Zoonosis Caused by Baylisascaris procyonis

The raccoon roundworm, Baylisascaris procyonis, is classified under the Phylum Nemathelminthes (the roundworms) and Class Nematoda. It is a member of Family Ascaridae and Superfamily Ascaridoidea, which represents intestinal worms with direct life cycles. Other, more familiar ascarids are Ascaris lumbricoides,Toxocara canis, and Toxocara cati, nematode parasites of humans, dogs, and cats, respectively.

Adult female worms in the small intestine of raccoons and produce between 115,000 and 179,000 eggs/worm/day. In nature, infected raccoons shed an average of 20,000 to 26,000 eggs per gram of feces, with higher shedding rates in juvenile raccoons than in adults, and can shed in excess of 250,000 eggs per gram of feces. Thus, infected raccoons can shed millions of B. procyonis eggs daily, leading to widespread and heavy environmental contamination. The numbers of eggs produced by infected raccoons combined with their defecation behaviour, ensures that latrine sites will become heavily contaminated. The eggs possess a sticky proteinaceous outer coat that enables them to adhere to objects and facilitates transmission. B. procyonis eggs become infective (secondstagelarva) in ∼2-4 weeks, depending on environmental temperature and moisture. B. procyonis eggs are very resistant to environmental conditions, especially in moist soil. Although they can be killed eventually by extreme heat and dryness, the eggs survive harsh winters, and under appropriate conditions, they can remain viable for years, contributing to the long-term danger posed by latrines.
Young raccoons become infected by ingesting infective eggs, whereas older raccoons become infected byingesting third-stage larvae (L3's) in paratecnic hosts, usually rodents. Young raccoons become infected at an early age by ingesting eggs fromtheir mother's contaminated teats or fur, from the contaminated den, or from raccoon latrines near their den.
In young raccoons, larvae hatching from eggs enter themucosa of the small intestine and develop there several weeks before reentering the intestinal lumen to mature, the worms reaching patency in 50-76 days. In older raccoons, larvae from intermediate hosts develop to adults in the intestinal lumen, reaching patency in 32-38 days.
The higherparasite burden of juvenile raccoons (mean burden, 48 to 62 worms) than in adults (mean burden, 12 to 22 worms) likely reflects differences in mechanisms of infection.
Like other parasite (Toxocara spp.) B. procyonis can be borne by intermediate hosts when eggs are swallowed by a different vertebrate. Most commonly the intermediate host is a rodent, birds or lagomorph. In these animals, parasite eggs hatch in the small intestine, penetrate the intestinal wall and are get to the bloodstream through the liver to the lungs, where they are eventually distributed via the blood to various organs. Larvae eventually become encapsulated within eosinophilic granulomas, where they remain viable until they are ingested by raccoonsor for the lifetime of the host (Kazacos& . B. procyonis larvae often invading the central nevous system (CNS). Invasion of the brain seems to be particularly common in rodents, rabbits, birds and primates. In mice, approximately 5-7% of B. procyonis larvae are estimated to enter the CNS. These larvae can www.intechopen.com cause considerable damage, both from mechanical damage during migration and from the inflammatory reaction they stimulate. Larvae in the eye can damage the retina and other structures. Eventually the larvae encyst, mainly in the connective tissues and muscles.  Humans are accidental intermediate hosts and infection typically occurs in young children with pica or geophagia after ingestion of infective B. procyonis eggs from environments or items contaminated with raccoon feces (Kazacos, 2000).
In humans, B. procyonis larvae have a tendency to invade the eyes, spinal cord, and brain, causing inflammatory reactions and tissue damage. The result can be blindness, neurological damage, and even death. B. procyonis infection typically results in fatal disease or severe sequelae.

