Revista - SciELO Colombia · Arteritis Viral Equina: perspectivas epidemiológicas y de intervención Arterite Viral Eqüina: Perspectivas Epidemiológicas e de Intervenção Julián

501Ruiz-Sáenz J. Equine Viral Arteritis

Rev Colomb Cienc Pecu 2010; 23:501-512

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¤ Tocitethispaper:RuizSáenzJ.EquineViralArteritis:epidemiologicalandinterventionperspectives.RevColombCiencPecu2010;23:501-512.* Correspondenceauthor:JuliánRuízSáenz.UnidaddeDiagnóstico,FacultaddeCienciasAgrarias.UniversidaddeAntioquia.Carrera75No.65-87.Ciuda-deladeRobledo.Medellín,Colombia.Tel:(574)2199160.E-mail:[email protected].

Equine Viral Arteritis: epidemiological and intervention perspectives¤

Arteritis Viral Equina: perspectivas epidemiológicas y de intervención

Arterite Viral Eqüina: Perspectivas Epidemiológicas e de Intervenção

JuliánRuiz-Sáenz1*,MV,MSc

1Grupo de Investigación CENTAURO, Facultad de Ciencias Agrarias, Universidad de Antioquia, A.A. 1226, Medellín, Colombia.

(Received: 10 march, 2009; accepted: 14 september, 2010)

Summary

Equine viral arteritis is an infectious viral disease of horses that causes serious economic losses due to the presentation of abortions, respiratory disease and loss of performance. After the initial infection males can become persistently infected carriers scattering infection through semen, a situation that brings indirect economic losses by restrictions on international trade of horses and semen from breeding and from countries at risk of infection with the virus. Outbreaks of infection have been reported in several American countries with which Colombia has active links of import and export of horses and semen. This article focuses on creating awareness of the key points of infection and on examples of how infection has been distributed in countries that have developed horse breeding, such as Argentina, a country aware of the origin and distribution of the infection. This study will help us understand how to assess, prevent and/or control the entry of such a devastating infection for the equine industry.

Key words: equine arteritis virus, prevention and control, viral infection.

Resumen

La arteritis viral equina es una enfermedad viral infectocontagiosa de los equinos que puede llegar a causar graves pérdidas económicas debidas a la presentación de abortos, enfermedad respiratoria y pérdida del desempeño; luego de la primoinfección los machos pueden resultar infectados persistentemente convirtiéndose en portadores los cuales diseminan la infección a través del semen; hecho que acarrea pérdidas económicas indirectas por las restricciones en el comercio internacional tanto de ejemplares como de semen procedente de criaderos y países con riesgo de infección con el virus. Se han reportado brotes de

Ruiz-Sáenz J. Equine Viral Arteritis502


infección en diversos países de América con los cuales Colombia tiene activos vínculos de importación y exportación de equinos y semen. El presente artículo trata de dar a conocer los puntos clave de la infección y ejemplos de cómo se ha distribuido la infección en países con sistemas de explotación de equinos tales como Argentina, en el cual se conoce el origen de la infección y su distribución; cuyo estudio, nos ayudará a entender cómo debemos evaluar, prevenir y/o controlar la entrada de una infección devastadora para la industria equina nacional.

Palabras clave: infección viral, prevención y mitigación, virus de la arteritis equina.

Resumo

A arterites viral eqüina é uma doença infecto-contagiosa dos eqüinos que pode chegar a causar graves perdas económicas devidas à apresentação de abortos, enfermidade respiratória e perda do desempenho; logo da primoinfecção, os machos podem resultar infectados persistentemente, convertendo-se em portadores e disseminam a infecção a través do sémen; fato que acarreia perdas económicas indirectas pelas restrições no comercio internacional de reprodutores e sémen procedente de criatórios e países com risco de infecção com o vírus. Tem-se reportado surtos de infecção em diversos países da América, com os quais Colômbia tem activos vínculos de importação e exportação de equinos e sémen. O presente artigo trata de dar a conhecer os pontos chave da infecção e exemplos de como tem-se distribuído a infecção em países com sistemas de exploração de eqüinos como Argentina, na qual se conhece a origem da infecção e sua distribuição; para o qual o estudo, nos ajudará a entender como devemos avaliar, prevenir e/ou controlar a entrada da infecção devastadora para a industria eqüina nacional.

