Migration behaviour and pathogenesis of five ascarid nematode species in the Mongolian gerbil Meriones unguiculatus

To understand the characteristic features of the Mongolian gerbil, Meriones unguiculatus, as an animal model of ascarid infections, the migration behaviour and pathogenesis of larvae were investigated in experimentally infected gerbils. Embryonated eggs from each of Toxocara canis, Baylisascaris procyonis, B. transfuga, Ascaris suum, and A. lumbricoides were orally inoculated into gerbils and larvae were recovered from various organs at designated periods. In T. canis-infected gerbils, larvae were present in the liver 3 days after infection and in the skeletal muscle and brain via the heart and lungs at a similar rate. In B. procyonis- and B. transfuga-infected gerbils, larvae were present in the lungs within 24 h after infection, with some having reached the brain by that time. After 24 h, larvae of B. procyonis tended to accumulate in the brain, while those of B. transfuga accumulated in skeletal muscles. In A. suum- and A. lumbricoides-infected gerbils, larvae remained in the liver on day 5 post-infection and elicited pulmonary haemorrhagic lesions, which disappeared 7 days after initial infection. Thereafter, no larvae of any type were recovered. Ocular manifestations were frequently observed in T. canis- and B. procyonis infected gerbils, but were rare in B. transfuga-infected gerbils. In the cases of A. suum and A. lumbricoides, migration to the central nervous system and eyes was extremely rare, and larvae had disappeared by 2 weeks post-infection. Fatal neurological disturbances were observed in B. procyonis-infected gerbils, whereas irreversible non-fatal neurological symptoms were observed in the case of B. transfuga.     

Baylisascaris procyonis (Nematoda: Ascaridoidea) eggs in raccoon (Procyon lotol) latrine scats in Orange County,California

Baylisascaris procyonis is the common intestinal nematode of the raccoon and is well-recognized as a cause of visceral, ocular, and neural larva migrans in many species of wild and domestic birds and mammals, including humans. To develop data on the prevalence of B. procyonis in Orange County, California, 800 distinct raccoon latrine sites were sampled in 4 spatial zones from 15 January to 31 December 2000. Counts of fecal eggs per gram (EPG) were determined and evaluated with reference to spatial zone and season of collection. No significant differences in EPG were noted among the spatial zones. However, EPG exhibited a significant rise (37,730 +/- 1,865) in the fall and a significant decline (26,204 +/- 1,446) in the winter (ANOVA, P = 0.045). The overall egg prevalence was 100%, and the overall mean EPG was 30,265 +/- 867.     

Distribution, prevalence, and genetic characterization of Baylisascaris procyonis in selected areas of Georgia

Baylisascaris procyonis is an intestinal nematode of raccoons (Procyon lotor) that can cause fatal larval migrans in numerous species of birds and mammals, including humans. Although this parasite has historically been absent in the southeastern United States, it has been found in isolated regions in the Appalachian Mountains and was recently documented in DeKalb County, Georgia. The first objective of the current study was to investigate the distribution and prevalence of B. procyonis in selected populations of raccoons in Georgia. Intestinal tracts of 312 raccoons from 25 Georgia counties were examined for B. procyonis. The only county where B. procyonis was detected was Clarke County, where 12 of 116 (10.3%) raccoons were infected. In Clarke County, significantly more juveniles (P = 0.049) were infected compared with adults, and no differences in prevalence were noted by sex, season of capture, or land use (rural vs. urban); however, significantly (P = 0.0370) higher worm burdens were found in infected raccoons from urban/suburban locations compared with rural areas. In addition, Toxascaris leonina , a morphologically similar ascarid, was found in 3 raccoons from Clarke County (n = 2) and Morgan County (n = 1). A second objective was to determine if sequence polymorphisms were associated with B. procyonis from different geographic regions. Because sequences from a single worm from Japan had been entered into GenBank, we obtained nematodes from Kentucky and Texas for comparison with our samples from Georgia. Sequence analysis of the 18S and 5.8S rRNA genes and the internal transcribed spacer (ITS) -1 and ITS-2 regions confirmed Georgia samples were B. procyonis. Although several polymorphic bases were observed within both ITS regions, none was associated with a particular geographic location. These data indicate that the distribution of B. procyonis within Georgia is increasing and only limited genetic variation is present in the rRNA and ITS gene regions among B. procyonis from the southern United States and introduced populations in Japan.     