Epidemiology
Raccoons, which are the definitive hosts for B. procyonis, are native to the Americas, where they can be found from Canada to Panama. They were introduced into Europe, the former U.S.S.R. and Asia for the commercial fur trade and into Japan as pets, and have become naturalized in some of these areas. The prevalence of B. procyonis infection is high in wild raccoons in Germany and those kept in zoos or as pets in Japan (Baeur &Gey, 2002;Kazacos, 2001;Miyashita,1993;Sato et al., 2001). Although surveys of feral raccoons in Japan have not detected this organism, it is possible that some pets released into the wild were infected.
In United State B. procyonis roundworms are most prevalent in the midwestern, northeastern, and Pacific western states. Numerous surveillance studies have been conducted in the southeastern United States, and parasite are most common in the mountainous regions of Virginia, Kentucky, and West Virginia (Kazacos, 2001;Souza et al., 2009). Geographic expansion of B. procyonis roundwormshas been recently documented in Georgia (Eberhard et al., 2003;Blizzard et al., 2010a) and into northwestern and southeastern of Florida (Blizzard et al., 2010b).Recently, and study about the prevalence of B. procyonis in raccoons in Portland, Oregon, showed that 58% of sampled raccoons were found to be infected with parasite (Yeitz et al., 2009).
In Canada, the prevalence of B. procyonis was estimated in 37.1% of the urban raccoon population of Winnipeg (Manitoba) (Sexsmith et al., 2009).
In most areas where raccoons occur, there should be no environmental limitationson the presence of B. procyonis, although conditions for optimal egg development and survival willvary based on temperature and humidity. Baylisascaris procyonis eggs become infective in 11-14 days at 22°C-25° C and 100% humidity (Sakla et al. 1989), similar to eggs of B. columnaris (11-16 d) (Berry, 1985). Under natural conditions, with cooler and/or fluctuating temperatures, egg development will beslower and will take several weeks to months.
Under sufficiently warm but fluctuating temperaturas (e.g., cooler nights), most eggs should reach infectivity in 3-4+ weeks. Embryonated B. procyonis eggs stored 9-12 years at4° C retained their infectivity and central nervous system pathogenicity for mice (Kazacos, 2001). Given adequate moisture, embryonated eggs will last years in the soil, including through harsh winters (Kazacos 1986(Kazacos , 1991.Conditions of extreme heat and dryness, as occur inbarn lofts and attics in summer months, will kill B. procyoniseggs by desiccation, probably in a few weeks ormonths ).
The epidemiology of Baylisascaris infection is linked to the defecation habits of raccoons. Presumably for communication or territorial reasons, individuals and groups of raccoons habitually defecate in focal areas called "latrines", where large amounts of feces and B. procyonis eggs accumulate (Kazacos, 2001;Roussere et al., 2003;Page et al., 1998;Page et al., 1999). Raccoon latrines are found directly on the ground, particularly at the base of trees; along and on the tops of fences; on roofs, decks, and stored firewood; and in outbuildings, attics, and various other locations (Kazacos, 2001;Roussere et al., 2003;Page et al., 1998). Homeowners are often unaware that there are latrines on roofs or hidden elsewhere on their property, thus increasing the risk of exposure to raccoon feces. Moreover, decomposition of the feces can occur rapidly under outdoor conditions, making it less obvious that these areas are contaminated.
The primary risk factors for human B. procyonis infection include contact with raccoon latrines, pica/geophagia, young age (<4 years), and male sex. Older persons who have pica or exhibit geophagia are alsoat risk for significant infection. People are commonly exposed to the eggsof this parasite in peridomestic areas where infected raccoons are common.

