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Richard E.A.,CIRALE ENVA | Richard E.A.,University of Liège | Fortier G.D.,Frank Duncombe Laboratory | Lekeux P.M.,University of Liège | Erck E.V.,University of Liège
Veterinary Journal | Year: 2010

Any disorder impairing a performance horse's ability to ventilate its lungs and exchange oxygen compromises exercise performance in any discipline. Since bronchoalveolar lavage was described in horses in the early 1980s, laboratory evaluation of respiratory fluids, along with clinical and functional assessment of the respiratory system, has become a relevant step in the diagnosis of respiratory disease affecting performance. The aim of this review is to provide objective information to assist clinicians in interpreting laboratory findings by (1) summarising published cytological references values in both clinically healthy horses and those with various airway diseases, (2) assessing the influence of physiological circumstances, such as exercise, on the cytological evaluation, (3) discussing the relationship between cytological and microbiological analyses, clinical signs and respiratory function, and (4) suggesting how this latter relationship may affect performance. © 2009 Elsevier Ltd.


Hans A.,Virology Unit | Gaudaire D.,Virology Unit | Manuguerra J.-C.,Institute Pasteur Paris | Leon A.,Frank Duncombe Laboratory | And 6 more authors.
Journal of Clinical Microbiology | Year: 2015

This study shows that an unbiased amplification method applied to equine arteritis virus RNA significantly improves the sensitivity of the real-time reverse transcription-quantitative PCR (RT-qPCR) recommended by the World Organization for Animal Health. Twelve viral RNAs amplified using this method were hybridized on a high-density resequencing microarray for effective viral characterization. Copyright © 2015, American Society for Microbiology. All Rights Reserved.


PubMed | Institute Pasteur Paris, Virology Unit, University Paris Est Creteil and Frank Duncombe Laboratory
Type: Journal Article | Journal: Journal of clinical microbiology | Year: 2014

This study shows that an unbiased amplification method applied to equine arteritis virus RNA significantly improves the sensitivity of the real-time reverse transcription-quantitative PCR (RT-qPCR) recommended by the World Organization for Animal Health. Twelve viral RNAs amplified using this method were hybridized on a high-density resequencing microarray for effective viral characterization.


Ramery E.,University of Liège | Fraipont A.,University of Liège | Richard E.A.,University of Liège | Richard E.A.,Frank Duncombe Laboratory | And 6 more authors.
Veterinary Clinical Pathology | Year: 2015

Background: Inflammatory airway disease (IAD) affects performance and well-being of horses. Diagnosis is primarily reached by bronchoalveolar lavage (BAL) cytology which is invasive and requires sedation. Objectives: The purpose of this study was to identify differential gene expression in peripheral blood of horses with IAD using species-specific expression microarrays. Methods: Equine gene expression microarrays were used to investigate global mRNA expression in circulating leukocytes from healthy, IAD-affected, and low-performing Standardbred and endurance horses. Results: Nine genes in Standardbred and 61 genes in endurance horses were significantly differentially regulated (P < .001). These genes were related to inflammation (eg, ALOX15B, PLA2G12B, and PENK), oxidant/antioxidant balance (eg, DUOXA2 and GSTO1-1), and stress (eg, V1aR, GRLF1, Homer-2, and MAOB). All these genes were up-regulated, except down-regulated Homer-2 and MAOB. DUOXA2, ALOX15B, PLA2G12B, MAOB, and GRLF1 expression was further validated by RT-qPCR. An increase in glutathione peroxidase (GPx) activity in heparinized whole blood of IAD-affected Standardbred (P = .0025) and endurance horses (P = .0028) also suggests a deregulation of the oxidant/antioxidant balance. There was good correlation (r = .7354) between BAL neutrophil percentage and whole blood GPx activity in all horses. Conclusions: This study showed that circulating blood cell gene expression reflects inflammatory responses in tissues. Whether any of the genes have potential for diagnostic applications in the future remains to be investigated. Although not specific for IAD, whole blood GPx activity appears to be correlated with BAL neutrophil percentage. This finding should be further assessed by testing a larger number of horses. © 2014 American Society for Veterinary Clinical Pathology.