Palavras chave: vírus, infecção viral, prevenção e mitigação.

Introduction

Equine viral arteritis (EVA) is a contagiousdisease of horses caused by an enveloped RNAvirus, knownasEquine arteritis virus (EAV).EAVis the prototype virus of the genus Arterivirus,familyArteriviridae, orderNidovirales (Cavanagh,1997; ICTVdB, 2006). Based on their geneticstructure and replication strategy, three additionalviruseshavebeenclassified in the samegenusandfamily.They are of great importance, one of themcausing Porcine Reproductive and RespiratorySyndrome(PlagemannandMoennig,1992).

Based on extensive comparative studies atthe genomic and antigenic level of EAV, onlyone serotype has been detected, usually calledthe Bucyrus strain (McCollum, 1969). However,temporal and geographic diversity among EAVisolates has been demonstrated (Murphy et al.,1992). There also exists some variability betweenthe strains based on their pathogenicity, somestrains being capable of causing a wide range ofclinical symptoms, collectively referred to as EVAinsusceptiblehorses,whereasothersproduceonlyaslightfever(Balasuriyaet al.,1995)

Although through the years it has beenrecognized that EAV causes contagious abortionin pregnantmares (Doll et al., 1957), and that thevirus causes abortion outbreaks (Balasuriya et al.,1998), some evidence suggests that not all strainsof the virus have abortigenic properties, somestrainscausingonlyclinicallymildorasymptomaticdiseases such that they are considered strongcandidates to be used as vaccine strains (Moore et al., 2002, Moore et al., 2003). However, froma practical standpoint, it is difficult to distinguishbetween EAV strains that can cause abortion andthose which do not; additionally, there are nostandard techniques toclassify thestrainsbasedontheir abortiveproperties.Therefore, fromaclinicalpoint of view, all strains of the virus should beconsidered potentially abortigenic unless provedotherwise(Timoney,2003).

EAV is not particularly resistant to exposureto the environment and is rapidly inactivated by awide rangeofphysico-chemical conditions,mainlytemperature,survivingonly20-30minutesat56ºCand2-3days at 37 °C.Additionally, it can surviveupto75daysat4ºC,andevenyears influidsandtissuespreservedat-70ºC.However, theviruscan



rapidly be inactivated by low humidity, sunlight,and theuseof lipid solvents, or byusing commoninactivating detergents (Burky, 1966, Shirai et al.,2000).Yet, since virus viability upon refrigerationor freezing is high, EAV can remain infective infrozen semen for long periods of time, even years(Timoney,2000).

Background

Morethanacenturyago,reportswerepublishedin the European veterinary literature concerninga horse disease whose clinical features wereconsistentwith thosedescribed forEVA.However,the virus was not isolated until 1953, during anoutbreak of abortions and respiratory illness in a“Standardbred”horsebreedingranchnearBucyrus,Ohio(Doll,1957)

The most important outbreak of EVA inAmericaoccurredin1984,whenthediseasestruckmany racing thoroughbred farms in Kentucky.This outbreak led to two very important findingsregarding the EVA: the efficiency of venerealtransmissionofthevirusbyaninfectedstallion,andthe high tendency of transmission as a carrier thata stallion has after a natural infection (Timoney,1984).The fear thatahighlypathogenicstrainhademerged, joined with the belief that most horsepopulations are susceptible to the virus, led toan increased evaluation of the importance of thedisease worldwide, imposing severe restrictionson the movement of horses with positive titersof antibodies against the virus (Timoney andMcCollum, 1993a). These measures weresubsequently gradually decreased, with today agreater emphasis on controlling the internationaltradeofcarrierstallionsandinfectedsemen,whichhave frequently been implicated in the spread ofthe virus within and between countries (Timoney,2000b)

According to the World Animal HealthOrganization-OIE, a carrier stallion has serologicalpositive for antibodies to the virus using the virus

neutralisation test or an appropriately validatedenzyme-linked immunosorbent assay (ELISA)and also positive virus isolation from the semen(Timoney,2004).

