---start--- parasit 10/28 dr hunter Toxoplasmosis (handout) and neospora toxoplasma gondii: a coccidian parasite which has a sexual cycle in the definitive host (cat, felids) and asexual cycle in int host (other mammals, birds) as well as in the cat. Toxoplasma and neospora: -understand the complex nature of the parasite life cycles -be able to tell the general features of disease transmission apart for the two parasites -understand the basis of dz -congenital vs reactivation these are similar parasites. can cause congenital or reaquired dz. t.gondii: infects any cell type complex life cycle sexual cycle in felids - schizogeny in epi cells of the gut, gives rise to infectious oocysts asexual cycle in other mammals and in cats - tachyzoite to bradyzoite transformation, important part of pathogenesis. tachyzoite- fast replicating form of parasite bradyzoite - forms latent cysts in many tissues, keeps infectino in chronic form. life cycle of toxoplasma: cat is definitive host. cat eats oocyst. schizogeny occurs in gut. cat passes oocysts. when oocysts first appear they are unsporulated. it takes 2-3 days for oocyst to sporulate and become infective. infective oocysts found in 2-3 day old cat feces. source of environmental contamination. if these oocysts are ingested by man, mouse,etc will enter asexual stage. oocysts become stage that invade epi cells. tachyzoites replicate within cells, lyse cells, disseminate through body. tachyzoites disseminate through body. hosts develop protective immune response. we see appearance of bradyzoite stage found in tissue cysts which persist in the host. once infected, you are infected for life. 10-30% of people in US are infected and have these tissue cysts. tissue cysts, if ingested by another host, are also infected. if cat eats infected mouse, it will become infected. stages we need to know: tachyzoite, bradyzoite, oocyst. if cat eats mouse bradyzoite leaves cyst, and can form new tissue cysts, or can give rise to tachyzoites, undergo schizogony, and form oocysts. slide: human fibroblast with toxoplasma tachyzoites within it. each rosette is a group arising from one tachyzoite. realize that tachyzoites can infect ANY nucleated cells. you name it, it can be infected. tachyzoites invade, replicate, fill cell, lyse cell, get released, infect new cells -yada yada- lather, rinse, repeat. this leads to widespread dissemination of parasite if immune response is lacking - will get large areas of necrosis in absence of immune response. immune response is critical to control the tachyzoite. slide: tissue cyst in mouse brain. within the cyst are thousands of bradyzoites. tachyzoite has invaded cell and instead of replicating has formed cyst - wall formed of parasite derived molecules and host material. tachyzoite forms bradyzoite - and makes many bradyzoites. mostly this occurs in brain, skeletal muscle, and heart. essentially, brain is immunocompromised. there are no T cells in the brain. the tissue cysts are long lived. they keep host chronically infected. they can spontaneously reactivate, parasites leave cysts, invade cells, and replicate. w/o immune response, parasite will go unchecked and cause disease, but usually immune response kicks in. infection occurs via ingestion of oocysts (which can survive over a year - highly infective, and pathogenic.) contaminated vegetables, water etc. there was an outbreak of water borne infection in vancouver a few years ago. or, can occur via ingestion of tissue cysts - most common method for humans. these tissue cysts occur in muscle. beef, pork, lamb, venison are all sources of contamination for humans. pork in USA - 23-42% of hogs in US seropositive in 1986. so do NOT eat undercooked pork. when oocysts first come out in cat, they are unsporulated and noninfective. in 2-3 days will sporulate, then contain infective sporozoites within them. these are about a third the size of an isospora cyst! there are many coccidians of cats. toxoplasma oocysts are much smaller than isospora felis. h.hammondi however is very similar looking to toxoplasma. so seeing coccidian cyst in cat feces isn't diagnostic. h.hammondi is not pathogenic. life cycle of toxoplasma wasn't known until the 1960s. cat wasn't known as host. when cat was identified as source of oocysts, people got really upset about it. many women's magazines had articles about what to do with cats if you're pregnant. cats normally get the infection through mice. also