Depecker M.,National School of Engineering in Agricultural and Food Industries | Richard E.A.,Frank Duncombe Laboratory | Pitel P.-H.,Frank Duncombe Laboratory | Fortier G.,Frank Duncombe Laboratory | And 3 more authors.
Veterinary Journal | Year: 2014

The aim of this study was to determine whether the lung side being sampled would significantly influence bronchoalveolar lavage (BAL) cytological profiles and subsequent diagnosis in Standardbred racehorses. One hundred and thirty-eight French Trotters in active training and racing were included in a prospective observational study. BAL was performed using videoendoscopy in both right and left lungs during summer meetings in 2011 (64 horses) and 2012 (74 horses). Cytological data performed 24. h later from right and left lungs were compared and specifically used to classify horses as affected with exercise-induced pulmonary haemorrhage (EIPH), inflammatory airway disease (IAD), or were 'controls'. For IAD, cytological definition was based on two different cut off values.Neutrophil percentages, haemosiderophage percentages and the haemosiderophage/macrophage (H/M) ratios were significantly higher in the right compared to the left lung. Measures of intra-class correlation coefficients revealed a fair agreement between left and right lungs for percentages of mast cells, eosinophils, and for the H/M ratio, and a moderate agreement for neutrophil percentages. Fair to moderate agreements were observed between left and right lungs for the diagnosis of IAD and/or EIPH based on kappa coefficients. When sampling one lung only, the risk of incorrectly classifying a horse as a 'control' increased with the use of the restraint cut-off values for IAD. As BAL from one lung is not representative of the other lung in the same horse, both lungs should be sampled for a better assessment of lung cellularity and for a precise diagnosis of lower airway diseases. © 2013 Elsevier Ltd.


PubMed | EQUI TEST, University of Caen Lower Normandy, National School of Engineering in Agricultural and Food Industries and Frank Duncombe Laboratory
Type: Journal Article | Journal: Veterinary journal (London, England : 1997) | Year: 2014

The aim of this study was to determine whether the lung side being sampled would significantly influence bronchoalveolar lavage (BAL) cytological profiles and subsequent diagnosis in Standardbred racehorses. One hundred and thirty-eight French Trotters in active training and racing were included in a prospective observational study. BAL was performed using videoendoscopy in both right and left lungs during summer meetings in 2011 (64 horses) and 2012 (74 horses). Cytological data performed 24h later from right and left lungs were compared and specifically used to classify horses as affected with exercise-induced pulmonary haemorrhage (EIPH), inflammatory airway disease (IAD), or were controls. For IAD, cytological definition was based on two different cut off values. Neutrophil percentages, haemosiderophage percentages and the haemosiderophage/macrophage (H/M) ratios were significantly higher in the right compared to the left lung. Measures of intra-class correlation coefficients revealed a fair agreement between left and right lungs for percentages of mast cells, eosinophils, and for the H/M ratio, and a moderate agreement for neutrophil percentages. Fair to moderate agreements were observed between left and right lungs for the diagnosis of IAD and/or EIPH based on kappa coefficients. When sampling one lung only, the risk of incorrectly classifying a horse as a control increased with the use of the restraint cut-off values for IAD. As BAL from one lung is not representative of the other lung in the same horse, both lungs should be sampled for a better assessment of lung cellularity and for a precise diagnosis of lower airway diseases.


Bidaud P.,Dozule Laboratory for Equine Diseases | Hebert L.,Dozule Laboratory for Equine Diseases | Barbey C.,Dozule Laboratory for Equine Diseases | Barbey C.,CNRS Laboratory of Microbiology Signals and Microenvironment | And 6 more authors.
PLoS ONE | Year: 2012

Rhodococcus equi is one of the most widespread causes of disease in foals aged from 1 to 6 months. R. equi possesses antioxidant defense mechanisms to protect it from reactive oxygen metabolites such as hydrogen peroxide (H2O2) generated during the respiratory burst of phagocytic cells. These defense mechanisms include enzymes such as catalase, which detoxify hydrogen peroxide. Recently, an analysis of the R. equi 103 genome sequence revealed the presence of four potential catalase genes. We first constructed ΔkatA-, ΔkatB-, ΔkatC-and ΔkatD-deficient mutants to study the ability of R. equi to survive exposure to H2O2 in vitro and within mouse peritoneal macrophages. Results showed that ΔkatA and, to a lesser extent ΔkatC, were affected by 80 mM H2O2. Moreover, katA deletion seems to significantly affect the ability of R. equi to survive within murine macrophages. We finally investigated the expression of the four catalases in response to H2O2 assays with a real time PCR technique. Results showed that katA is overexpressed 367.9 times (±122.6) in response to exposure to 50 mM of H2O2 added in the stationary phase, and 3.11 times (±0.59) when treatment was administered in the exponential phase. In untreated bacteria, katB, katC and katD were overexpressed from 4.3 to 17.5 times in the stationary compared to the exponential phase. Taken together, our results show that KatA is the major catalase involved in the extreme H2O2 resistance capability of R. equi. © 2012 Bidaud et al.


