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F Thiaucourt

Publications and source records attributed to F Thiaucourt.

At least 19 recordsLinked to original sources
PubMed2003

Molecular epidemiology of contagious bovine pleuropneumonia by multilocus sequence analysis of Mycoplasma mycoides subspecies mycoides biotype SC strains.

Contagious bovine pleuropneumonia is a bacterial disease caused by Mycoplasma mycoides subsp. mycoides SC (MmmSC), and included in list A of the Office International des Epizooties. It is one of the major constraints to cattle raising in sub-Saharan and south-western Africa and also a threat to all countries currently free of the disease. MmmSC strains were considered very homogeneous until 1995, when various techniques such as enzymatic restriction of whole DNA or Southern blotting showed that this was not the case. These techniques are unfortunately difficult to standardize and require the extraction of DNA from an MmmSC culture. We therefore decided to investigate the possibility of constructing a molecular epidemiology tool based on multilocus sequence analysis (MLSA) with PCR amplification of various loci followed by sequencing. Six loci were found suitable for this purpose and an additional PCR was designed to detect the presence of an 8.8kb deletion described by others in some strains. Fifteen different MLSA profiles were evidenced in our study. They allowed a clear distinction between European, south-western African and sub-Saharan strains. In addition, the results obtained on strain PO1967 confirmed its European origin, even though it does not exhibit the 8.8kb deletion. This new tool for contagious bovine pleuropneumonia may prove particularly useful for identifying MmmSC strains in countries at risk from contamination. It can also easily be refined by adding more strains or other loci of interest.

Africa↗
PubMed2003

Abattoir-based survey of contagious bovine pleuropneumonia in cattle in Turkey.

Blood samples collected from 945 cattle at four local abattoirs in Turkey were examined for contagious bovine pleuropneumonia (CBPP) by the complement fixation test (CFT) and competitive ELISA (cELISA). In addition, the carcases of the animals were examined macroscopically at the abattoirs and 62 lung samples which had lesions suggestive of CBPP were collected for bacteriological culture. To identify suspicious isolates the PCR was used in addition to the routine biochemical tests. By the CFT, two of the 945 serum samples were seropositive, and by the cELISA, four of them were seropositive. In the bacteriological culture of the lungs, growth was observed in 18 (29 per cent) of the samples by the observation of turbidity in the broths. However, when these broths were inoculated into an agar base, growth was observed in only three (4.8 per cent) samples. These isolates were identified as Mycoplasma species on the basis of biochemical tests. In the PCR analysis of DNA extracted from the broths, none of the isolates was identified as Mycoplasma mycoides subspecies mycoides small colony or one of the members of the M mycoides cluster, but amplification was obtained in only eight (44.4 per cent) of 18 samples, using Mycoplasma-genus specific primers. These DNA samples were examined further with primers specific to 16S rRNA and were then sequenced and compared with the databanks; DNA homologies at different levels were observed in five samples, with Mycoplasma alkalescens, Mycoplasma canadense, Mycoplasma bovis and Mycoplasma bovigenitalium.

Abattoirs↗
PubMed2002

Genetic evolution of Mycoplasma capricolum subsp. capripneumoniae strains and molecular epidemiology of contagious caprine pleuropneumonia by sequencing of locus H2.

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 amplification 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 modifications 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 modifications 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 defined, 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 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 finer identification 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.

Animals↗
PubMed2001

Phage displayed peptides and anti-idiotype antibodies recognised by a monoclonal antibody directed against a diagnostic antigen of Mycoplasma capricolum subsp. capripneumoniae.

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 filamentous phage was screened by panning on the immobilised Mab 4.52 and anti-idiotype antibodies were generated by immunising a chicken with the F(ab')(2) fragments of the antibody. Analysis of the peptide sequences displayed by the isolated phages identified 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 1mg/ml. After absorption, to remove isotypic and allotypic reactivities, the anti-idiotype IgY was specifically 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-specific 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 does not recognise exactly the same antigenic site as antibodies in the goat antisera.

Amino Acid Sequence↗
PubMed2000

Phylogeny of the Mycoplasma mycoides cluster as shown by sequencing of a putative membrane protein gene.

