Evaluation des effets de l’extrait aqueux des fleurs de Nymphaea lotus Linn. (Nymphéacées) sur la fonction de reproduction des rats normoglycémique et diabétique de type 1

In traditional medicine of Cameroon, Nymphaea lotus Linn. (Nympheaceae) is used for treatment of male sexual disorders. The aim of this study was to evaluate the effects of the N. lotus flowers aqueous extracts on general mating behavior, fertility and some androgenic parameters of normoglycemic and diabetic adult male rats. Mating behavior assessment was carried out with primiparous and with oestrus female rats. Male rats were divided into 5 groups and subjected once in a day to the following treatment: distilled water (10 mL/kg), Sildenafil citrate (5mg/kg), Testosterone enanthate (5mg/kg) and plant extract (75 mg/kg and 150 mg/kg). Treatment lasted for 8 and 55 days before sacrifice. Organ weight, epididymal sperm counts, motility, viability, histomorphometric analysis, muscular strength, seminal fructose, cholesterol level, epididymal and serum proteins testicular were determined. Results showed a dose- dependant influence of the treatment in sperm count and motility which significantly increases compared to the distilled water-administered control. An increase in some sexual performance parameters (mount and intromission frequencies) was observed in both diabetic and normal rats compared to respective controls. The latencies of mount and intromission were significantly reduced. Computed frequencies of penile licking, index of libido and sexual motivation were higher in normal and diabetic extract-treated animals; which suggest an enhancement of motivational parameters. The treatment also caused significant increases in the weight of the testis, prostate, epididymis, in serum cholesterol and epididymal protein level in normal rats compared to the distilled water-administered control. These results indicate an androgenic pro-sexual potential of N. lotus in male rats and justify the empirical use of N. lotus in the management of erectile dysfunction and fertility disorders in males. Key-words: Nymphaea lotus, prosexual, androgenic, fertility, diabetic, male rat. //Selon les tradipraticiens de l’Ouest-Cameroun, Nymphaea lotus Linn. (N. lotus), de la famille des Nymphéacées, est utilisé pour le traitement des dysfonctionnements sexuels chez les hommes. Dans cette étude, l’effet de l’extrait aqueux des fleurs de N. lotus sur la fonction de reproduction des rats mâles adultes normaux et diabétiques a été évalué. Dans le but d’évaluer les effets de l’extrait aqueux des fleurs de N. lotus sur les paramètres généraux de copulation, d’androgénicité et de fertilité, les rats normoglycémiques et diabétiques ont été divisé en 3 groupes contrôle [Groupe I- recevant de l’eau distillée, Groupe II et III recevant respectivement du citrate de sildénafil (5 mg/kg) et l’énanthate de testostérone (5 mg/kg)] et deux groupes expérimentaux traités à l’extrait aux doses 75 mg/kg (Groupe IV) et 150 mg/kg (Groupe V). L’eau distillée, l’extrait et la substance de référence était administré per os une fois par jour. Pour analyser le comportement sexuel et la fertilité, des femelles primipares et en oestrus étaient utilisées. Le traitement a duré 8 et 55 jours avant le sacrifice. Le poids relatif des organes, le nombre de spermatozoïde, la motilité, les analyses morphométriques, la force musculaire, le taux de cholestérol, le taux de protéines sériques et épididymaires étaient déterminé. Le temps de latence de monte et d’intromission a diminué significativement alors que la fréquence d’éjaculation a augmenté. L’index de libido, la fréquence de monte, d’intromission, d’éjaculation, d’orientation des mâles vis-à-vis d’eux même et l’index de motivation sexuelle a augmenté chez les animaux traités l’extrait comparés à ceux ayant reçu de l’eau distillée aussi bien chez les normaux que chez le diabétiques qui n’enregistrent d’ailleurs aucune éjaculation. Le traitement a augmenté significativement (P < 0,01) le poids des testicules, de la prostate et de l’épididyme. Il est observé une augmentation dose-dépendante du nombre et de la motilité des spermatozoïdes (P < 0,05), ainsi qu’une augmentation significative (P < 0,001) temps-dépendant du taux de cholestérol sérique et de protéines épididymaires. Ces résultats indiquent un potentiel androgénique pro-érectile de N. lotus chez les rats mâles et justifient l’utilisation empirique des fleurs de N. lotus dans le traitement des dysfonctions érectiles et des problèmes de fertilité chez les hommes. Mots-clés: Nymphaea lotus, pro-érectile, androgénique, fertilité, diabétiques, rat male