can get it through raw meat. the cat mouse life cycle is a natural life cycle. toxo dz in cats: in most cases this is an asymptomatic infection, in most hosts. however, in cats under a year old you can see acute dz not uncommonly. is dx'd by persistent high fever, unresponsive to abx. within 2 wks of infxn, cat sheds cysts for 1-3 weeks. so the cat only sheds oocysts for 1-3 wks of its life. oocyst shedding can reoccur though following marked stress, immunosuppression, or superinfection with other coccidians like isospora. these are exceptions, however, in general, cats only shed onece, then get protective immunity that deals with parasite, prevents reinfection. pathology can occur during acute dz, and in immunosuppressed cats that reactivate (associated with FIV) slide: lung of cat with toxoplasmosis - lungs are full of blood with white patches where the parasites are replicating. slide: bronchial wash from infected cat - mononuclear cells stuffed full of parasites. these parasites replicate in macrophages as well. pancrease, liver, lungs, brain can all be affected like this. in cats the funny thing about it is there are cases where acute dz occurs in mature cats that are not immune suppressed - but this is unusual. normally it's asymptomatic. vertical transmission is pathogenic for sheep and humans. this is transmission from mom to fetus. in sheep, it is an important cause of abortion and perinatal mortality. you don't see acute dz in adult sheep. but if ewe becomes infected while pregnant, vertical transmission can occur. during early gestation, can lead to death and expulsion of fetus. often undetected if occurs in field. during midgestation more readily detected. will see stillborns, mummified fetus, typical white lesions in placenta. ewes generally lamb normally in ensuing years. in a flock, you may see "abortion storms" - wild infected cat sheds oocysts on the pasture - large numbers of sheep are infected during gestation - 30-50% can abort (assuming previously uninfected herd). when parasite invades ewe for first time, disseminates through body, multiplies in placenta, crosses into fetus which has no immune system yet. so parasite can go unchecked, and kill the fetus. in humans: several forms of disease congenital disease: abortion or sequelae in newborns acute toxo: rare but when occurs can be severe. can kill immunocompetent individuals. thought some strains are more virulent. most people who get infected show no lcinical signs. 10% can get acute toxo - which can be misdiagnosed as mononucleosis, severe flu, hodgins dz - fever, lymphadenopathy. ocular dz: rare, can be consequence of acquired or congenital dz loss of T cell function - very important predisposing factor for reactivation of dz- reactivation of tissue cysts - so, since early 80's and emergence of AIDS, toxo has been more of a problem. also cancer patients and transplant patients are at risk for reactivation of preexisting toxoplasmosis. in one case, an uninfected human got a heart from a seropositive donor - and developed toxoplasmosis disease because cysts in heart were reactivated. eye is another immunocompromised site like the brain. you may see loss of retinal cells "ketchup and mayonnaise" type lesion. toxo in pregnancy: in humans: early infection is more severe (abortion, hydrocephalus, blindness) 85% of infected infants are NORMAL at birth, however. It is important to knwo that they are infected, because if not treated, will see development of chorioretinitis, hearing loss, and developmental delays. if a woman is infected for the first time while pregnant, there is a good chance of it affecting the fetus. but if they diagnose the new infection, they can treat the problem. mothers infected during first trimester and left untreated - 15% of infants got disease. 