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Biomedical subjects

I Kempf

Publications and source records attributed to I Kempf.

At least 19 recordsLinked to original sources
PubMed2004

Evidence for natural horizontal transfer of tetO gene between Campylobacter jejuni strains in chickens.

AIMS: The transfer of tetO gene conferring resistance to tetracycline was studied between Campylobacter jejuni strains, in the digestive tract of chickens. METHODS AND RESULTS: In vitro conjugation experiments were first performed in order to select donor/recipient couples for further in vivo assay. Then, chickens were inoculated with a donor/recipient couple of C. jejuni strains displaying spontaneous in vitro tetracycline resistance gene transfer. The donor was a tetracycline-resistant ampicillin-susceptible strain, and the recipient was a tetracycline-susceptible ampicillin-resistant strain. Chicken droppings were streaked on antimicrobial selective media and bi-resistant Campylobacter isolates were further characterized according to their donor or recipient flaA gene RFLP profile. The acquisition of tetracycline-resistance gene by the recipient C. jejuni strain from the donor C. jejuni strain was confirmed by tetO PCR. CONCLUSIONS: The study showed that transfer of tetO gene occurs rapidly and without antimicrobial selection pressure between C. jejuni strains in the digestive tract of chickens. SIGNIFICANCE AND IMPACT OF THE STUDY: The rapid and spontaneous transfer of tetO gene may explain the high prevalence of tetracycline resistance in chicken Campylobacter strains.

Animals↗
PubMed2002

A reverse transcription-PCR assay to detect viable Mycoplasma synoviae in poultry environmental samples.

In order to study horizontal transmission of Mycoplasma synoviae an avian pathogen, a reverse transcription-polymerase chain reaction (RT-PCR) assay was developed to detect viable Mycoplasma in environment. The test was based on the RT-PCR of the 16S ribosomal RNA (rRNA) of Mycoplasma genus. Results showed that Mycoplasma 16S rRNA was stable up to 23 h after cell death. Therefore, the test allowed detection of viable or very recently (less than 23 h) dead mycoplasmas. M. synoviae survival in artificially contaminated water, food and soil and in the environment of M. synoviae experimentally infected turkeys was estimated by culture and RT-PCR. The RT-PCR method was then applied in a naturally infected laying hen farm showing problems of recurrent mycoplasmosis in the hens. Results confirmed the usefulness of RT-PCR in checking the efficiency of biosecurity measures and in improving cleaning and disinfection protocols.

Air Microbiology↗
PubMed2002

In vitro development of resistance to enrofloxacin, erythromycin, tylosin, tiamulin and oxytetracycline in Mycoplasma gallisepticum, Mycoplasma iowae and Mycoplasma synoviae.

The in vitro emergence of resistance to enrofloxacin, erythromycin, tylosin, tiamulin, and oxytetracycline in three avian Mycoplasma species, Mycoplasma gallisepticum, Mycoplasma synoviae and Mycoplasma iowae was studied. Mutants were selected stepwise and their MICs were determined after 10 passages in subinhibitory concentrations of antibiotic. High-level resistance to erythromycin and tylosin developed within 2-6 passages in the three Mycoplasma species. Resistance to enrofloxacin developed more gradually. No resistance to tiamulin or oxytetracycline could be evidenced in M. gallisepticum or M. synoviae after 10 passages whereas, resistant mutants were obtained with M. iowae. Cross-sensitivity tests performed on mutants demonstrated that mycoplasmas made resistant to tylosin were also resistant to erythromycin, whereas mutants made resistant to erythromycin were not always resistant to tylosin. Some M. iowae tiamulin-resistant mutants were also resistant to both macrolide antibiotics. Enrofloxacin and oxytetracycline did not induce any cross-resistance to the other antibiotics tested. These results show that Mycoplasma resistance to macrolides can be quickly selected in vitro, and thus, providing that similar results could be obtained under field conditions, that development of resistance to these antibiotics in vivo might also be a relatively frequent event.

Animals↗
PubMed2002

Antimicrobial susceptibility of Streptococcus suis isolated from swine in France and from humans in different countries between 1996 and 2000.