Transmission

EVA is primarily a respiratory disease,transmitted through inhalation of viral particlesbetween horses during the acute stage of infectionmainly during transport of the animals for trade,exhibition or racing.The virus can be excreted bynasal secretions until 16 days post-infection, andthis is a source of infection for horses that are inclosecontact(McCollum,1971).

However,unlikeotherrespiratorydiseases,EAVcanalsobetransmittedasavenerealdiseaseduringinsemination, either by natural mating or artificialinsemination, because the virus may be excretedin the semen,which is amajor sourceof infection(Figure 1).When a mare, a gelding or a sexuallyimmature colt catches the disease, the animalexcretes the virus through respiratory airwaysand develops a strong immunity to re-infection(Fukunagaet al.,1981).Incontrast,adultmalesarevery likely tobecomecarriersof thevirusfor longperiodsoftimeandcantransmitthevirustomaresduringmating(Neu et al.,1988).Althoughthemareeasilyeliminatesthevirus,apregnantEAV-infectedmarecantransmitthevirustothefetus.

Verticaltransmissioncanoccurbythecongenitalroute, and depending on how advanced thepregnancyis,thefetuscanbecomeinfected,dieandbe aborted between 9 and 30 days after infection.At times a live but congenitally infected and sickfoal is born (Vaala et al., 1992); in such cases,the placenta, the placental fluids and the fetus areimportant sources of infectious virus (Figure 1).Additionally, indirect transmission can occur in aniatrogenic manner by using utensils contaminatedwith nasal secretions/excretions or abortion tissuesorsecretions(TimoneyandMcCollum,1993b)



Figure 1. EAV transmission in horses on farms and during an outbreak (Adapted from Holyoak et al., 2008)

Clinical findings

There are considerable variations in theclinical symptoms and in severity of infection,and although many horses infected with EAV donot show symptoms, it has been demonstratedthat experimental infection can be fatal (Glaseret al., 1996,MacLachlanet al., 1996).As inmostinfectious diseases, certain host factors determinethe severity of the disease, such as age, immunestatus,sexwhichdeterminesthepossibilityofbeingapotentialcarrier,etc.Other factorsdependon theagentsuchastherouteofexposure,viralstrainandinfectivedose.

After an incubation period varying from2-14 days (6-8 days after venereal exposure) themost prominent symptom is fever of about 41°C that may last between 2 and 12 days, usuallyconcomitant with depression and anorexia,

conjunctivitis and rhinitis with nasal and oculardischarge, leucopenia, peripheral or supraorbitaledema,andinflammationandedemaofthescrotum,prepuce, mammary glands and limbs (Figure 2).Urticariathatmaybelocalizedinthefaceandneckorgeneralizedinmostof thebodyisalsocommon(TimoneyandMcCollum,1993b)

Another symptom of infection in pregnantmares is abortion, that is not preceded by anycharacteristic or premonitory sign and may occureither at the end of the acute phase of illness orwhen the recovery phase begins (Coignoul andCheville, 1984; Vaala et al., 1992). Abortionshave been observed between 3 and 10 months ofgestationinbothnaturalandexperimentalinfectionsand abortion rates in the different outbreaksreported have varied from low levels of 10% orup to dramatic levels of 50-60% (Timoney andMcCollum,1993b).



During acute infection, stallions may showtemporal subfertility/infertility, associated withdecreased libido, low spermmotility/concentrationand low percentage of morphologically normalsperm; these changes are mainly due to the highscrotal temperatureandnot toapathologicaleffectinduced by the virus. These changes persist forabout16weeks,atwhich time thehorse returns toits normal seminal parameters (Neu et al., 1992).Additionally, carrier stallions can excrete virusin the semen and exhibit a normal semen quality(Timoney et al., 1987, Timoney and McCollum,1993b). As for other members of the Equidae family, outbreaks have not been reported amongdonkeys or mules, and although antibodies havebeen identified, there is little information on theclinical signs observed in these species (TimoneyandMcCollum,1993b).