Presence of new mecA and mph(C) variants conferring antibiotic resistance in Staphylococcus spp. isolated from the skin of horses before and after clinic admission

Because of the frequency of multiple antibiotic resistance, Staphylococcus species often represent a challenge in incisional infections of horses undergoing colic surgery. To investigate the evolution of antibiotic resistance patterns before and after preventative peri- and postoperative penicillin treatment, staphylococci were isolated from skin and wound samples at different times during hospitalization. Most staphylococci were normal skin commensals and belonged to the common coagulase-negative group. In some cases they turned out to be opportunistic pathogens present in wound infections. MICs were determined for 12 antibiotics, and antibiotic resistance genes were detected by microarray. At hospital admission, horses harbored staphylococci that were susceptible to antibiotics or resistant to one group of drugs, mainly due to the presence of new variants of the methicillin and macrolide resistance genes mecA and mph(C), respectively. After 3 days, the percentage of Staphylococcus isolates displaying antibiotic resistance, as well as the number of resistance genes per isolate, increased moderately in hospitalized horses without surgery or penicillin treatment but dramatically in hospitalized horses after colic surgery as well as penicillin treatment. Staphylococcus species displaying multiple resistance were found to harbor mainly genes conferring resistance to beta-lactams (mecA and blaZ), aminoglycosides [str and aac(6')-Ie-aph(2')-Ia], and trimethoprim [dfr(A) and dfr(D)]. Additional genes conferring resistance to macrolides [mph(C), erm(C), and erm(B)], tetracycline [tet(K) and tet(M)], chloramphenicol [cat(pC221) and cat(pC223)], and streptothricin (sat4) appeared in several strains. Hospitalization and preventive penicillin use were shown to act as selection agents for multidrug-resistant commensal staphylococcal flora.

Management, feeding, production, reproduction and udder health on organic and conventional Swiss dairy farms

Organic dairy farms (OP; n=60) and conventional dairy farms (integrated production, IP; n=60), matched in size, location, and agricultural zone (altitude), were studied for possible differences in management, feeding, production, reproduction and udder health. OP and IP farms were similar in size (17.7 and 16.9 ha), milk quota (65900 and 70,000 kg/year), cow number (14 and 15), cow age (5.3 and 5.2 years), housing of cows of the Simmental x Red Holstein or Holstein breeds (87 and 75%; 45 and 60%), but differed significantly with respect to loose housing systems (18 and 7%), outside paddocks (98 and 75%), energy-corrected 305-d milk yield (5,695 and 6,059 kg), milk protein content (31.8 and 32.7 g/kg), use of bucket milking systems (73 and 33%), observance of regular (12-h) milking intervals (47 and 68%), routine application of the California-Mastitis-Test (10 and 28%), teat dipping after milking (25 and 43%) and blanket dry cow treatments (0 and 45%). Milk somatic cell counts on OP and IP farms (119 000 and 117,000/mL) and reproduction data were similar and there were no significant differences between OP and IP farms as concerns available feeds, planning and management of feeding. Alternative veterinary treatments were used more often on OP than IP farms (55 and 17%). Main causes for cow replacements on OP and IP farms were fertility disorders (both 45%), age (40 and 42%), sale (30 and 37%) and udder health (35 and 13%).Between OP and IP Swiss dairy farms thus relatively few larger differences were found.

[Evaluation of the quality of husbandry of Franches-Montagnes horses in their breeding farms]

The quality of husbandry of Franches-Montagnes horses (FM) in Switzerland is evaluated on the basis of an investigation carried out in 2002 by the Swiss FM breeding federation. Questionnaires were sent to 3500 of its members and the results include data from 968 breeding enterprises, housing a total of 3965 FM: 46.1% were breeding mares (61.0% with foal at foot), 26.5% young stock, 1.3% stallions and 26.0% non breeding stock (74.6% of which were pleasure horses and 25.4% working horses). 57.6% of the FM were housed in individual boxes with or without permanent outdoor access, 25.4% were hold in groups with or without permanent outdoor access, the remaining 17.0% were kept in standing stalls. 95.0% of the FM had at least visual contact with other equines and 99.2% had sufficient light in their stable. 88.1% were stabled on long stalk straw, while only 4.3% were bedded on other materials other than straw. The average time spent at pasture per horse and per week ranged from 96.5 +/- 51.6 hours in summer to 27.2 +/- 26.7 hours in winter. On average, a FM is used for 8.3 +/- 6.5 hours per week. Horses with an paddock at their disposal spend an average of 39.8 +/- 45.9 hours there per week.