Infections in humans
Baylisascaris procyonis causes neurologic disease in wild, zoo, and domestic animals as well as human beings. The full clinical spectrum of human baylisascariasis is unknown but includes VLM, NLM, and OLM. In addition, preliminary evidence suggests that asymptomatic infection also occurs.
The severity of CNS disease is related to the number of eggs ingested, the extent and location of larval migration, and the severity of ensuing inflammation and necrosis (Kazacos 2000(Kazacos , 2001.When infective B. procyonis eggs are ingested, infective larvae emerge from the eggs, penetrate the gut, and after migrating through the liver and lungs, become distributed via the bloodstream to various somatic tissues, including skeletal muscles, the viscera, brain, and eyes; here, they continue to migrate and eventually become encapsulated in granulomas (Kazacos, 1997(Kazacos, , 2001. The pathogenicity of the larvae is related to their aggressive migratory behavior in the tissues and the fact that they molt and grow considerably during migration (Kazacos, 1997(Kazacos, , 2000(Kazacos, , 2001Goldberg et al., 1993). Only 5%-7% of ingested larvae enter the CNS (Kazacos, 2000(Kazacos, , 2001; thus, although theyare not neurotropic per se, their large size, aggressive migration, and stimulation of intense eosinophilic inflammatory reactions cause extensive damage to nervous (and ocular) tissues (Kazacos, 1997). Baylisascaris larvae entering the brain migrate there for extended periods before becoming walled off by host reactions.In heavy infections, the brain undergoes postinflammatory atrophy, leading to the progressive neurologic impairment and severe incapacitation seen in surviving patients with neural LM (Gavin et al., 2002a;Rowley et al., 2000a). Pathologic changes are further exacerbated by diagnostic and treatment delays.
The incubation period in humans is uncertain, but NLM may occur as soon as 2 to 4 weeks after ingestion of the eggs.
www.intechopen.com Zoonosis 290 NLM occurs when the parasites migrate through the CNS, and the symptoms vary with the location and number of the migrating larvae. The initial signs may be mild, with subtle behavioral changes, lethargy, somnolence or irritability, weakness, speech defects and/or mild changes in vision, but they can rapidly become severe. A variety of symptoms including ataxia, paresis or paralysis, developmental regression, tremors, torticollis, nystagmus and coma have been reported. Seizures are common and can be severe. Ocular signs, including blindness, also occur in many cases.
NLM is associated with eosinophilic meningoencephalitis, an elevated peripheral cerebrospinal fluid eosinophilia can be detected in cases of meningoencephalitis.
Some cases of NLM are fatal, almost all surviving patients have been left with serious neurological defects despite treatment. It has been reported only a one case of a child who developed relatively mild symptoms (headache, right arm pain, vomiting, mild upper extremity tremors and dysmetria, progressing to ataxia) and appeared to recover completely.
OLM has been reported more frequently than neural larva migrans, and can occur without neurological signs. Inflammatory and degenerative changes are mainly seen in the retina and optic disk, usually only in one eye. The clinical signs may include transient obscuration of the vision, photophobia, other signs of diffuse unilateral subacute neuroretinitis (DUSN) and loss of vision. Some visual defects can be permanent.
VLM by B. procyonis showed non-specific signs such as low-grade fever, nausea and lethargy. Invasion of the liver can result in hepatomegaly, and migration through the lung may cause symptoms of pneumonitis. A macular rash, seen mainly on the face and trunk, has also been reported. VLM is associated with eosinophilic cardiac pseudotumors (cardiac myofibroblastic tumors with high percentage of eosinophils). Subclinical cases might also occur in infection with B. procyonis.
Human infections with B. procyonis have been documented most often in the U.S., but suspected cases have been reported from Europe, and a patient with neural larva migrans was reported in Canada in 2009. One case of ocular larva migrans in Brazil was reported as a probable B. procyonis infection, but the identification was not definitive. No exposure to raccoons was documented in the latter case, although the patient had been exposed to skunks.
Other Baylisascaris species have been less well studied, but probably occur in most areas where their definitive hosts are found.