Enterovirus 71 Infection Causes Severe Pulmonary Lesions in Gerbils,Meriones unguiculatus,Which Can Be Prevented by Passive Immunization with Specific Antisera

Neurogenic pulmonary edema caused by severe brainstem encephalitis is the leading cause of death in young children infected by Enterovirus 71 (EV71). However, no pulmonary lesions have been found in EV71-infected transgenic or non-transgenic mouse models. Development of a suitable animal model is important for studying EV71 pathogenesis and assessing effect of therapeutic approaches. We had found neurological disorders in EV71-induced young gerbils previously. Here, we report severe pulmonary lesions characterized with pulmonary congestion and hemorrhage in a gerbil model for EV71 infection. In the EV71-infected gerbils, six 21-day-old or younger gerbils presented with a sudden onset of symptoms and rapid illness progression after inoculation with 1×10^5.5 TCID₅₀ of EV71 via intraperitoneal (IP) or intramuscular (IM) route. Respiratory symptoms were observed along with interstitial pneumonia, pulmonary congestion and extensive lung hemorrhage could be detected in the lung tissues by histopathological examination. EV71 viral titer was found to be peak at late stages of infection. EV71-induced pulmonary lesions, together with severe neurological disorders were also observed in gerbils, accurately mimicking the disease process in EV71-infected patients. Passive transfer with immune sera from EV71 infected adult gerbils with a neutralizing antibody (GMT=89) prevented severe pulmonary lesion formation after lethal EV71 challenge. These results establish this gerbil model as a useful platform for studying the pathogenesis of EV71-induced pulmonary lesions, immunotherapy and antiviral drugs.     

Frequency of deposition and location of Baylisascaris procyonis eggs in raccoon feces

Baylisascaris procyonis is a large ascarid nematode found in the small intestine of raccoons (Procyon lotor). Infection with larvae of B. procyonis can produce visceral, ocular, and neural larval migrans in humans. Infected raccoons can shed millions of eggs a day in their feces. However, it is unknown whether eggs are consistently shed or whether eggs occur at irregular intervals by the population of female nematodes within a host. We trapped, infected, and collected daily fecal samples from 11 raccoons maintained in captivity. Eggs from B. procyonis were obtained from anterior, central, and posterior sections of raccoon feces, isolated by flotation, and quantified under 100× magnification. Naturally infected raccoons were collected and used as a comparison with the experimentally infected group. All raccoons in the experimental group (n=11) became infected with B. procyonis after consuming one infected mouse. Additionally, differential egg deposition rates were observed among individual raccoons from the experimental and naturally infected groups. Mean number of eggs per gram of feces (means±SE) was 16,563±4,321, which was less than previously reported for the species. However, no differences (F(2,30)=0.84, P=0.45) were noted in mean number of eggs per gram of feces among fecal sections. Wildlife biologists, veterinarians, health officials, and researchers of B. procyonis should collect daily fecal samples for a minimum of 3 days before identifying a raccoon as negative for B. procyonis infection. However, it does not matter where within the fecal matter the sample is obtained.     

Physicochemical characterization and monoclonal and polyclonal antibody recognition of Baylisascaris procyonis larval excretory-secretory antigens

Baylisascaris procyonis larval excretory-secretory (ES) antigens consisted of complex glycoproteins ranging from 10 kDa to over 200 kDa as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis and lectin binding. Five monoclonal antibodies (Bapr1-Bapr5) produced against B. procyonis ES antigens were assayed by western blotting with larval ES antigens from B. procyonis, Baylisascaris melis, Baylisascaris transfuga, Ascaris suum, and Toxocara canis. Bapr1 and Bapr2 recognized periodate-sensitive epitopes on 14-kDa ES components of B. procyonis, B. melis, and B. transfuga, whereas Bapr4 and Bapr5 recognized periodate-resistant epitopes present on 55-kDa ES components of B. procyonis and B. melis. Bapr3 primarily recognized periodate-resistant epitopes on 33-45-kDa components of B. procyonis and B. melis ES. Heterologous rabbit antisera cross-reacted with many B. procyonis ES antigens on western blots, but recognition of the 33-45-kDa components was genus-specific. Normal human sera and T. canis-positive human sera also cross-reacted with many B. procyonis ES antigens, including those of 33-45 kDa. However, periodate oxidation markedly decreased cross-reactions and allowed for differential immunodiagnosis of B. procyonis versus T. canis. These studies demonstrated that antibody recognition of carbohydrate epitopes on ES components is an important cause of cross-reactions in antibody detection assays. Recognition of periodate-resistant (protein) epitopes on the 33-45-kDa B. procyonis ES components appears to be useful for genus-specific immunodiagnosis of larva migrans caused by Baylisascaris spp.     

