A LOVAK AGY-GERINVELŐ FOLYADÉKÁNAK MINTAVÉTELE, ELEMZÉSE ÉS EGYES PARAMÉTEREKRE VONATKOZÓ SAJÁT REFERENCIATARTOMÁNY MEGHATÁROZÁSA

Az agy-gerincvelő folyadék (liquor) egy színtelen, tiszta, fiziológiásan nem alvadó, a vízéhez hasonló viszkozitású folyadék, mely körülveszi és határolja a központi idegrendszert, szerepe van annak védelmében, tápanyaggal való ellátottságában, valamint a koponyaűri nyomásingadozások kiegyenlítésében. A liquor egyes paramétereinek laboratóriumi kiértékelése hasznos diagnosztikai eszköz a központi idegrendszer megbetegedéseinek meghatározásában, vagy a betegség eredetének lokalizációjában. Ahhoz, hogy az egyes betegségek okozta, egyes paraméterekre jellemző elváltozásokat vizsgálni tudjuk a liquorban, szükséges adott laboratóriumra jellemző referencia tartományok meghatározása. Ez mindinkább szükséges, mivel bizonyos paraméterekre vonatkozó referencia adatok nem, vagy csak hiányosan szerepelnek a nemzetközi szakirodalomban. Szakirodalmi útmutatás alapján, a referencia tartomány meghatározásához 20, egészséges lóból gyűjtöttünk agy-gerincevlői folyadék mintát. 12 minta származott atlanto-occipital (AO), és 8 darab lumbo-sacral (LS) punkcióból. Az AO mintavétel általános anesztéziában, az LS mintavétel álló helyzetben, bódításban történt. A kapott minták fizikális vizsgálatát követően, meghatároztuk azok vörösvérsejt-, magvas sejtszám és fehérje tartalmát, biokémiai összetevőit (aszpartát-transzamináz, alkalikus-foszfatáz, glükóz, tejsav, kreatin-kináz, laktát-dehidrogenáz), valamint egyéb, mérhető összetevőit (nátrium, kálium, kalcium, klorid, anorg. foszfát, magnézim). A mintából készített kenetet külső laboratóriumba küldtük citológiai vizsgálatra. A referencia tartomány meghatározása a következő képpen történt. Korábbi szakirodalmi referencia tartományokhoz hasonlítottuk a kapott adatokat. Amennyiben a 20 eredményből 18 beleesett a publikált tartományba, akkor ezt, a már meglévő referencia tartományt elfogadtuk saját laboratóriumunk részére. Amennyiben kevesebb, mint 18 mért adat felelt meg a leírtaknak, akkor erre az adatra vonatkozóan azt nem elfogadhatónak kell tekinteni és újabb 20 mintát gyűjteni a jövőben. Eredményeink közül a kapott magvas sejtszám, a vörösvérsejtszám, az ultraszenzitív összfehérje, a glükóz, az aszpartát transzamináz, a nátrium, a kálcium és a kálium értékek beleestek a leírt tartományba, azonban esetünkben jóval magasabb tejsav és laktát dehidrogenáz, valamint eltérő kreatinin-kináz értékeket mértünk. Vizsgált adataink közül legtöbb esetben sikeresen tudtunk referencia tartományt meghatározni. A maradék paraméterekre további 20 egészséges minta gyűjtésére van szükség ahhoz, hogy megtudjuk, ezek az értékek a mintagyűjtés folyamán emelkedtek meg a liquorban, vagy esetleg egy teljesen új, magasabb értékeket tartalmazó referencia táblára van esetükben szükség. A liquorban a CK és LDH izoenzimek külön mérése, illetve ezen enzimek és a laktát párhuzamos vizsgálata a vérben szintén segítene az eredményeink további értékelésében. Köszönet a vizsgálatok elvégzésének lehetőségéért Némedi Istvánnak, Sári Viktornak, a Haszonállat Tanszék és Klinika laboratóriumának. A vizsgálatok elvégzését az NKB 2013/15918 számú pályázata tette lehetővé

Equine multinodular pulmonary fibrosis (EMPF): Five case reports

Equine multinodular pulmonary fibrosis (EMPF), a progressive fibrosing interstitial lung disease has been associated with gammaherpesviruses. This case series describes five horses with EMPF. Three of the horses (two in Hungary, one in the Czech Republic) were diagnosed with EMPF ante mortem. They presented with typical clinical signs of EMPF including dyspnoea and weight loss. Arterial blood gas analysis revealed hypoxaemia. Blood work showed signs of inflammation like neutrophilia and hyperfibrinogenaemia. An endoscopic examination of the respiratory tract including cytology and culture of tracheobronchial secretion and bronchoalveolar lavage were performed, revealing secondary bacterial infection in one case. A suspected diagnosis of EMPF was made on the basis of a positive EHV-5 PCR from bronchoalveolar lavage and the findings of thoracic radiographs and ultrasound examination. In one case the diagnosis was confirmed by lung biopsy. All horses died or had to be euthanised despite treatment. Two horses (from Austria) were diagnosed with EMPF post mortem. They not only had EMPF but also concurrent other diseases which seemed to be associated with immunosuppression. Three horses showed the discrete form and two horses the diffuse form of EMPF. EHV-5 DNA was identified in lung tissue of all horses by PCR.</jats:p