30% in second and 60% in third. if mothers were treated, dz incidence dropped remarkably. serodiagnosis of T.gondii: 10-30% of US population, 40-80% of western europe. IgM is associated with acute dz, recently acquired IgG is associated with chronic dz IgM titer can persist for over a year, so serial testing is required. all seropositive people are at risk for reactivation if they become immunosuppressed. if a woman is suspected of having contracted toxo and she has high IgG level, it's a chronic problem. but if she has IgM, it could have been anytime in the past year. So you test again, to see if it is increasing or decreasing. increasing is associated with a more recently acquired infection. with recent/active infections, titers are rising. dx and tx: in cat, b/c cat only sheds 1-3 wks, oocyst detection is rare. can use serology to detect acute vs chronic infections. serology useful to detect naive status. tx rarely warranted. vaccination: anti-abortion: S48 strain has lost ability to form cysts and is used in NZ routinely to immunize sheep. cysts are the stage that's infective to the next host to eat the sheep. so you won't get congenital dz either. anti-transmission: T263 does form tissue cysts but not oocysts. infection of kittens results in mild dz but no oocysts. challenge infection led to no oocysts in 84% of kittens. company thinks we should use this to infect cats to reduce environmental contamination control: cook meat properly. freeze meat. sanitation. don't feed raw meat to anyone or anything. pregnant women should not handle cat litterpans (although risk is really minimal if it's changed daily...since it takes 2-3 days to sporulate). change pans daily and wash with boiling water. 10% ammonia will kill oocysts but clorox won't. pigs are a big source of tissue cysts in america. pigs don't just get infected by cat feces containing oocysts. they also eat small furry rodents. cows do not eat cats, but they can be near cats and can get oocysts from feces. for many years, people thought toxoplasma caused abortion in cattle, but it turned out to be neospora caninum. neospora caninum is important in dogs and cattle. toxoplasma like organism - 10-23% of dogs formerly thought to have t.gondii actually had neospora caninum. Lifecycle unknown, but cysts are resistant to pepsin-HCl. transmission can occur congenitally or by carnivorism. important cause of congenital dz in dogs and cattle. we don't have to be able to tell these apart. they look similar even at EM level. looking at tissue cysts...toxoplasma cysts and neospora cysts are very similar, but the wall is much more well defined and thick with neospora cysts. dx of neospora caninum: serology: IFA, ELISA. postmortem: immunohistochemistry, isolation of parasite in tissue culture or mice. puppy with ascending hindlimb paralysis PA 1987 4 lab puppies got asymmetric paraparesis at 4 wks. 3 progressed to death, last one was euthed. necropsy revealed meningoencephalitis and neospora. bitch was rebred and had 7 normal pups - but at 5-6 weeks, all pups developed hind limb paralysis. 3 killed by owner,3 by vet, and one lived with minimal ataxia. so this parasite exhibits repeated congenital transmission unlike neospora. a second bitch in household gave birth to a litter in which 3 of 7 were also affected by neospora caninum. calfs also have HL paralysis. histo of brain shows loss of cellularity. the parasite has the same stages as toxo - is very destructive to tissue. you see areas of necrosis. first reported in cattle in 1989 when a herd in NM had persistent abortions and no toxo was found. dx made on immunohistochemistry. in cattle, abortion is only clinical sign in adults. few cases of clinical dz - subclinical congenital dz more common. estimations of abortion are 5-50% of all bovine abortions. can also see abortion storms. repeat abortions can occur but at less than 5% rate. less common than in dogs. in CA about 20,000 cattle were checked - 266 abortions occured, 43% were neospora related and 6% were suspected to be neospora - 14% bacteria/mycosis, and 37% other. in PA, about 5% of bovine abortions. other animals - one report of congenital dz in sheep. can infect sheep experimentally. goats - congenital dz horse - rare but has occured cats- no report of natural dz, experimental dz will occur coyotes,deer, monkeys, foxes. does it infect humans? dunno. not sure. retrospectives are being done. ---break---- 3-4 dr farrell parasit SARCOCYSTOSIS sarcocystis are within the apicomplexa along with crypto, isospora,and toxo. if you do a fecal on dogs or cats you will find potentially a number of coccidia. in fact, what you see in feces with sarcocystis is not a fully differentiated oocyst, but a sporocyst containing sporozoites. the organism makes an oocyst, but oocyst wall is thin, it breaks down, maturation is rapid, so in teh feces are the sporocysts which have broekn out of oocyst, and sporocysts are fully differentiated and infective. many spp in feces - s.suicanis in dogs, s. hirsuta, s.leporum and others in cats, others in dogs too. life cycle: requires two hosts. one