Pathogenesis

Two days after aerosol infection, EAV spreadsrapidly to the bronchial and pulmonary lymph

nodes, reaching the bloodstream and spreadingthroughout the body. It is therefore possible toisolate the virus in a wide variety of tissues andbody fluids,making isolation of the virus possible2 days post-infection in nasopharyngeal swabsamples, 19 days in white cells, and up to 9 daysin serum or plasma (Figure 2). Virus clearancecoincides with the development of specificneutralizing antibodies against EAV (Timoney andMcCollum,1993b).

During primary viremia, EAV infects andreplicates in endothelial cells causing strongmulti-vessel damage to the endothelium, thesubsequent internal elastic lamina and affectingthemiddlemuscular layerof thevessels.Vasculitisis characterized by fibrinoid necrosis of smallarteries with extravasation of red blood cells andproteinaceous material. Additionally, the virusinfects macrophages, which in conjunction withendothelialcelldamageleadstoincreasedreleaseofproinflammatorycytokinesthatareheavilyinvolvedin the pathogenesis of “arteritis” (Moore et al.,2003).

Figure 2. Clinical and laboratory findings, and sequential pathogenesis of EAV after respiratory infection. DPI: days post-infection; vertical bars represent the chronological occurrence of each clinical or laboratory finding and virus distribution in tissues and body fluids (Adapted from Holyoak et al., 2008).



Experimentally,ithasbecomeclearthatabortionis primarily the result of fetal infection, other thanmyometritis or damage to the placenta, leading toexpulsionofthefetus;inaddition,thereisevidencethat the tissues of the aborted fetus tissues havehigherviraltiterscomparedtothemother,stressingthe high level of virus replication in the fetus(MacLachlanet al.,1996).

Virus reservoir

The natural reservoir of EAV infection is thecarrier stallion, which ensures the permanence oftheinfectioninequinepopulations.Thecarrierstatehasbeenidentifiedonlyinthestallion,notinmares,geldingsorsexuallyimmaturefoals(Timoneyet al.,1987). Virus can persist in the stallion for weeks,monthsoryears, even for life in some individuals.About 60% of the stallions that acquire the virusby the respiratory route may become persistentlyinfected; that is why the frequency of carriers inanystallionpopulationcanvaryfromlessthan10%to over 70% (Timoney andMcCollum, 1993b). Inthe persistently infected stallion, EAV is locatedin the accessory sexual glands and establishmentand persistence of the carrier state is testosterone-dependent (Little et al., 1992). Carrier stallionsconstantly eliminate the virus in the semen andconsequently the risk of transmission of infectionis limitedtothetimeofmating.Transmissionratesrangefrom85to100%,whethermatingisnaturalorartificialinseminationisperformed(Figure1).

A lesser percentage of long-term carrierstallions can clear the virus spontaneously fromthe reproductive tracts and do not show risk fortransmissionofinfection(TimoneyandMcCollum,2000). However it has been shown that carrierstallions are the natural source of genetic andphenotypicdiversityofthevirus,creatingnewrisksofemergenceofviralvariantswithnewpathogenicpotential (Balasuriya et al.,1999, Hedges et al.,1999,Balasuriyaet al.,2004).

Diagnosis

Given the clinical similarity of EVAwith otherequine infectious and non-infectious diseases, any

presumptive diagnosis should always be confirmedby laboratory tests (Holyoak et al., 2008). Thepresence of EVA must be suspected if respiratorysymptoms are accompanied by abortions. Virusisolates can be obtained from nasopharyngealswabs or washes, conjunctival swabs, samplesof ejaculates, essentially the sperm-rich fraction,placentas,fetalfluidsandtissuesofabortedfetuses,suchaslung,spleenandlymphnodes,andthroughbloodsampleswithEDTAasanticoagulant.