Infections in animals
Susceptibility to Baylisascaris larva migrans varies among animal groups and species (Wirtz 1982;Sheppard & Kazacos 1997). Animal groups particularly susceptible to parasite NLM include rodents, rabbits, primates, and birds, based on the number of cases and species affected. For example, in 2007 a Baylisascaris procyonis infection in a Moluccan cockatoo (Cacatua moluccensis), was reported. An adult female Moluccan cockatoo was evaluated for a 10-day history of progressive ataxia and weakness. The bird had been exposed intermittently over a 3-day period to a cage that had previously housed juvenile raccoons. Results of diagnostic tests were inconclusive and, despite supportive care, the bird died 7 days after the initial presentation. Histopathologic examination revealed a single nematode larva in the midbrain that was consistent with Baylisascaris species and multifocal granulomas in the left ventricle of the heart (Wolf et al., 2007).
Some animal groups and species are only marginally susceptible, with limited migration occurring in the intestinal wall or viscera; others appear to be resistant. For example, no cases of B. procyonis NLM have been documented in opossums, which are commonly exposed through foraging at raccoon latrines (Page et al., 1998(Page et al., , 1999, or in adult domestic livestockor zoo hoofstock, which are commonly exposed through contaminated hay. Very limited or no migration was seen in sheep, goats, and swine experimentally infected with B. procyonis (Dubey, 1982;Snyder, 1983;. No cases have been documented in cats or raptors, which eat rodents possibly contaminated with eggs and/or containing L3's. (Kazacos, 2001). The apparent species limitations to Baylisascaris infection should be regarded with caution.
Except in very heavy infections with intestinal obstruction, raccoons infected with B. procyonis appear clinically normal with no outward signs of infection. Similarly, other species with Baylisascaris larva migrans usually are asymptomatic if no larvae enter the brain. The severity and progression of central nervous system disease in NLM depends on the number of eggs ingested, the number of larvae entering the brain, the location and extent of migration damage and inflammation in the brain, and the size of the brain.Thus, clinical disease will vary from mild, insidious, slowly progressive central nervous system disease with subtle clinical signs to acute, fulminating, rapidly progressive central nervous system disease with marked clinical signs. Although larvae enter the somatic tissues, eyes, and brain of some species as early as 3 days postinfection, clinical central nervous system disease is not usually apparent before 9-10 days postinfection, and in many cases not until 2-4+ weeks postinfection, due to the lag time in causing central nervous system damage and inflammation (Kazacos, 1997;Sheppard& Kazacos, 1997).

Diagnostic
The diagnosis of baylisascariasis is difficult in live patients; there is no widely available, non-invasive definitive test. The combination of encephalopathy with cerebrospinal fluid (CSF) and peripheral eosinophilia and diffuse white matter disease on neuroimaging, with or without eye disease, in a patient from North America or Europe should strongly suggest the diagnosis of Baylisascaris NLM, and a history of exposure to raccoons or their feces should be sought.
Unless a brain biopsy is done and a larva is found, antemortem diagnosis usually depends on serology, with supportive evidence from other tests.

