Cultivation of Schwann cells from fresh and non-fresh adult equine peripheral nerves.

BACKGROUND: Over the past 25 years, acquired equine polyneuropathy (AEP) has emerged as a neurological disease in Scandinavian horses. This condition is characterized by histopathological features including the presence of Schwann cell (SC) inclusions. Cultivated equine SCs would serve as a valuable resource for investigations of factors triggering this Schwannopathy. Ideally, cells should be sampled for cultivation from fresh nerves immediately after death of the animal, however the availability of fresh material is limited, due to the inconsistent case load and the inherent technical and practical challenges to collection of samples in the field. This study aimed to cultivate SCs from adult equine peripheral nerves and assess their ability to survive in sampled nerve material over time to simulate harvesting of SCs in field situations. NEW METHODS: Peripheral nerves from five non-neurological horses were used. After euthanasia, both fresh and non-fresh nerve samples were harvested from each horse. Flow cytometry was employed to confirm the cellular identity and to determine the SC purity. RESULTS: The results revealed successful establishment of SC cultures from adult equine peripheral nerves, with the potential to achieve high SC purity from both fresh and non-fresh nerve samples. COMPARISON WITH EXISTING METHOD: While most SC isolation methods focus on harvest of cells from fresh nerve materials from laboratory animals, our approach highlights the possibility of utilizing SC cultures from field-harvested and transported nerve samples from horses. CONCLUSIONS: We describe a method for isolating SCs with high purity from both fresh and non-fresh peripheral nerves of adult horses.

Epidemiological Aspects of Equid Herpesvirus-Associated Myeloencephalopathy (EHM) Outbreaks.

Equid Herpesvirus Myeloencephalopathy (EHM) is a multifactorial disease following an EHV-1 infection in Equidae. We investigated a total of 589 horses on 13 premises in Europe in search of risk factors for the development of EHM. We found that fever (p < 0.001), increasing age (p = 0.032), and female sex (p = 0.042) were risk factors for EHM in a logistic mixed model. Some breeds had a decreased risk to develop EHM compared to others (Shetland and Welsh ponies; p = 0.017; p = 0.031), and fewer EHV-1-vaccinated horses were affected by EHM compared to unvaccinated horses (p = 0.02). Data evaluation was complex due to high variability between outbreaks with regards to construction and environment; viral characteristics and the virus's transmissibility were affected by operational management. This study confirms earlier suspected host-specific risk factors, and our data support the benefit of high vaccine coverage at high-traffic boarding facilities.

The first reported Florida clade 1 virus in the Nordic countries, isolated from a Swedish outbreak of equine influenza in 2011.

Equine Influenza Virus (EIV) is a major cause of respiratory disease in horses and the virus constantly undergoes antigenic drift. Here we characterize and describe the HA1 and the NA genes of H3N8 within samples obtained from outbreaks in Sweden during November-December 2011. Both clade 1 and clade 2 viruses of the Florida sublineage were identified. The index case of clade 2 was transported to Sweden from Spain through the Netherlands, whereas the clade 1 had its origin from a Swedish stud farm. The clade 1 virus was efficiently spread between training yards by unvaccinated young horses, but vaccinated horses were also presented with clinical signs of respiratory disease. No virus of the Eurasian lineage was isolated during this outbreak. Clade 1 has previously been described in outbreaks in numerous of other countries, but this is the first time it has been detected in Sweden. The results from this study shows the importance of including both clade 1 and clade 2 of the Florida sublineage in equine influenza vaccines, supporting the ESP and OIE recommendations.

Spatial and temporal distribution of incidence of acquired equine polyneuropathy in Norway and Sweden, 1995-2012.

BACKGROUND: Acquired equine polyneuropathy (AEP) is an emerging disease in horses in Sweden, Norway and Finland since 1995. Affected horses show bilateral pelvic limb knuckling and weakness, sometimes progressing to recumbency and euthanasia. The aetiology is unknown but is thought to be non-infectious and non-genetic, though possibly toxic or toxico-infectious. The objectives of this study were to describe the spatial, temporal and spatio-temporal features of AEP in Norway and Sweden for the period of 1995 to 2012. Data from all documented case farms (n = 136) were used. Space-time interaction clustering of case farms was investigated with a retrospective space-time scan statistic with a space-time permutation model, the space-time K-function and the Jacquez k nearest neighbour (kNN) test. RESULTS: There was a clear seasonality in disease occurrence, as 123 case farms presented their first case from January to May. However, there was large variation in the number of case farms between years. Case farms were more numerous in certain regions. Despite the larger horse population in Sweden, 120 of the case farms were in Norway. Space-time clustering was supported by the K-function and partly by the space-time scan, but not by the Jacquez k nearest neighbour (kNN) test. CONCLUSIONS: The results suggest an aetiology for AEP where the exposure is not consistent in time, but varies during and between years, assuming that the incubation period does not vary greatly. The results further suggest that the exposure varies between regions as well. Two out of three different analytical methods supported spatio-temporal clustering of case farms, which rendered inconclusive results. The negative result in the kNN test might be explained by lack of power, which is due to the small number of outbreaks in relation to the size of the study area and length of the study period, and further by the low to moderate power of methods to detect space-time clustering when the background population is unknown. Further research is needed to study how management, meteorological variables and other factors with local or regional differences may explain outbreaks of AEP.

Automated counting of nucleated cells in equine synovial fluid without and with hyaluronidase pretreatment.

BACKGROUND: Microscopy is usually used to obtain manual total and differential cell counts in equine synovial fluid. A faster, more precise method is desirable. OBJECTIVES: The objectives were to compare an automated impedance method with a manual method for obtaining total and differential cell counts in equine synovial fluid and to evaluate the effect of pretreatment with hyaluronidase on automated results. METHODS: Synovial fluid samples (n=48) were collected into EDTA and analyzed within 48 hours. Automated total and differential cell counts were evaluated using a Medonic CA620-VET hematology analyzer before and after pretreatment for 5-30 minutes with hyaluronidase (final concentration 0.01 mg/mL). A hemacytometer count and microscopic evaluation of a direct smear were used as the reference method. Intra-assay coefficients of variation (CV) were determined. RESULTS: Thirty-one of 46 untreated samples and 0/46 hyaluronidase-treated samples were error-flagged by the analyzer. Correlation between automated (ANCC) and manual (MNCC) nucleated cell counts in untreated samples (n=15; R(2)=0.93) and pretreated samples (n=46; R(2)=0.94) was high, and pseudomedian difference was low. Intra-assay CVs for samples with medium and high cellularity were significantly lower for ANCC (1.5-2.7%) compared with MNCC (6.1-15.7%) (P<.01). Valid automated differential cell counts were not obtained. CONCLUSIONS: Automated total cell counts obtained on the Medonic analyzer correlate well with manual counts in equine synovial fluid; however, pretreatment with hyaluronidase is required to minimize error flags. Automated differential counts are not accurate for synovial fluid.