The most widely used method for diagnosis isthe evaluation of neutralizing antibodies, althoughthe presence of such antibodies does not indicateactiveinfection,butratherthattheanimalhasbeenexposed to EAV.An active infection is diagnosedwhen high levels of antibodies in a single sampleare found or when an increase in antibody titersoccurs in two paired blood samples, obtained 14-28 days apart (Holyoak et al., 2008). Althoughseveraltypesofimmunoassays(ELISAs)havebeendeveloped, given the high specificity of the virusneutralization test to detect individuals with lowserumantibodytiters, this testhasbeenestablishedby the World Animal Health Organization-OIEas the gold standard for international transport ofhorses(Timoney,2004)

Treatment

Since there is no specific treatment for EVA,oncetheinfectionisconfirmedclinicalmanagementshouldincluderest,fluidsandinsomecases,broad-spectrumantibioticstoreducetheriskofsecondarybacterialinfections.Adulthorsesfullyrecoverfromthe disease, leaving only the recovered stallions ascarriersandsourcesofinfection.

Prevention and control

Although in Colombia, EVA is an exoticdisease, some prevention and control measureshavebeenwidelyreported, thatcanmakeinfectiona manageable disease, and most importantly, canreduce the direct economic losses produced bydisease outbreaks. There is currently an avirulentlive vaccine (Arvac®, Fort Dodge Animal HealthLaboratories, Iowa, USA) that is safe, effective



and economical, and has been used in otherLatin-American countries. The combination ofvaccination with the use of isolation measures ofunvaccinated animals can prevent the transmissionof EAV. Since EAV-negative and properlyimmunized stallions cannot be carriers, everynegative foal under 9 months of age should bevaccinated. It is worth mentioning that the useof this vaccine in pregnant mares has not beenapproveddue to thepossibilityofcausingabortion(USDA-APHIS, 2004). In addition, vaccinationhas been successfully used as a tool to controlthe spread of the disease during an outbreak,controlling the spreadof infection and the severityof the symptoms (Timoney, 1988). It is necessaryto mention that due to EVA is an exotic disease,vaccinations are not recommended in our countryyet.

In breeders and sites dedicated to reproduction,it is necessary to performbleeding and serologicalassays of all horses prior tomating; viral isolationshouldalsobeattemptedonimportedsemenbeforeuse. It is very important tomaintain strict hygieneand disinfection of instruments and equipment tominimizetheriskoftransmissionofthevirus.EAV-negativemares shouldmateonlywith semen fromEAV-seronegative stallions, but not with semenfromcarriersthatcouldbeinfected.

If the results of blood tests are positive for astallion, but there is no official documentation ofa negative state prior to vaccination, the stallionshouldbeanalyzedtodetermineitspossiblecarrierstate. Virus isolation should be attempted usingsemen of two separate ejaculations, or matingtwo EAV-negative mares with the same stallion.Twenty-eightdaysaftermating,serologicalsurveysshould be performed on themares to determine ifneutralizingantibodiesagainstEAVhavedeveloped(Holyoaket al.,2008).

Carrier stallions should mate only with EAV-positive or properly vaccinated mares. When acarrierstallionispairedwithapositivemareorwitha vaccinated mare, both mares should be isolatedfor 24 hours after mating to prevent mechanicaltransmission of the virus through traces of semen.If it is thefirst time that themare has been paired

with a carrier stallion, themare should be isolatedfromotherhorsesfor21daysduetothepossibilityof virus dispersion trough traces of semen. Allvaccinated horses should receive yearly boosterstoprotect themagainst infectionand in thecaseofstallions, to prevent the development of the carrierstate. In a generation or two, these practices couldeliminatethepopulationofcarrierstallions(USDA-APHIS,2004).

Epidemiological status in America

In America, the presence of EAV has beenreportedindifferentcountriesofSouth,CentralandNorth America, mainly in Argentina, the UnitedStates and Canada where the virus has causedsubstantialeconomiclossesandistodayoneofthemainpathogensofveterinaryimportance(Figure3).

Figure 3. EAV distribution in America. In light grey, countries with positive serology. Dark grey, country with the most recent EAV outbreak. Cross-filled, EAV positive result by surveillance in borders, no cases reported in the country.