Serology
In neural larva migrans, antibodies to Baylisascaris can be found in serum and CSF; a rising titer is usually seen. An enzyme linked immunosorbent assay (ELISA), indirect immunofluorescence and immunoblotting (Western blotting) have been developed to detect anti-Baylisascaris antibodies. These serological assays are not commercially available, but they may be provided by university research laboratories. In the U.S., an ELISA is available from the Department of Comparative Pathobiology at Purdue University, West Lafayette, IN. Indirect immunofluorescence tests use frozen sections of B. procyonis third-stage larvae as an antigen. Enzyme-linked immunosorbent assay and Western blotting use excretory-secretory products from in vitro cultures of B. procyonis larvae as the antigen . Larval excretory-secretory antigens have been characterized as complex glycoproteins, with molecular masses of 10 kDa to 200 kDa, that contain several different sugar residues . Protein epitopes of 33-kDa to 45-kDa antigens appear to be recognized selectively by antibodies from B. procyonis-infected humans and animals but not by normal human or T. canis antibody-positive sera . In addition, children with clinical B. procyonis NLM are strongly positive for anti-Baylisascaris antibodies in CSF and serum and have consistently been negative for anti-Toxocara antibodies (Cunningham et al., 1994;Fox et al., 1985;Gavin et al., 2002;Kazacos, 2001;Moertel et al., 2001;Murray, 2002;Park et al., 2000;Rowley et al., 2000). In several of these cases, positive B. procyonis serology was confirmed by brain biopsy or at autopsy (CDC, 2002;Fox et al., 1985;Huff et al., 1984;Rowley et al., 2000). Acute and convalescent-phase titers characteristically demonstrate several fold increases in both serum and CSF anti-Baylisascaris antibody levels (Gavin et al., 2002;Moertel et al., 2001).
Although B. procyonis excretory-secretory (BPES) antigen-based ELISA and Western blot assays are useful in the immunodiagnosis of this infection, cross-reactivity remains a major problem. Recently, a recombinant B. procyonis antigen, BpRAG1, was reported for use in development of improved serological assays for the diagnosis of Baylisascaris larva migrans (Dangoudoubiyam et al., 2010). In a recent study, authors tested a total of 384 human patient serum samples in a BpRAG1 ELISA, including 20 patients with clinical Baylisascaris larva migrans, 137 patients with other parasitic infections (8 helminth and 4 protozoan), and 227 with unknown/suspected parasitic infections. A sensitivity of 85% and specificity of 86.9% was observed with the BpRAG1 ELISA, compared to only 39.4% specificity with the BPES ELISA. In addition, the BpRAG1 ELISA had a low degree ofcross-reactivity with antibodies to Toxocara spp. infection (25%), while the BPES antigen showed 90.6% cross-reactivity. Based on these results, BpRAG1 antigen has a high degree of sensitivity and specificity and should be very useful and reliable in the diagnosis and seroepidemiology of Baylisascaris larva migrans by ELISA (Dangoudoubiyam et al., 2011)

Laboratory tests
Although no routine laboratory test is considered diagnosticof B. procyonis NLM by itself, a number of studies provide additional supporting evidence. Most importantly, the presence of eosinophilia, particularly eosinophilic meningitis, should alert the physician to the possibility of a parasitic etiology (Lo Re& Gluckman, 2003;Rothenberg, 1998;Weller, 1993). Eosinophils are not normally present in CSF; their presence narrows the differential diagnosis of CNS disease and provides an early or theonly etiologic clue. In documented cases of NLM, the peripheral white blood cell count is usually mildly elevated, but eosinophilia may be marked. Cerebrospinal fluid cell counts may be normal at presentation and generally demonstrate only mild leukocytosis, again with eosinophilia. Notably, even in the absence of pleocytosis, demonstrable CSF eosinophilia may be evident. Because eosinophils are easily missed in unstained or Gram-stained CSF, it may be necessary to request Wright's orGiemsa stain of cytocentrifuged CSF specimens. In documented cases of NLM, CSF protein is generally normal or only mildly elevated, while www.intechopen.com Zoonosis Caused by Baylisascaris Procyonis 295 CSF glucose levels are normal. Although the finding of elevated serum isohemagglutinins, caused by cross-reactions between larval glycoproteins and human blood group antigens, is not specific for baylisascariasis, it does provide an additional clue to the diagnosis .

Other techniques
Imaging techniques and encephalography provide supportive evidence and help rule out other causes. In ocular larva migrans, an ophthalmoscopic examination may occasionally reveal large, motile larvae in the retina, as well as choroidioretinitis and other signs of DUSN. The presence of Baylisascaris larvae in the eye is also suggestive in cases with neurological signs. Biopsies of the CNS are occasionally definitive, but larvae are often absent from the sample. A definitive diagnosis can also be made retrospectively from CNS samples taken at autopsy. Baylisascaris larvae are much larger (up to 80 μm in diameter and up to 1900 μm long) than Toxocara larvae, and can also be distinguished by their morphology. However, parasite larvae can be difficult to identify within tissues, and misidentification is common. In tissues, the third stage larvae of B. procyonis cannot be differentiated from B. columnaris or B. melis. Epidemiological evidence, such as a history of exposure to raccoons but not skunks or badgers, can be suggestive.