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 flanking 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 define a primer pair that enables the amplification by PCR of two fragments 302 and 1298bp long, respectively. The 302bp long fragment contains an intergenic sequence that can be used for phylogenetic studies or for identification 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 identified as M. capricolum subsp. capricolum, M. capricolum subsp. capricolum and M. sp Bovine Group 7. However M. capricolum subsp. capripneumoniae strains can readily be identified by three specific nucleotide positions or by sequencing the 1298bp 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 distinguished as they bear an insertion sequence 15bp downstream from the stop codon of the membrane protein gene.

Animals↗
PubMed2000

Specific PCR identification of the T1 vaccine strains for contagious bovine pleuropneumonia.

A specific PCR test for the identification of the vaccine strains T1, T1/44 and T1sr, was developed. This PCR reaction is based on variations of DNA sequences in a region flanking one IS1296 copy. The specific primer pair MmmSCP1-T1M2 amplifies a 700-bp long DNA fragment in the T1 vaccine strains and gives no amplification with the 60 other Mycoplasma mycoides subsp. mycoides SC strains tested. This PCR will permit to distinguish the T1 strain from all other vaccine strains and therefore avoid possible confusions. In addition, it should enable better investigations of post-vaccinal reactions.

Animals↗
PubMed2000

Contagious bovine pleuropneumonia. A reassessment of the efficacy of vaccines used in Africa.

Contagious bovine pleuropneumonia is a major threat for cattle in Africa. Since 1956 the T1/44 strain has been used as a vaccine, and later on, T1sr, a streptomycin-resistant variant that gives fewer post-vaccinal reactions had been developed. These vaccines are known not to be very efficient but they normally should provide protection for about eight months. However, recent emergency vaccinations, performed in various countries in the southern part of the continent apparently met with failure, casting doubts on the identity as well as the protection afforded by the T1sr strain. A vaccine trial has been designed to reassess the real protection afforded by these vaccines in face of recently isolated pathogenic strains. Great care has been taken to test the original vaccinal strains at a dose corresponding to the minimum requirement by international standards. The test was performed in Cameroon, Kenya, and Namibia as to take into account the genetic diversity that exists among the pathogenic strains. In those conditions, the protection rate at three months varied from 33 to 67%, whatever the strain used, T1/44 or T1sr. These results call for additional research for vaccine development and careful planning of strategies in the fight against CBPP.

Africa↗
PubMed2000

Comparison between c-ELISA and CFT in detecting antibodies to Mycoplasma mycoides mycoides biotype SC in cattle affected by CBPP in Botswana.

Contagious bovine pleuropneumonia (CBPP), caused by Mycoplasma mycoides mycoides biotype small colony (SC) (MmmSC) appears to be making a serious comeback in Africa after successful control programs in many parts of the continent during the 1960s and 1970s. Botswana, a country that has been free from the disease for more than 50 years, was affected in 1995. An eradication policy was adopted by the Government of Botswana in which 320,000 cattle in the affected district of Ngamiland, Northwestern Botswana were slaughtered. This was followed by a restocking exercise in which 70,000 cattle were sent to the outbreak areas as replacement stock. It became necessary to carry out serosurveillance in order to ensure that the disease did not reenter Botswana and to ensure that the replacement stock remained free from the disease. The specificity and sensitivity of the complement fixation test (CFT) in Botswana was assessed in 82 cattle affected by the disease and held in a double fenced quarantine camp. The newly developed competitive ELISA was made available to the National Veterinary Laboratory (NVL) through the FAO/IAEA Joint Division in Vienna, Austria. Using postmortem lesions as the gold standard and a 2 x 2 contingency table, the two tests were compared in terms of their sensitivity and specificity in detecting antibodies to MmmSC. The CFT was found to be slightly more sensitive than the c-ELISA, and this could be related to the stage of the disease. A long-term study comparing the progression of the disease with the two tests is, therefore, essential.

Animals↗
PubMed1999

Comparative analysis of the lppA locus in Mycoplasma capricolum subsp. capricolum and Mycoplasma capricolum subsp. capripneumoniae.

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 of M. capricolum subsp. capripneumoniae which seems not to express an LppA analogue.

Animals↗
PubMed1999

Genetic and serological analysis of lipoprotein LppA in Mycoplasma mycoides subsp. mycoides LC and Mycoplasma mycoides subsp. capri.