Rahelinirina S.,Pasteur Institute | Leon A.,Frank Duncombe Laboratory | Harstskeerl R.A.,OIE and National Leptospirosis Reference Center | Sertour N.,Institute Pasteur Paris | And 9 more authors.
PLoS ONE | Year: 2010

Background: Leptospirosis has long been a major public health concern in the southwestern Indian Ocean. However, in Madagascar, only a few, old studies have provided indirect serological evidence of the disease in humans or animals. Methodology/Principal Findings: We conducted a large animal study focusing on small-mammal populations. Five field trapping surveys were carried out at five sites, from April 2008 to August 2009. Captures consisted of Rattus norvegicus (35.8%), R. rattus (35.1%), Mus musculus (20.5%) and Suncus murinus (8.6%). We used microbiological culture, serodiagnosis tests (MAT) and real-time PCR to assess Leptospira infection. Leptospira carriage was detected by PCR in 91 (33.9%) of the 268 small mammals, by MAT in 17 of the 151 (11.3%) animals for which serum samples were available and by culture in 9 of the 268 animals (3.3%). Rates of infection based on positive PCR results were significantly higher in Moramanga (54%), Toliara (48%) and Mahajanga (47.4%) than in Antsiranana (8.5%) and Toamasina (14%) (p = 0.001). The prevalence of Leptospira carriage was significantly higher in R. norvegicus (48.9%), S. murinus (43.5%) and R. rattus (30.8%) than in M. musculus (9.1%) (p<0.001). The MAT detected antibodies against the serogroups Canicola and Icterohaemorrhagiae. Isolates were characterized by serology, secY sequence-based phylogeny, partial sequencing of rrs, multi-locus VNTR analysis and pulsed field gel electrophoresis. The 10 isolates obtained from nine rats were all identified as species L. interrogans serogroup Canicola serovar Kuwait and all had identical partial rrs and secY sequences. Conclusions/Significance: We present here the first direct evidence of widespread leptospiral carriage in small mammals in Madagascar. Our results strongly suggest a high level of environmental contamination, consistent with probable transmission of the infection to humans. This first isolation of pathogenic Leptospira strains in this country may significantly improve the detection of specific antibodies in human cases. © 2010 Rahelinirina et al.


Fortier G.,Frank Duncombe Laboratory | Fortier G.,University of Liège | van Erck E.,University of Liège | Pronost S.,Frank Duncombe Laboratory | And 2 more authors.
Veterinary Journal | Year: 2010

Equine gammaherpesviruses (γEHV) have been widely studied over the past 45 years and many isolates have been characterised. Despite this, the diagnosis of γEHV infection remains difficult to establish as its clinical manifestations lack specificity, ranging from mild respiratory signs in a small number of animals to outbreaks in large groups of young horses. This review focuses on the epidemiology, pathogenesis, clinical manifestations and diagnosis of equine herpesvirus (EHV)-2 and -5 infections, as well as on the genetic variation of these viruses. Study of these variations has resulted in hypotheses relating to viral re-infection and re-activation. Interestingly, the viruses were found to contain genetic sequences identical to those of eukaryotic cells which are considered central to the development of viral latency through interfering with host immune and inflammatory responses. Future molecular biological studies will further elucidate the virulence mechanisms of these equine pathogens. © 2009 Elsevier Ltd.


Leon A.,Frank Duncombe Laboratory | Pronost S.,Frank Duncombe Laboratory | Fortier G.,Frank Duncombe Laboratory | Andre-Fontaine G.,Leptospira Medical and Molecular Bacteriology Unit | Leclercq R.,University of Caen Lower Normandy
Journal of Clinical Microbiology | Year: 2010

Fifty-three strains belonging to the pathogenic species Leptospira interrogans and Leptospira kirschneri were analyzed by multilocus sequence analysis. The species formed two distinct branches. In the L. interrogans branch, the phylogenetic tree clustered the strains into three subgroups. Genogroups and serogroups were superimposed but not strictly. Copyright © 2010, American Society for Microbiology. All Rights Reserved.

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