[The influence of the socio-economic structure of the breeding farms of Franches-Montagnes horses on the conditions of husbandry]

The socio-economic structure of the breeding farms of Franches-Montagnes horses (FM) in Switzerland is evaluated on the basis of an investigation carried out in 2002 by the Swiss FM breeding federation. Questionnaires were sent to 3500 of its members and the results include data from 968 breeding enterprises, housing a total of 3965 FM. The quality of the husbandry of FM varies according to factors such as the altitude and the geographical situation of the farms and studs. Socio-economic parameters, such as the role of FM in the business, their use (breeding, driving, riding) and the age and level of professional education of the owners may also have an effect on standards of husbandry. The results show that the owners for whom FM represent a source of income more frequently keep their horses in standing stalls, but give them more time to exercise at liberty than the horses belonging to amateur breeders. Younger and better educated breeders are more likely to house their animals in groups.

Evaluation of an in vitro sulphidoleukotriene release test for diagnosis of insect bite hypersensitivity in horses

REASONS FOR PERFORMING STUDY: Insect bite hypersensitivity (IBH) is an IgE-mediated allergic dermatitis caused by bites of Culicoides and Simulium species, and improved means of diagnosis are required. OBJECTIVES: The cellular antigen simulation test (CAST) with C. nubeculosus and S. vittatum extracts was assessed in a population of IBH-affected and healthy horses. Variations in test results over a one year period and possible cross-reactivity between different insect extracts was studied. METHODS: A total of 314 mature horses were studied using the CAST. Influence of severity of clinical signs, gender and age were evaluated, and 32 horses were tested repeatedly over one year. The kappa reliability test was used to assess agreement of the test results with different insect extracts. RESULTS: Horses with IBH had significantly higher sLT release than controls with C. nubeculosus and S. vittatum. The highest diagnostic sensitivity and specificity levels were attained when using adult C. nubeculosus extracts with the CAST (78% and 97%, respectively), suggesting that most horses with IBH are sensitised against Culicoides allergens. A proportion of IBH-affected horses was found to be sensitised to allergens of Simulium spp. in addition to those of C. nubeculosus. The CAST with C. nubeculosus had positive and negative predictive values > or = 80% for a true prevalence of IBH of 12-52%. In the follow-up study, the proportion of IBH-affected horses with a positive test result ranged from 90% in November to 68% in March. Severity of clinical signs or age did not influence test results significantly. However, IBH-affected males achieved significantly more positive test results than IBH-affected females. CONCLUSIONS: The CAST with adult C. nubeculosus has high specificity and good sensitivity for diagnosis of IBH. Horses with IBH are mainly sensitised to Culicoides allergens, and some horses are additionally also sensitised to allergens in Simulium spp. POTENTIAL RELEVANCE: The CAST is likely to be a useful test for diagnosis of IBH, even allowing the identification of IBH-affected but asymptomatic horses. This test may also help in further characterisation of allergens involved in this condition.

Magnetic resonance imaging, ultrasonography and histology of the suspensory ligament origin: a comparative study of normal anatomy of warmblood horses

REASONS FOR PERFORMING STUDY: The diagnosis of lameness caused by proximal metacarpal and metatarsal pain can be challenging. Magnetic resonance imaging (MRI) offers the possibility for further diagnosis but there have been no studies on the normal MRI appearance of the origin of the suspensory ligament (OSL) in conjunction with ultrasonography and histology. OBJECTIVES: To describe the MRI appearance of the OSL in fore- and hindlimbs of sound horses and compare it to the ultrasonographic and histological appearance. The findings can be used as reference values to recognise pathology in the OSL. METHODS: The OSL in the fore- and hindlimbs of 6 sound horses was examined by ultrasonography prior to death, and MRI and histology post mortem. Qualitative evaluation and morphometry of the OSL were performed and results of all modalities compared. RESULTS: Muscular tissue, artefacts, variable SL size and shape complicated ultrasonographic interpretation. In MRI and histology the forelimb OSL consisted of 2 portions, the lateral being significantly thicker than medial. The hindlimb SL had a single large area of origin. In fore- and hindlimbs, the amount of muscular tissue was significantly larger laterally than medially. Overall SL measurements using MRI were significantly higher than using histology and ultrasonography and histological higher than ultrasonographic measurements. Morphologically, there was a good correlation between MRI and histology. CONCLUSIONS: MRI provides more detailed information than ultrasonography regarding muscle fibre detection and OSL dimension and correlates morphologically well with histology. Therefore, ultrasonographic results should be regarded with caution. POTENTIAL RELEVANCE: MRI may be a diagnostic aid when other modalities fail to identify clearly the cause of proximal metacarpal and metatarsal pain; and may improve selection of adequate therapy and prognosis for injuries in this region.