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 members of the M. mycoides cluster. Analysis of the PCR-amplified lppA genes with frequently cutting restriction enzymes showed a certain degree of genetic variability which, however, did not cluster the two subspecies. This PCR therefore allows a rapid identification of M. mycoides subsp. mycoides LC and M. mycoides subsp. capri but does not distinguish between these two closely related subspecies. LppA was expressed in Escherichia coli K-12 and used for the production of polyclonal mouse antiserum. Antibodies against recombinant LppA[MmymyLC] reacted with a 62-kDa protein in all M. mycoides subsp. mycoides LC and M. mycoides subsp. capri type strains and field strains tested but not with the other members of the M. mycoides cluster, thus showing the antigenic specificity of LppA and further supporting the concept that a close relationship exists between these two mycoplasmas.

Bacterial Proteins↗
PubMed1998

Vaccination against contagious bovine pleuropneumonia and the use of molecular tools in epidemiology.

Contagious bovine pleuropneumonia is a serious threat to cattle not only in Africa but also in southern Europe and possibly Asia. It is now present in countries that had been free of the disease for many years, giving rise to doubts about the efficiency of the control strategies. In Africa CBPP is controlled mainly by a vaccination policy that uses variant strains of Mycoplasma mycoides subsp mycoides biotype SC, called T1/44 or T1sr. Until recently, it was not possible to differentiate the various strains within the biotype and consequently to identify the vaccine strains. Restriction analysis of mycoplasma DNA with HindIII and Pst1 has been applied to 24 strains of African origin and one European strain. Each enzyme gave rise to different restriction profiles and the combination of the results permitted subdivision of these strains into 9 groups. Interestingly, some profiles of pathogenic strains seem to be restricted to certain geographical areas. The profile of the poorly immunogenic vaccinal strain KH3J is also very peculiar, and it is easily distinguished from that of the other vaccine strains originating from T1. This technique is simple once the strains are isolated. Efforts are now under way to use molecular tools based on PCR products to alleviate the difficulty of isolation.

Africa↗
PubMed1998

A competitive ELISA for the specific diagnosis of contagious bovine pleuropneumonia (CBPP).

A competitive ELISA, using a specific monoclonal antibody, was designed to detect antibodies to Mycoplasma mycoides subsp. mycoides SC, the agent of contagious bovine pleuropneumonia. One monoclonal antibody was found suitable for such a test, '117/5', it does not cross-react with any of the other mycoplasma species tested, furthermore, its binding is inhibited by positive sera. The cutoff, 50% of inhibition, was determined using a set of negative sera from CBPP-free areas. The sensitivity was controlled with sera from artificially infected animals as well as from sera from areas where CBPP is enzootic. In both cases, cELISA compared favorably with CFT. The precocity of detection was similar but cELISA detected more positives and the positive titers seemed to persist longer than in the case of CFT. Lysis of the antigen used to coat the ELISA plates reduced the variability of fixation and improved the repeatability of the test. A field evaluation is now in progress which will determine the true sensitivity and specificity of the test and also check if antibodies are detected after vaccination.

Animals↗
PubMed1998

Molecular evolution of Mycoplasma capricolum subsp. capripneumoniae strains, based on polymorphisms in the 16S rRNA genes.

Mycoplasma capricolum subsp. capripneumoniae belongs to the so-called Mycoplasma mycoides cluster and is the causal agent of contagious caprine pleuropneumonia (CCPP). All members of the M. mycoides cluster have two rRNA operons. The sequences of the 16S rRNA genes of both rRNA operons from 20 strains of M. capricolum subsp. capripneumoniae of different geographical origins in Africa and Asia were determined. Nucleotide differences which were present in only one of the two operons (polymorphisms) were detected in 24 positions. The polymorphisms were not randomly distributed in the 16S rRNA genes, and some of them were found in regions of low evolutionary variability. Interestingly, 11 polymorphisms were found in all the M. capricolum subsp. capripneumoniae strains, thus defining a putative ancestor. A sequence length difference between the 16S rRNA genes in a poly(A) region and 12 additional polymorphisms were found in only one or some of the strains. A phylogenetic tree was constructed by comparative analysis of the polymorphisms, and this tree revealed two distinct lines of descent. The nucleotide substitution rate of strains within line II was up to 50% higher than within line I. A tree was also constructed from individual operonal 16S rRNA sequences, and the sequences of the two operons were found to form two distinct clades. The topologies of both clades were strikingly similar, which supports the use of 16S rRNA sequence data from homologous operons for phylogenetic studies. The strain-specific polymorphism patterns of the 16S rRNA genes of M. capricolum subsp. capripneumoniae may be used as epidemiological markers for CCPP.