Treatment
The prognosis for B. procyonis NLM is grave with or without treatment; among documented cases,there are no neurologically intact survivors. In this parasite is very important early clinical suspicion of raccoon roundworm meningoencephalitis.
The majority of cases have been treated with anthelmintics and corticosteroids. Empirical anthelmintic treatment with thiabendazole, fenbendazole, tetramisole, or ivermectin has failed to prevent death or unfavorable outcomes. Animal data suggest that albendazole and diethylcarbazinehave the best CSF penetration and larvicidal activity (Kazacos, 2001).
Treatment with albendazole is protective in animal models if eggs have been ingested, but symptoms have not yet developed. In humans, albendazole has been used prophylactically after exposure to raccoon latrines or other sources of eggs. Whether it is helpful in patients with clinical signs is uncertain, because the death of the parasite might worsen the inflammation.
Albendazole appears to have the more favorable pharmacologic profile, with good absorption, high serum concentrations of the active metabolite, good penetration across the blood-brain barrier, and minimal toxicity (de Silva et al., 1997;Jung et al., 1990).
Most clinical cases have been treated concurrently with anthelmintics and corticosteroids; the corticosteroids are used to suppress inflammation caused by the death of the larvae, as well as to dampen the existing inflammatory response. In a recent case a early intervention with both, albendazole and steroids, may have contributed to patient's partial recovery (Hajek et al., 2009) and a mild case of suspected B procyonis infection, with apparent early complete response after cerebellar edema, was treated with early corticosteroids and later albendazole (Pai et al., 2007).
Other supportive therapy may also be given. Recently, an acute eosinophilic meningoencephalitis, caused by Baylisascaris procyonis in a previously healthy teenager with a history of substance abuse, was treated with methylprednisolone; no antihelmintic drugs were administered (Chun et al., 2009).
In many cases, significant damage has already occurred by the time treatment is begun, and improvement is not seen. The best chance of recovery is expected with a very early diagnosis and treatment.

Prevention and control
In intermediate hosts, the risk of infection can be decreased by avoiding contact with raccoons, other definitive hosts and their feces. Raccoons can be discouraged from visiting homes and farms with the same measures that are used to prevent disease in humans.
Infections are difficult to prevent completely in pets allowed outdoors, as the infective eggs can survive for long periods in the environment. In dogs, monthly heartworm/nematode preventatives appear to decrease the risk of intestinal infection with B. procyonis. In high-risk areas, dogs that are not on these preventatives should receive regular fecal examinations to decrease the risk that they will shed eggs.
In zoos and other facilities, the housing for intermediate hosts should be designed to minimize exposure to raccoons, skunks and other definitive hosts. Captive raccoons and skunks should be kept in dedicated cages that can be cleaned, if necessary, with the harsh methods required to destroy Baylisascaris eggs. They should be tested regularly and dewormed when necessary, and they should not be fed wild animals that might carry larvae. Newly acquired definitive hosts should be quarantined and dewormed. Once contamination has occurred, it can be difficult to remove completely. Intermediate hosts in exhibits are sometimes treated prophylactically with pyrantel tartrate or ivermectin. Similarly to humans, animals with recent exposure might also be treated with albendazole to prevent the development of clinical signs.
In humans, risk of infection is greatest when infants or toddlers with geophagia or pica come in contact with raccoon latrines or an environment contaminated by infected raccoon feces. Young infants and toddlers, particularly those with pica or geophagia, should be kept away from potentially contaminated areas.
Raccoon latrines in and around homes and play areas shouldbe cleaned up and decontaminated. However, the longevity of B. procyonis eggs and their resistance to disinfection or decontamination makes successful environmental cleanup difficult. Recently a publication (Shafir et al., 2011) showed that eggs survived complete desiccation for at least 6 months at room temperature. Total loss of viability was observed after 7 months of desiccation. Eggs frozen at -15°C for 6 months demonstrated no reduction in viability. Freeze-thaw, including exposure to 5 cycles, did not affect survival. Although they did not specifically design experiments to assess the effect of chlorine on inactivation of B. procyonis eggs, exposure to undiluted household bleach for 90 min toremove the mammillated layer