Anti-inflammatory and immunosuppressive drugs and reproduction

Rheumatic diseases in women of childbearing years may necessitate drug treatment during a pregnancy, to control maternal disease activity and to ensure a successful pregnancy outcome. This survey is based on a consensus workshop of international experts discussing effects of anti-inflammatory, immunosuppressive and biological drugs during pregnancy and lactation. In addition, effects of these drugs on male and female fertility and possible long-term effects on infants exposed to drugs antenatally are discussed where data were available. Recommendations for drug treatment during pregnancy and lactation are given.

[Antirheumatic therapy and reproduction. The influence on fertility, pregnancy and breast feeding]

Antirheumatic drugs can have a negative effect on reproduction in both men and women. Possible negative effects are impairment of fertility, harmful effects on the fetus and adverse effects on the breastfed child. In women non-steroidal antiinflammatory drugs (NSAID) and cyclophosphamide can impair fertility. In men infertility can result from the use of salazopyrine and cyclophosphamide. A desire for children should be taken into account before the start of disease modifying drugs (DMARD). Treatment with NSAID is possible at some stages of pregnancy as well as during lactation. A limited number of DMARD is compatible with pregnancy and is presented. Cytostatic drugs and leflunomide must be prophylactically withdrawn before a planned pregnancy. TNF alpha antagonists should be discontinued at the start of pregnancy. Safe birth control must be practised during therapy with drugs that are gonadotoxic or teratogenic. Treatment with immunosuppressive drugs during lactation is limited because of insufficient documentation of safety for the breastfed child.

Histomorphologic evaluation of extracorporeal shock wave therapy of the fourth metatarsal bone and the origin of the suspensory ligament in horses without lameness

OBJECTIVE: To determine via histologic examination and scintigraphy the effect of focused extracorporeal shock wave therapy (ESWT) on normal bone and the bone-ligament interface in horses. ANIMALS: 6 horses without lameness. PROCEDURE: Origins of the suspensory ligament at the metacarpus (35-mm probe depth) and fourth metatarsal bone (5-mm probe depth) were treated twice (days 0 and 16) with 2,000 shocks (energy flux density, 0.15 mJ/mm2). One forelimb and 1 hind limb were randomly treated, and the contralateral limbs served as nontreated controls. Bone scans were performed on days -1 (before ESWT), 3, 16, and 19. Histomorphologic studies of control and treated tissues were performed on day 30. RESULTS: ESWT significantly increased the number of osteoblasts but caused no damage to associated soft tissue structures and did not induce cortical microfractures. A significant correlation between osteoblast numbers and radiopharmaceutical uptake was noticed on lateral views of the hind limb on days 3 and 16 and on caudal views of the forelimb on day 3. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that ESWT has the potential to increase osteoblast numbers in horses. The correlation between increased osteoblast numbers and radio-pharmaceutical uptake 3 days and 16 days after the first ESWT suggested that stimulation of osteogenesis occurred soon after ESWT. No damage to bone or the bone-ligament interface should occur at the settings used in this study, and ESWT can therefore be administered safely in horses.

Production of monoclonal antibodies specific for native equine IgE and their application to monitor total serum IgE responses in Icelandic and non-Icelandic horses with insect bite dermal hypersensitivity

Immunoglobulin E forms a minor component of serum antibody in mammals. In tissues IgE is bound by FcvarepsilonRI receptors on the surface of mast cells and mediates their release of inflammatory substances in response to antigen. IgE and mast cells have a central role in immunity to parasites and the pathogenesis of allergic diseases in horses and other mammals. This paper describes the production of several novel monoclonal antibodies that detect native equine IgE in immunohistology, ELISA and Western blotting. An antigen capture ELISA to quantify equine IgE in serum has been developed using two of these antibodies. The mean serum IgE concentration of a group of 122 adult horses was 23,523ng/ml with a range of 425-82,610ng/ml. Total serum IgE of healthy horses was compared with that of horses with insect bite dermal hypersensitivity (IBDH) an allergic reaction to the bites of blood feeding insects of Culicoides or Simulium spp. IBDH does not occur in Iceland where Culicoides spp. are absent, but following importation into mainland Europe native Icelandic horses have an exceptionally high incidence of this condition. In the present study Icelandic horses with IBDH had significantly higher total IgE than healthy Icelandic horse controls (P<0.05). By contrast in horses of other breeds the difference in total serum IgE between those affected with IBDH and healthy controls was not statistically significant. Total serum IgE was also monitored in a cohort of Icelandic horses prior to import into Switzerland and for a period of 3 years thereafter. High levels of serum IgE were present in all horses at the start of the study but dropped in the first year after import. Thereafter the total serum IgE remained low in Icelandic horses that remained healthy but rose significantly (P<0.05) in those that developed IBDH. These results support the conclusion that IBDH is a type I hypersensitivity response to insect allergens but indicate that IBDH in Icelandic horses may have a different pathogenesis from the same condition in other breeds.