TheprevalenceofEAVvariessignificantlyamonghorsebreeds.IntheU.S.,thisinfectionisparticularlycommonamongStandardbreds. In a surveydone in1998, 24% of unvaccinated Standardbreds, 4.5%of Thoroughbreds, 0.6% of Quarter horses, and3.6% of Warmblood horses had antibodies to thisvirus. Breed-related differences in seroprevalencemight be due to genetic differences, but they aremore likely to be caused by different managementpractices (USDA-APHIS, 2004). In experimentallyinfected horses, the breed has no apparent effect



on susceptibility to infection or the establishmentof carriers (Neu et al., 1992). For LatinAmericancountriestherearenotanofficialreportofprevalencedue to there is only few studies andEAV serologicsurveyofhorse’spopulationsdoesnotexist.

InBrazil, infectionwasfirstreportedin1993inIbiúna,stateofSãoPaulo(Fernandeset al., 1997),and now prevalence rates have been reported,ranging from 0.85% in the state of Minas Gerais(Belloet al.,2007),to18.2%inSãoPaulo(Laraet al.,2002)and to23% inhorses in the stateofRioGrande do Sul (Diel et al., 2006 ), indicating thatthe level of infection can be high if no adequatecontrolmeasuresareadopted.

However, studies conducted in the provinceofUrara inBrazil and in serum samples of horsesbelonging to different breeds and uses in Peru didnot reveal seropositive animals (Rodríguez andRivera,1998,Heinemannet al.,2002).Ontheotherhand, theviruswasnever isolatedinChile,but theborder surveillance program detected the presenceof two positive animals (Figure 3), one of whichwas killed and the other returned to its country oforigin(Berrios,2005).

In 2005, using neutralization assays, out of1,008equineserumsamples takenfrom5differentstates of Venezuela, 2.48% seropositivity wasreported, with a significantly greater proportionin imported horses compared with horses born inthe country; this was the first laboratory evidencefor the presence of EVA in the northern regionof South America (Perozo, 2005), and highlightsthe importance of horse exchange on the borderbetweenVenezuelaandColombia.

ThecaseofArgentinehasbeenlargelydiscussedand documented in the scientific and technicalliterature, as it is the SouthAmerican country withthe largest number of reported cases. Until 1998,there were no reported cases of the disease inArgentina and the virus had never been isolated insamples from nasal swabs or abortions in horsessuffering from respiratoryor reproductivedisorders.Inparallelwiththeapprovalof theimportofhorsesfrom EVA-infected countries, Argentina initiatedserologicalmonitoringofimportedhorsesandsemen

as part of the EVA Epidemiological SurveillanceProgram;thisrevealedonepositivesampleofsemenimportedinApril1998,andledtothedestructionofthe entire lot of semen, and to official interventionof the two farms involved in the import and use ofthesemen.Inpairedsamplescarriedoutinallofthehorses existing in the two farms, no infection wasfound,andthisdeterminedtheriseofhealthactivitiesand restrictivemeasures in themovement of horses(movementrestriction)(DelaSotaet al.,2008).

In October 1998, of three stallions importedto Argentina, one was seropositive, so that thefarm was restricted and required sanitation. Inthis site and during the implementation of thesehealth activities a high serological prevalence wasdetectedandthetwootherstallionswereconfirmedby biological assays as virus reservoirs and wereconsequently castrated toprevent the spreadof theinfection(DelaSotaet al.,2008).

Therehavebeendifferentserologicaltests,totrytoassessthepresenceofinfectionindifferentareasandhorsepopulationsfromArgentina,mostofthemyielding negative results, such as in breeders thathad imported horses toArgentina since 1995, andalsoinhorsespresentinfarmsadjacent(inaradiusof 10 km) to the first farm in which the infectionwasfirstdetected(DelaSotaet al.,2008).

Threeyearlater,inOctober2001,therequirementto diagnose EVA in native, imported horses orseminal material was established. Out of the 1774samples included in this study, 99.83% were freeof infection; 14 stallionswerepositive forEVA.Ofthese,10werethoroughbredsimportedfromtheU.S.and one was aWelsh Pony imported from France;allofthemhadbeenvaccinated,thethreeremainingstallions were Show Jumping and Dressage horses,andwereremovedfromthereproductiveactivityandcastrated to prevent the reproductive spread of theinfection(Gonzálezet al.,2003).