Cerebrospinal fluid parameters of horses with West Nile virus encephalomyelitis

West Nile virus (WNV) is a mosquito-borne zoonotic arbovirus transmitted in natural cycles between mosquitoes and wild birds. Horses and humans are incidental, dead-end hosts, but can develop severe neurological disorders. By its close contact with the extracellular fluid of the brain, analysis of cerebrospinal fluid (CSF) composition can reflect biological central nervous system (CNS) impairments enabling the diagnosis and understanding of various neurodegenerative CNS disorders. Fifteen CSF samples were collected from horses with acute neurologic symptoms and positive WNV IgM ELISA on their sera. CSF samples of twenty healthy horses without any neurologic disease were used as controls. Biochemical and cytological parameters were evaluated and compared. Most of the data obtained from the WNV affected horses did not seem to follow a normal distribution, but protein, creatine-kinase, aspartate-aminotransferase, lactate-dehydrogenase, alkaline-phosphatase, magnesium, glucose, and lactate showed abnormal levels in a number of cases. None of the 6 horses with elevated glucose levels survived (<=0.36, modified Wald method with 90% CI). Opposite to previous equine studies we have found neutrophilic pleocytosis in 54% of cases. Measured data also indicates that CSF neutrophilia is more likely to be found parallel with high protein content (Fisher exact test, two tailed, p = 0.1026). The CSF findings with WNV neuroinvasive disease are nonspecific and variable. Neutrophils are likely play a role in the development of inflammatory response and brain damage. Increased enzyme levels reflect rather CNS injury than blood-brain barrier damage. Elevated glucose levels might be secondary to increased plasma levels and predict outcome

Cerebrospinal fluid parameters of horses with West Nile virus neuroinvasive disease

Objective: To compare biochemical and cytological findings of cerebrospinal fluid (CSF) samples in horses with acute neuroinvasive West Nile virus (WNV) infections with those of control healthy horses. Design: Retrospective case-control study. Samples: Fifteen CSF samples from horses with acute WNV neuroinvaisve disease (WNVND)and twenty from healthy horses. Procedures: WNVND was diagnosed based on acute neurologic symptoms and positive IgM ELISA results. CSF samples were collected either from the atlanto-occipital or the lumbosacral sites. Results: CSF results of the WNV affected group did not follow normal distribution. Protein,creatine-kinase, aspartate-aminotransferase, lactate-dehydrogenase, alkaline-phosphatase,magnesium, glucose, and lactate concentrations showed abnormal levels in a number of WNV cases. None of the 6 horses with elevated glucose concentrations survived (<=0.36, modified Wald method). Opposite to previous equine studies we have found neutrophilic pleocytosis in 54% of cases. Measured data also indicates that CSF neutrophilia is more likely to be found parallel with high protein content (Fisher exact test, p = 0.1026). Conclusions and clinical relevance: The CSF findings with WNVND are nonspecific and variable. Neutrophils likely play a role in the development of inflammatory response and brain damage. Increased enzyme activities and changes in the electrolyte concentrations reflect CNS cellular injury rather than blood-brain barrier leakage. Although elevated glucose levels reliably predicted outcome, these results might be the consequences of increased plasma levels and reflectgeneral stress rather than any CNS pathophysiology. Examination of CSF is most useful when the results are correlated with history, clinical findings and ancillary laboratory studies

EQUINE HERPESVIRUS TYPE 5 IN BRONCHOALVEOLAR LAVAGE FLUID OF HORSES WITH EQUINE MULTINODULAR PULMONARY FIBROSIS (EMPF) AND WITH OTHER CHRONIC RESPIRATORY DISORDERS.