Africa↗
PubMed1997

ISCOM vaccine against contagious bovine pleuropneumonia (CBPP). 1. Biochemical and immunological characterization.

A better vaccine than the existing ones against contagious bovine pleuropneumonia (CBPP) caused by Mycoplasma mycoides subsp. mycoides small colony type (MmmSC) would improve the chances for eradication of CBPP. In such an effort, immunostimulating complexes (ISCOMS) have been prepared from the whole detergent-solubilized cells of MmmSC and characterized biochemically and immunologically. The most efficient detergent for solubilization of the mycoplasma was MEGA-10 which yielded a high recovery of proteins in the ISCOMS. The ISCOMS showed the typical cage-like structure by EM and sedimented as 19S by sucrose gradient centrifugation. The protein pattern of the ISCOMS, analyzed in SDS-PAGE, revealed a great number of bands distributed along the gel as high and low molecular weight polypeptides. The Western blot developed with a serum from a CBPP infected animal detected a reduced number of polypeptides. In samples from whole mycoplasma cells and in ISCOMS, lectin blots revealed more than 20 carbohydrate structures. The ISCOMS induced a strong primary antibody response in mice measured by ELISA and the boost resulted in a 6-fold increase of the serum antibody response. The IgG response was distributed into various IgG subclasses with high IgG1, IgG2a and IgG2b titres while the IgG3 response was low. In cattle the ISCOM vaccine induced strong primary and long lasting secondary antibody responses of similar magnitudes as those of naturally infected animals as recorded by ELISA which persisted more than a year. IgG response was equally distributed in IgG1 and IgG2 subclasses. Also a cell-mediated immune response measured by proliferation assay was induced by low dose of ISCOMS. In the growth inhibition test, sera from vaccinated cattle readily inhibited colony growth already after the first immunization.

Animals↗
PubMed1996

Diagnosis and control of contagious caprine pleuropneumonia.

The diagnosis of contagious caprine pleuropneumonia (CCPP) has often been considered difficult. This is because of the confusion that can arise with other mycoplasmoses of small ruminants. Symptoms and lesions can be similar and the isolation of M. capricolum subsp. capripneumoniae (MccF38) requires skilled technicians. Once MccF38 strains are isolated, their identification should not be difficult. New techniques, such as polymerase chain reaction, now offer the possibility of identifying MccF38 directly from dried samples. However, the isolation of MccF38 strains is always required for an official declaration of infection. Until now, the official serological test has been the complement fixation test; the main drawbacks being lack of sensitivity and specificity and also the short persistence of antibodies detected by this technique. The specific competition enzyme-linked immunosorbent assay has now been developed and should enable wide serological enquiries to determine the real prevalence of the disease. Antibiotic treatments are effective but may not prevent persistence in latent carriers. An inactivated vaccine with saponin as an adjuvant has been produced in Kenya, which protects goats for approximately one year.

Animals↗
PubMed1996

Contagious caprine pleuropneumonia and other pulmonary mycoplasmoses of sheep and goats.

Contagious caprine pleuropneumonia (CCPP) is now a well-defined disease that is caused by Mycoplasma capricolum subsp. capripneumoniae. CCPP is infectious, contagious and fulfils the classic Koch postulates that characterise such types of disease. The distribution of the disease is not exactly known, but reports of mycoplasma isolation and official declarations to the Office International des Epizooties (OIE) enable a probable distribution map to be obtained. There are many other mycoplasmas that can infect goat and sheep lungs and induce pleuropneumonia. However, pleuropneumonia is often restricted to young animals and the prominent symptom is mastitis in lactating does. Other symptoms may also occur, contributing to a syndrome that has been tentatively described in this paper as 'MAKePS syndrome' for mastitis, arthritis, keratitis, pneumonia and septicaemia.

Animals↗