InJune2002,twofarmsepidemiologicallyrelatedwith the initial outbreak of EVA were evaluatedby virus neutralization. Bleeding was performedonall thehorses inorder todetectpossiblecasesofseroconversion. Eight of the 439 horses evaluatedtested positive, corresponding to eight mares that



camefromthefarminwhichtheinitialoutbreakhadoccurred(Gonzálezet al.,2003).

As a result of positive serological diagnoses,interventionswereconductedinsixestablishments.Serological surveys were conducted in all thehorses, two serial diagnostic tests with an intervalof 14 days, to verify the absence of viral activity.Of a total of 1080 equines sampled in nine farms,306(30.6%)werepositive,indicatingthatoncetheinfection enters a herd it can easily be distributedwithinit.However,noneof thesymptomsreportedthe presence of reproductive or respiratory clinicalsigns consistent with those caused by EAVinfection. A consolidated analysis of the resultsobtained between 1998 and 2002 showed that of7722 samples tested, 319 were positive (4.13%).In addition, when the origin of the horses wasexanimated,thepresenceofinfectionwasrestrictedto three of 23 tested provinces, Buenos Aires,CordobaandEntreRios(DelaSotaet al.,2008).

From the epidemiological investigations carriedout, it was concluded that the possible entry ofthe virus that occurred through a stallion importedin 1996, could have spread the infection to othersusceptiblehorseson the farm.Additionally itwasconcluded that EAV has a restricted geographicdistributioninArgentina,beingpresentinjustafewprovinces,equinecategoriesandproductionsystemsdue mainly to high risk factors such as horseimportations(DelaSotaet al.,2008).

Phylogeneticanalysesofstrainsisolatedin2002fromsemensamplesofanimalsbelonging toherdswith a history of import of animals from Europeand from a testicle stored at -20 ºC since 2000,showed that the Argentinian strains of EAV havestrong phylogenetic relationships with Europeanstrainsandthatthereareatleasttwodifferentvirus“populations” circulating in Argentina (Echeverriaet al., 2007, Metz et al., 2008b). Other recentand larger studies identified two major groups ofsequences confirming that SouthAmerican strainshavemultipleorigins,oneof thembeingEuropeanstrains(Metzet al.,2008a,Metzet al.,2010).

Additionally, studies of viral neutralizationin vitro have shown that the strains isolated in

Argentina have different neutralization patternscompared with a reference strain, and that thereference strain used in diagnostic tests andvaccines, is neutralized to a lower lever byheterologous antisera (Echeverría et al., 2010).This result is very important if one considersthat American commercial vaccines use strainswhichmay not generate adequate immunity in ourcountries.

EVA multi-state outbreak in U.S.

As in Latin America, in U.S. EVA has causesignificant economic loses; in the last years astrong multi-state outbreak reaches 18 states. Thisoutbreak, began with an outbreak of abortions inearly June 2006 in a Quarter Horse ranch in thestate of New Mexico where one of four stallionsdevelopedfever,depression,preputialandforeheadlimb edemas, with subsequent loss of fertility andvirus isolationfromsemen.Subsequently,asecondstalliondevelopedpyrexia,butmaintainedadequatefertility. Prior to the diagnosis of EAV, semenhadbeen sent todifferent states in theUS, that allreportedpositiveforEAVaftershipmentanduse.Inthe samebreeder farm, the two remaining stallionswere confirmed as positive, but only one showedlightpyrexiaandbothexcretedvirus in thesemen.Intotalandassociatedwiththefirstcase,therewere30 reported abortions, representing about half thepopulation of pregnant mares exposed to the firstinfectedstallion(Timoneyet al.,2006).