Genome-wide search for markers associated with osteochondrosis in Hanoverian warmblood horses

A genome-wide scan was performed to detect quantitative trait loci (QTLs) for osteochondrosis (OC) and osteochondrosis dissecans (OCD) in horses. The marker set comprised 260 microsatellites. We collected data from 211 Hanoverian warmblood horses consisting of 14 paternal half-sib families. Traits used were OC (fetlock and/or hock joints affected), OCD (fetlock and/or hock joints affected), fetlock OC, fetlock OCD, hock OC, and hock OCD. The first genome scan included 172 microsatellite markers. In a second step 88 additional markers were chosen to refine putative QTLs found in the first scan. Genome-wide significant QTLs were located on equine chromosomes 2, 4, 5, and 16. QTLs for fetlock OC and hock OC partly overlapped on the same chromosomes, indicating that these traits may be genetically related. QTLs reached the chromosome-wide significance level on eight different equine chromosomes: 2, 3, 4, 5, 15, 16, 19, and 21. This whole-genome scan was a first step toward the identification of candidate genome regions harboring genes responsible for equine OC. Further investigations are necessary to refine the map positions of the QTLs already identified for OC.

Mapping quantitative trait loci for osteochondrosis in fetlock and hock joints and palmar/plantar osseus fragments in fetlock joints of South German Coldblood horses

The aim of this study was to identify quantitative trait loci (QTL) for osteochondrosis (OC) and palmar/plantar osseous fragments (POF) in fetlock joints in a whole-genome scan of 219 South German Coldblood horses. Symptoms of OC and POF were checked by radiography in 117 South German Coldblood horses at a mean age of 17 months. The radiographic examination comprised the fetlock and hock joints of all limbs. The genome scan included 157 polymorphic microsatellite markers. All microsatellite markers were equally spaced over the 31 autosomes and the X chromosome, with an average distance of 17.7 cM and a mean polymorphism information content (PIC) of 63%. Sixteen chromosomes harbouring putative QTL regions were further investigated by genotyping the animals with 93 additional markers. QTL that had chromosome-wide significance by non-parametric Z-means and LOD scores were found on 10 chromosomes. This included seven QTL for fetlock OC and one QTL on ECA18 associated with hock OC and fetlock OC. Significant QTL for POF in fetlock joints were located on equine chromosomes 1, 4, 8, 12 and 18. This genome scan is an important step towards the identification of genes responsible for OC in horses.

A polymorphism within the equine CRISP3 gene is associated with stallion fertility in Hanoverian warmblood horses

Fertility of stallions is of high economic importance, especially for large breeding organisations and studs. Breeding schemes with respect to fertility traits and selection of stallions at an early stage may be improved by including molecular genetic markers associated with traits. The genes coding for equine cysteine-rich secretory proteins (CRISPs) are promising candidate genes because previous studies have shown that CRISPs play a role in the fertilising ability of male animals. We have previously characterised the three equine CRISP genes and identified a non-synonymous polymorphism in the CRISP1 gene. In this study, we report one non-synonymous polymorphism in the CRISP2 gene and four non-synonymous polymorphisms in the CRISP3 gene. All six CRISP polymorphisms were genotyped in 107 Hanoverian breeding stallions. Insemination records of stallions were used to analyse the association between CRISP polymorphisms and fertility traits. Three statistical models were used to evaluate the influence of single mutations, genotypes and haplotypes of the polymorphisms. The CRISP3 AJ459965:c.+622G>A SNP leading to the amino acid substitution E208K was significantly associated with the fertility of stallions. Stallions heterozygous for the CRISP3 c.+622G>A SNP had lower fertility than homozygous stallions (P = 0.0234). The pregnancy rate per cycle in these stallions was estimated to be approximately 7% lower than in stallions homozygous at this position.