is a carnivore (dog) and another is a herbivore (cow). there are multiple carnivorious hosts - cats, humans, pigs. also herbivores - cows, sheep, goats, etc can all be intermediate hosts. carnivore is definitive host. within GI Tract of carnivore you have sexual cycle - male and female gametocytes form, oocysts form, sporocysts pass in feces, sporocysts containing four sporozoites each are eaten by herbivore, in intestinal tract sporocyst wall breaks down, sporozoites penetrate into intestinal tissue, into lamina propria, enter into blood vessels. once there, enter into endothelial cells. first cell affected are endothelial cells of large blood vessels. the organism then undergoes schizogony in the endo cells of large vessels. after a period of division they break out into blood stream, reinvade cells generally of very small vessels, capillaries, the endothelial cells of them, and divide again - then when organism breaks out again, it enters WBCs and may undergo a third cycle of division - this is unclear. but shortly after this,it can enter muscle cells and infect them as toxoplasma would, and we have a slowing of the rate of division of organism, to form bradyzoite like cysts. so early rapid division is equivalent to tachyzoite stage, adn then cysts are like bradyzoite stage, fairly quiescent within tissue. these persist for extended periods of time, years if cow lives that long. these cysts are infective for the carnivore. a dog eating meat from an infected cow would ingest bradyzoites in tissue cysts. instead of undergoing cycles of schizogony, in the dog, gametes immediately form. there's no asexual cycle. you get fertilization, production of oocysts, and excretion of sporocyts. only sexual cycles occur in carnivore hosts. only asexual w/in herbivore hosts. small sporocyst seen on fecal float - can see sporozoites within it. there's no maturation outside the host. are infective when passed. the bradyzoite like stages, the sarcocysts, in int hosts, can be very large. can see the fully matured cyst containing the sarcocysts or bradyzoites - just huge compared to say, trichinella. these are visible to naked eye. after sporozoites invade endothelial cells in large vessels and divide, there is some pathology associated with this - generally if animal eats enough it will get fever, general lethargy, and some mild inflammation. at the same time, we're amplifying number of parasites. the second time they enter endothelium this time of capillaries, you have more pathology - marked inflammatory response. this occurs about 4 weeks after infection. animals may have fevers, anemia, and more. then organisms may enter WBC - then they will eventually invade muscle cells. young maturing sarcocyst with developing bradyzoites seen on slide -- similar to toxoplasma bradyzoite cysts in that not much inflammation occurs. they are quiescent, don't elicit immune response. just sit andwait to be eaten by carnivore. no dz associated with sarcocysts. if animal ingests a very very heavy dose of sporocysts, it can become clinically ill. 40 yrs ago in canada, an outbreak occured where many animals died. cows stopped making milk. abortions occured. there was massive internal damage to multiple organ systems due to growth of these things in capillaries. petechial hemorrhages seen all over. so, any capillary bed was vulnerable to this. any capillary bed can be damaged by this. it's basically rupturing large numbers of endothelial cells which lead to the hemorrhages. animals can die of the infection if severe enough. animals can be stillborn. calves/lambs can be stillborn. but generally signs are not severe or are absent. first generation of schizogony in lg vessels - slight fever, maybe. 2nd generation of schizogony in cap beds - fever, anemia, may see other signs eg anorexia, wt loss, decreased milk, hair loss, abortion, or even death. most however recover. and most don't show any signs. there ahas been a syndrome which is called chronic sarcocystosis. these animals have continued inflammatory responses in their muscles. not sure of why this happens. people at USDA estimate that 90% or more of cattle in US are infected with one or more spp of sarcocystis. unless animals ingest a really high number of sporocysts, they probably won't show any signs at all. theremay be subclinical disease - failure to gain wt as much as they wouldhave, failure to reach potential - but we can't attribute this directly to