The aim of the study was to estimate the prevalence and the potential role of equine herpesvirus type 5 (EHV-5) in the bronchoalveolar lavage fluid (BALF) samples of horses with chronic respiratory signs. Altogether 60 horses with chronic respiratory signs of minimum of 2 weeks duration were involved in the study. Horses were clinically examined, respiratory endoscopy, thoracic radiography, ultrasonography, tracheal culture and evaluation of BALF cytology was performed. EHV-5 PCR assay was carried out on BALF samples of 15 horses in a commercial laboratory (IDEXX-VetMedLab Germany), 54 BALF samples were tested in the research laboratory of Szent Istvan University, Faculty of Veterinary Science and samples of 9 horses were tested by both laboratories. PCR testing by the two different laboratories gave homogenous results. Altogether there were 7 horses (prevalence: 11,6%) with positive PCR results. Three of them were diagnosed with EMPF based on histologic results of lung biopsy specimens or post-mortem tissue samples. These three horses were genetically closely related warmbloods. Two other horses suffered of suspected EMPF based on thoracic radiograpy with nodular interstitial pattern, EHV-5 positivity and intranuclear inclusion bodies but pulmonary biopsy was not performed on any of them. One positive horse was diagnosed with inflammatory airway disease and one with systemic granulomatous disease. Presence of EHV- 5 in BALF significantly (Fisher test, p < 0,001) correlated with the diagnosis of EMPF. BALF testing for EHV-5 is an important examination when establishing the diagnosis of EMPF. Genetic predisposition might render the patient more susceptible to EMPF or viral infection

Comparison of assays for the detection of West Nile virus antibodies in equine serum after vaccination

Introduction: The West Nile virus (WNV) mainly infects birds, horses and humans. Outcomes of the infection range from light uncharacteristic symptoms to fatal neurologic disease. The Hungarian equine WNV outbreak reported in 2008 was the first caused by a lineage 2 sub-Saharan strain in Europe. To protect horses from serious disease an inactivated virus vaccine is available in Hungary since 2009. Objectives: The main objectives of the present study were to measure serum IgG and IgM antibodies in naturally exposed and vaccinated horses, and to compare hemagglutination-inhibition test (HIT) results with those of competitive and IgM antibody capture (MAC) enzyme-linked immunosorbent assays (ELISA). Methods: Altogether 268 animals were tested with HIT for WNV antibodies and 34 horses were examined for both IgG and IgM with HIT and ELISA simultaneously. After primary screening for WNV antibodies, all horses were vaccinated. Samples were taken immediately before and 3-5 weeks after each vaccination. McNemar chi-squared and percent agreement tests were used to detect concordance between HIT and ELISA. Results: Analyses by HIT confirmed the presence of WNV antibodies in 38/123 (30,89%) from naturally exposed horses. Sera from 63/72 (87,5%) vaccinated animals were positive before the first booster and from 11/11 (100%) before the second booster. HIT and ELISA tests were non-concordant with a low positive percentage agreement value concerning the IgM measurements. HIT was less sensitive when detecting IgG antibodies. We could detect post vaccination IgM in 9 cases with MAC ELISA and in 7 cases with HIT. Discussion and conclusions: WNV is endemic in Hungary causing regular natural infections. Antibodies could not be detected in each individual case 12 months after primary injections, protection is more reliable after the first yearly booster. Based on our findings it may not be possible to differentiate infected horses from recently vaccinated horses using the MAC ELISA. HIT does not substitute ELISA when detecting IgM in acute infections. The study was permitted by the Animal Health and Welfare Directorate of National Food Chain Safety Office (22.1./1606/003/2009). Aknowledgement to Bólyai János research Grant

Comparison of assays for the detection of West Nile virus antibodies in equine serum after natural infection or vaccination

West Nile virus (WNV) mainly infects birds, horses and humans. Outcomes of the infection range from light uncharacteristic signs to fatal neurologic disease. The main objectives of the present study were to measure serum IgG and IgM antibodies in naturally exposed and vaccinated horses and to compare results of hemagglutination-inhibition test (HIT), enzyme-linked immunosorbent assay (ELISA) and plaque reduction neutralization test (PRNT). Altogether 224 animals were tested with HIT for WNV antibodies and 41 horses were simultaneously examined with ELISA and PRNT. After primary screening for WNV antibodies, horses were vaccinated. Samples were taken immediately before and 3-5 weeks after each vaccination. McNemar chi-squared and percent agreement tests were used to detect concordance between HIT, ELISA and PRNT. Analyses by HIT confirmed the presence of WNV antibodies in 27/105 (25.71 %) from naturally exposed horses. Sera from 57/66 (86.36%) vaccinated animals were positive before the first booster and from 11/11 (100%) before the second booster. HIT was less sensitive when detecting IgG antibodies. We could detect post vaccination IgM in 13 cases with MAC-ELISA and in 7 cases with HIT. WNV is endemic in Hungary causing regular natural infections. Protective antibodies could not be measured in each individual case 12 months after primary injections; protection is more reliable after the first yearly booster. Based on our findings it was not be possible to differentiate infected horses from recently vaccinated horses using IgM antibody capture ELISA (MAC-ELISA). HIT does not substitute ELISA or PRNT when detecting IgG, but was useful tool in this study to gain statistical information about the tendencies within a fixed population of horses