The susceptibility of 135 Streptococcus suis strains isolated from pigs (n = 110) and from humans (n = 25) to 13 antimicrobial agents was studied by microdilution and disc diffusion methods using Mueller-Hinton Agar II (MH) supplemented with either defibrinated sheep blood (MHSB) or horse serum (MHHS). Results were similar for both methods used except for penicillin G whose zone diameters were reduced with MHSB compared with MHHS. When MH was supplemented with sheep blood, 39% of S. suis strains classified as penicillin susceptible by MHHS microdilution showed intermediate susceptibility. Nearly all strains were susceptible to penicillin G (except by disc diffusion in MHSB), amoxicillin, ceftiofur, florfenicol, gentamicin and bacitracin. The least active antimicrobial agents were doxycycline and macrolides/lincosamides. High-level resistance (MIC > 500 mg/L or zone diameters < 10 mm) to streptomycin and kanamycin was detected in only a few strains. The virulence of strains did not seem to be related to antimicrobial resistance because no statistical difference was reported between the proportion of resistant strains of S. suis isolated from pigs with meningitis, septicaemia and arthritis, and those from tonsils and nasal cavities. However, significant differences were found in the proportions of macrolide- or doxycycline-resistant strains between S. suis serotype 2 and other serotypes. The results of antibiotic susceptibility testing presented in this study indicate that beta-lactams can be used in empirical treatment of human and pig S. suis infections in France.

Animals↗
PubMed2002

Fluoroquinolone resistance in Mycoplasma gallisepticum: DNA gyrase as primary target of enrofloxacin and impact of mutations in topoisomerases on resistance level.

Resistant mutants of Mycoplasma gallisepticum were selected in vitro by passaging strains 10 times in increasing concentrations of enrofloxacin. The regions of gyrA/gyrB and parC/parE, encoding the quinolone resistance-determining regions (QRDRs) of DNA gyrase and DNA topoisomerase IV, respectively, of the mutants obtained during different passages were sequenced. Several mutations were found in the four fluoroquinolone targets. Substitution of Ser-83-->Arg in GyrA and Ser-80-->Leu or Trp in ParC QRDRs seem to have the greatest impact on resistance to fluoroquinolones. The results obtained also suggest that the preferential target of enrofloxacin in M. gallisepticum is DNA gyrase.

Anti-Infective Agents↗
PubMed2002

Characterization of mutations in DNA gyrase and topoisomerase IV Involved in quinolone resistance of Mycoplasma gallisepticum mutants obtained in vitro.

Mycoplasma gallisepticum enrofloxacin-resistant mutants were generated by stepwise selection in increasing concentrations of enrofloxacin. Alterations were found in the quinolone resistance-determining regions of the four target genes encoding DNA gyrase and topoisomerase IV from these mutants. This is the first description of such mutations in an animal mycoplasma species.

4-Quinolones↗
PubMed2001

Comparison of pulsed-field gel electrophoresis with random amplified polymorphic DNA for typing of Mycoplasma synoviae.

Pulsed-field gel electrophoresis (PFGE) analysis was developed and compared with random amplified polymorphic DNA (RAPD) method to type 18 Mycoplasma synoviae (MS) strains. All analysed strains were typeable by RAPD but only 89% of MS strains were typeable by PFGE because of DNA degradation. The discriminatory power of RAPD was greater than that of PFGE but the two techniques had a discriminatory index superior to 0.95, the threshold value for interpreting typing results with confidence. The in vitro, in ovo and in vivo reproducibility of both typing techniques was 100%. However, the interpretation of RAPD patterns was complicated because of inconsistent band intensity. Thus, these molecular typing techniques should be helpful for epidemiological studies of avian mycoplasma infections.

Animals↗
PubMed2001

Development of a blocking enzyme-linked immunosorbent assay for detection of turkey antibodies to Mycoplasma meleagridis.