sarcocystis. disease outbreaks are rare. when that does occur, that's usually associated with 2nd generation of schizonts, and local hemorrhaging in capillary beds. so, why is this of any importance? there are occasional outbreaks of infection. there are economic losses due to meat condemnation - if you see the sarcocysts, the meat is condemned. most sarcocysts are probably microscopic though, so most infected animals probably pass inspection. the large visible sarcocysts are rare. so we have life cycles that go from cow to dog. other species infect cat and cow, others dog and sheep, others dog and horse, multiple spp. probably 100 or more sarcocystis spp using diff definitive/int host combinations. speciation isn't that clear, actually. no known dz in carnivore host. there is an important sarcocystis in horses. the disease is generally called equine protozoal myeloencephalitis. 30 yrs ago it was observed that horses got a clinical syndrome of hind end paralysis, altered gait, neuro signs. some of these horses had small toxo like organisms found - and in past ten years or so we found out these are sarcocystis. called it s. neurona, b/c it produces neuro dz. about a year and a half ago they figured out life cycle - in fact, it was already known - s. somethingelse - possum/bird life cycle. it's in cow birds and other birds in bradyzoite stage. possum is definitive host. possum sheds sporocysts which are infective to birds. horses are accidental hosts. they are a dead end host. can show severe pathology which isn't normally seen in birds, other definitive hosts. hind end paralysis, unilateral problem usually. infected horses with clinical dz are very resistant to backing up. they have problems primarily with hind limbs and prefer to remain balanced and don't want to back up. may also show loss of muscle tone, exhibited in face, usually unilateral, or tongue - may see altered structure due to loss of tone on one side. infection may be fatal to horse. most sarcocystis spp infect intermediate host, invade lamina propria, blood vessels, multiply in endothelial cells, and eventually encyst. but we aren't sure what happens in the equine host. we know animals are infected, organisms probably disseminate through blood, may infect multiple cell types we aren't aware of...but if it invades CNS it causes severe dz. lesions are seen in brain - unilateral swelling, edema in brain. also in cervical or thoracic spinal cord. usually unilateral lesions, leading to asymmetrical clinical signs. paralysis of one HL, loss of tone on one side, etc. ability of aberrant parasite in horse to invade into neural tissue probably as tachyzoite like stage and cause focal lesions leads to the dz. at NBC there may be from 20-40 cases/yr. we're not sure. very hard to dx. few antemortem diagnostic tests. usually dx is based on clinical signs. those signs generally relate to some kind of unilateral limb paralysis or loss of tone. in kentucky, probably 10-15% of neuro admissions in horses are due to sarcocystis infections. we don't have evidence of that here. there are diagnostic tests to use - not done here. basically this involves detecting antibody for sarcocystis using PCR or western blot from CNS fluid. serology using serum is not effective b/c many animals have antibody but no disease. people are thinking maybe in many areas even around here, up to 50% of horses may be exposed and have circulating Ab and be seropositive, with no dz. but animals wouldn't have Ab in CNS fluid unless there is infection there, so you have to look there either via PCR for organism, or by western blot for Ab. generally, animals coming in showing these signs are immediately treated for presumed sarcocystosis. tx is longterm, up to twelve weeks. it's expensive - $500-1000 per animal, and usually based just on signs and presumptive dx, rather than real dx. problem is if you wait to tx til you have dx, you will risk severe permanent damage. if you tx early, you may prevent permanent damage. often NSAIDs are given as well. control is very difficult - keep birds out of stalls, barns, etc. don't have nests within barns. can try to fence areas against possums but that is really hard. in terms of other effects - don't feed raw meat from domestic animals to anything carnivorous. don't feed sheep to dogs, etc. that's just perpetuation of the life cycle. don't keep cats/dogs in barns where they can defecate near herbivore food sources. ---end----