ratio R 0 , cumulative clinical incidence and risk of herd infection) were compared given different hypotheses on the carrier infectiousness. The late seroconversions observed in the field data were fitted correctly only for the highest carrier infectiousness we considered (mean chronic duration of 1 year and carriers 50-times less infectious than clinical cases). Although sensitivities (in terms of disease impact in the herd) were in general negligible when the carrier infectiousness was low (e.g. when carriers were assumed to be
Preventive Veterinary Medicine, 2004
Contagious bovine pleuropneumonia (CBPP) is a major threat for cattle health and production in Af... more Contagious bovine pleuropneumonia (CBPP) is a major threat for cattle health and production in Africa. This disease is caused by the small-colony type of Mycoplasma mycoides subspecies mycoides (MmmSC). Transmission occurs from direct and repeated contacts between sick and healthy animals. Veterinary services recently reported a resurgence of CBPP in the province of West Wellega, in the Ethiopian highlands. A research program was set up to estimate the epidemiological parameters of the within-herd infection spread. A follow-up survey was implemented in 71 sampled herds of the Boji district (West Wellega province). Fifteen herds were classified as newly infected and used in a serological-and clinical-incidence study. The overall 16-month cumulative sero-incidence risk was 34%. Clinical cases were recorded for 39% of the seropositive cattle; case-fatality risk was 13%. There was no evidence of benefit on infection spread of CBPP-control measures used locally by farmers (isolation or antibiotic treatments of sick animals). This might be related to a lack of power in the statistical analyses or to a quality problem for the medications used (and more generally, for health-care delivery in the Boji district).
PLoS ONE, 2012
Mycoplasma mycoides subsp.
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Papers by François Thiaucourt