A blocking enzyme-linked immunosorbent assay (B-ELISA) was developed to detect antibodies to Mycoplasma meleagridis (MM) in turkey sera. This assay was based on two mouse monoclonal antibodies recognising all MM strains tested but none of seven avian mycoplasmal species tested. Furthermore, their binding to the Tween 20 antigen was inhibited by serum from MM-infected birds. The B-ELISA test format was optimized. The cut-off was determined using a set of sera from MM-free turkeys. This B-ELISA was then compared with a commercial indirect ELISA (I-ELISA). Specificities of the two ELISA tests were not significantly different (100 or 99%, respectively). The sensitivity of B-ELISA was significantly higher than the I-ELISA when I-ELISA suspicious results were considered as negative. Testing sera from experimentally MM-infected animals showed that serum plate agglutination (SPA) test detected positive birds before both ELISA methods. Samples were collected in MM-infected commercial flocks and analyzed by SPA, ELISAs, MM-PCR or culture. Results showed that the sensitivity of the B-ELISA appeared superior to the I-ELISA. Moreover, the ability to detect maternal antibodies makes it a useful tool for eradication or control of MM infections.

Animals↗
PubMed2001

Antigen heterogeneity and epitope variable expression in Mycoplasma meleagridis isolates.

Immunoblotting was used to check the antigenic profiling of 27 Mycoplasma meleagridis strains isolated in different countries. Hyperimmune polyclonal rabbit antiserum as well as monoclonal antibodies (MAbs) raised against M. meleagridis (MM) showed antigen heterogeneity among strains. Five anti-MM MAbs were selected for lack of reaction against heterologous avian mycoplasma. Three of these five Mabs did not cross-react with 63 mycoplasma strains from six species affecting turkeys other than M. meleagridis. The five Mabs used to analyse the epitopes of 30 M. meleagridis strains indicated that some epitopes were not expressed in all strains. Moreover, other epitopes were located on proteins which differed according to number or molecular mass from strain to strain. The five Mabs therefore, recognised variable surface proteins, among which two were amphiphilic membrane proteins. Three of the selected Mabs recognised 29 or 30 of the 30 tested strains. The in vitro expression of surface epitopes in M. meleagridis ATCC 25284 was investigated by colony immunobinding and allowed demonstration of a variable antigenic system.

Animals↗
PubMed2001

Molecular differentiation of Mycoplasma gallisepticum and Mycoplasma imitans strains by pulsed-field gel electrophoresis and random amplified polymorphic DNA.

Pulsed-field gel electrophoresis (PFGE) and random amplified polymorphic DNA (RAPD) analysis were used to compare 21 Mycoplasma gallisepticum strains and five M. imitans strains. Each strain of M. gallisepticum typed by PFGE and RAPD methods was genetically quite unique and RAPD and PFGE fingerprinting enabled strain characterization. Relationships between the M. gallisepticum and M. imitans strains were established and dendrograms were drawn from PFGE and RAPD patterns. PFGE group A and RAPD group D were significantly associated with M. imitans strains (P < 0.05). Three M. imitans strains shared the same PFGE and RAPD patterns. The two M. gallisepticum vaccine strains had singular PFGE and RAPD patterns. Thus, PFGE and RAPD can be used to investigate disease outbreaks in vaccinated flocks or for epidemiological tracking. For M. gallisepticum, the RAPD and PFGE discriminatory powers were superior to 0.95 and the in vitro, in ovo and in vivo reproducibility of RAPD and PFGE was 100%. The RAPD drawback was the inconsistent band intensity complicating the interpretation of patterns, while the PFGE limit was its low typeability (86%). Thus, these two molecular typing methods seemed complementary for M. gallisepticum epidemiological studies.

Animals↗
PubMed2000

Detection of Mycoplasma synoviae in poultry environment samples by culture and polymerase chain reaction.

Successful detection of Mycoplasma synoviae (MS) by culture and PCR from samples collected in the environment of experimentally infected chickens and turkeys, or under field conditions, is described. Results showed that in the experimental infection, 10/96 and 46/96 samples of food, drinking water, feathers, droppings or dust were positive by culture and Mycoplasma-PCR. In field conditions, the number of positive results for environmental samples were respectively 7/28 and 17/28. These observations highlight the high disseminating capacities of this mycoplasma and show the usefulness of the PCR method for epidemiological studies.

Animals↗
PubMed1999

[C-reactive protein and transthyretin in early diagnosis of infection after open fractures of the lower limbs (a preliminary study)].