Additionally, due to shipping of semen ormovement of mares, the outbreak spread to 18states: 69.5% corresponded to those infectedthrough direct exposure to insemination withinfected semen, 29% corresponded to mares andfoals that were exposed indirectly by visitinginfectedherdsduringtheoutbreakand1.5%inbothinsemination and indirect exposure. Moreover, insome states such asUtah, the outbreak resulted inrespiratory illness, abortions, neonatal pneumoniaand death of several animals. It should be notedthat initial studies of the outbreak concludedthat the infection reached approximately 90%seroprevalence in the infectedfarms, indicating theefficiency of spread of infection in a susceptiblepopulation(Timoneyet al.,2006).



Recent phylogenetic analyses performed onsamples of the same outbreak, confirmed that thenationalcirculationofthevirusoriginatedfromthedispersion of the original strain fromNewMéxicoand later that the virus mutated and changed itsneutralizing phenotype, demonstrating the strongcapacityofthevirustoevolveandadaptthroughthepersistenceofthevirusincarrierhorsesandthroughthe passage fromone animal to another during theoutbreak(Zhanget al.,2010)

Is Colombia at risk?

Nowadays,Colombiahasactive import linksofhorses with countries in which EAV is present orhas been reported, and has imported many horsesfrom those countries. Examples of this are themassive import fromArgentina of more than 700CreoleArgentinehorsesundertakenbytheNationalPolice in 1997, or the Paso FinoWorld Cups thathave taken place inMedellín in 2003 and Pereirain2009, using thefigureof “temporary import”ofanimalsfortheinternationalcompetitors.

Theimportofhorsesisaveryimportantelementof Colombia’s livestock trade. Only during 2005and2006,1079horseswereimportedfromMexico,Panama,Perú,Portugal,PuertoRico,theDominicanRepublic, Guatemala, Honduras, Venezuela,Argentina, Aruba, Bolivia, Chile, Costa Rica,Ecuador, USA, France, Italy and the Netherlandswith a value of USD 1.200.000. This is not tomention the importance of exports which duringthesameperiodwasoverUSD22.000.000(Source:ColombianAgriculturalInstitute,ICA).

According to a 2008 equine census, publishedby ICA, Colombia currently has a population of1.882.730horses settledmainly in thedepartmentsof Cordoba, Antioquia, Casanare, Tolima andCundinamarca, where it concentrates 42% ofthe national equine census. Given these figures,an infection could clearly be devastating to oursusceptiblepopulationsandtheriskistoohightobeignored.

Among the animal health requirements forpermanent entrance of horses into Colombia, ICArequires that stallions be negative after two viral

neutralization tests carried out during the 28 daysprior to shipment and a 14 day interval betweenthem; for seropositive stallions, the requirement isthattheyhavematedtwoseronegativemareswhichremained negative after two neutralization tests,carried out first, the day ofmating and second 28dayslater.Forfemalesanegativeresultorstabilityof neutralizing titers in two serological, evaluated28daysprior toshipment isrequired.Additionally,import is also allowed of geldings that werenegativebetween theagesof6and12monthsandwerethenvaccinated.

However“illegal”entryofequinesacrossborderssuchastheVenezuelanborder,foreventsthatgrouppopulationssuchasfairsorforuseastransport,canrepresentastrongriskofdiseaseintroductionintothecountry. This means that infection can be acquiredvia European strains imported to the Americasfrom South America, or via strains causing strongoutbreaksinthenorthernhemisphere.

ThebackgroundintheLatinAmericancountriespresented above highlights the importance ofserological surveillance, which informs us that anewagenthascomeincontactwiththesusceptiblepopulation of horses, and allows us to establishpreventivemeasurestoavoidspreadofthevirus.

It is noteworthy that the ICA has recentlytemporarilysuspendedtheentryintoColombiafromhorses from Argentina as a result of the nationalhealth warning issued by the National HealthService and Food Quality-SENASA following apositivediagnosisofEVAintheprovinceofBuenosAires(ICA,2010).

Therefore we propose to strengthen technical-scientific investigations and mainly diagnosticcapacities to be prepared for early diagnosisand control, and avoid an infection that can bedevastatingforsuchalargeandimportantindustryaszootechnicalexploitationofequinesinourcountry.

Acknowledgment

The author thanks Dr. Anne-Lise Haenni forrevision of English and critical reading of themanuscript.



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