PURPOSE OF THE STUDY: The authors investigated the value of C-reactive protein (CRP) and transthyretin (TTR) in the early diagnosis of infection after open fractures of the lower limb in an open, prospective study. MATERIAL AND METHODS: Eighty patients were treated with acute debridement and bone fixation. Follow-up included clinical, radiological, bacteriological and biological assessment: white cell blood count (WBC), erythrocyte sedimentation rate (ESR), CRP and TTR. Diagnosis of infection was based on macroscopic presence of pus. RESULTS: Post-operative reference biological profiles were defined in 74 cases without infection. Reference profiles of WBC and ESR showed unreliable interindividual variations and could not be considered for the diagnosis of infection. Reference profiles of CRP and TTR showed a respective increase (for CRP) and decrease (for TTR) in the early post-operative course, with return to normal values after 12 days. In 6 infected patients, CRP concentrations were suddenly increased and TTR concentrations decreased at the time (3 cases) or even before (3 cases) clinical diagnosis of infection. These variations were mostly simultaneous. No unusual profile was found. The ratio of CRP/TTR concentrations experienced also a sudden increase in infected cases. DISCUSSION: Because of not specifical and unreliable variations in the post-operative outcome of non infected patients, WBC and ESR cannot be considered for the early diagnosis of infection. CRP and TTR concentrations with a respective cut-off value of 100 mg/L and 120 mg/L were found efficient for the early diagnosis of infection, and preceded clinical diagnosis in three of them. A CRP/TTR ratio over 60 p. 100, 8 days or more after initial surgery was found to be very specific (93 p. 100) and sensitive (100 p. 100) for the diagnosis of infection. CONCLUSION: Serial quantifications of CRP and TTR should be performed every four days during the follow-up of open fractures in order to early diagnose a post-operative infection. Comparison of both CRP and TTR could allow a higher accuracy, because of the possible lack of variation of one the two markers.

Adolescent↗
PubMed1998

Utility of an internal control for evaluation of a Mycoplasma meleagridis PCR test.

Mycoplasma meleagridis, a turkey pathogen, can be detected by PCR directly from tracheal or genital swabs. However, up to 40% samples may contain inhibitory substances. A DNA fragment, that can be amplified with M. meleagridis primers and in the same cycling conditions, was constructed to use as an internal control (IC) to check for these inhibitors. This IC can easily be distinguished from the M. meleagridis amplicon after agarose gel electrophoresis since it is longer. Use of this IC in PCR amplifications revealed that more than 35% of turkey tracheal swabs and more than 45% of turkey cloacal swabs contained inhibitors. In most cases, dilution (1:100) of swab lysates allowed amplification of the internal control but DNA purification may be necessary to eliminate inhibitors (20% of tracheal swabs and 5% of cloacal swabs). Use of this internal control DNA allowed assessment of the efficiency of each individual reaction and ensured that the reaction was not inhibited by interfering substances.

Animals↗
PubMed1998

Comparison of serological tests for detection of Mycoplasma gallisepticum antibodies in eggs and chicks hatched from experimentally infected hens.

Specific pathogen free hens and males were experimentally infected with Mycoplasma gallisepticum. Eggs were then collected, and a part was incubated and set for hatching. Mycoplasma cultures were performed on infected adults and antibodies to MG were analysed by use of slide agglutination (SA) test and commercial ELISA tests on adults and chicks sera and on yolks from non incubated eggs. Both ELISA tests could detect antibodies in yolks from non incubated eggs laid three weeks after infection. SA and the three ELISA tests revealed positive sera in chicks hatched from eggs laid as soon as one week after infection.

Animals↗
PubMed1998

Efficacy of difloxacin in growing broiler chickens for the control of infection due to pathogenic Mycoplasma gallisepticum.

Chickens 14 days old were experimentally inoculated with Mycoplasma gallisepticum (MG) R-P10 strain. After development of respiratory symptoms, birds were left unmedicated or medicated for 5 consecutive days with Difloxacin 5, 7.5 or 10 mg/kg body weight per day or Enrofloxacin at the dose level of 10 mg/kg body weight per day. Evaluation of efficacy was based on body weight, symptoms, post-mortem findings, re-isolation of MG and serology. Results indicated that under the conditions of this experiment, treatment with 7.5 mg Difloxacin per kg body weight for 5 days was already effective against pathogenic MG infection. The dose of 10 mg/kg Difloxacin was equally effective as a dose of 10 mg/kg Enrofloxacin in treating respiratory symptoms.

Animals↗