Larva migrans by Baylisascaris transfuga: fatal neurological diseases in Mongolian jirds, but not in mice

Raccoon roundworms (Baylisascaris procyonis) and other Baylisascaris species cause patent or latent larva migrans (LM) in a variety of mammals and birds, including humans. It is not clear whether LM by Baylisascaris transfuga, roundworms of bears, is associated with clinical neurological disorders. To clarify this issue, ICR and BALB/c mice as well as Mongolian jirds (Meriones unguiculatus) were orally inoculated with 2,000-5,000 embryonated eggs of B. transfuga. In mice, the ascarid caused symptomatic LM of limited extent and duration, whereas the infection was fatal in jirds; i.e., they exhibited general signs such as severe depression and emaciation on days 8-11 postinfection (PI) and died, or they developed progressive and fatal neurological disorders after day 14 PI. Histological examination showed B. transfuga larvae in the brain of all mice and jirds examined, and the larvae collected from them developed to a size comparable with that of B. procyonis. There existed, however, critical differences in host reactions against larvae localized in the brain of mice and jirds; B. transfuga larvae found in mice were surrounded by granulomatous reactions and immobilized, whereas larvae found in jirds were free from any host reaction and mobile, causing extensive malacia.     

Land-use effects on prevalence of raccoon roundworm (Baylisascaris procyonis)

The raccoon (Procyon lotor) is the definitive host of Baylisascaris procyonis, a large intestinal roundworm that is zoonotic and can result in fatal or severe central nervous system disease in young children. Prevalence of infection among raccoon populations often is high, and in the midwestern United States, B. procyonis has been reported in 68-82% of raccoons. Raccoon populations have increased in response to changes in human land use, and often reach higher densities in urban and suburban landscapes than rural landscapes. However, shifts in foraging behavior among urban raccoons could impact the transmission of B. procyonis if small vertebrate intermediate hosts are not a significant part of the raccoon diet. The objective of this study was to compare prevalence of B. procyonis infection between urban and rural raccoon populations on a regional scale. Necropsy was done on 204 raccoons collected from September through February during 2000-2005 from seven states across the Midwest (regional sample). Baylisascaris procyonis was found in 54% of examined raccoons. Prevalence differed between land-use types (chi2=11.56, df=1, P=0.0007), and was higher among animals collected from rural locations (65%) than those collected in urban locations (41%). Intensity of infection also differed (F=5.52, df=1, P=0.02), with rural raccoons having greater worm burdens (x=29.63+/-36.42) than urban raccoons (x=13.85+/-18.47). Despite high densities of raccoons in urban landscapes, fewer urban raccoons were infected with B. procyonis, suggesting decreased dependence on intermediate hosts as a food source. This possible explanation was supported by a similar trend in prevalence among subsamples of raccoons collected from three Chicago-area populations (local samples) with differing levels of urbanization, population densities, and foraging behavior that had been intensively monitored during 1995-2002. Decreased transmission of B. procyonis in urban landscapes may be due to decreased predation of intermediate hosts, and contact of juvenile raccoons with B. procyonis eggs may be an important factor in maintaining infections within such populations.     

Multiple in vivo passages enhance the ability of a clinical Helicobacter pylori isolate to colonize the stomach of Mongolian gerbils and to induce gastritis

The Mongolian gerbil is an excellent animal model for Helicobacter pylori-induced gastritis in humans. In this study, initially low colonization rates of the H. pylori strains ATCC 43504, SS1, or HP87 inoculated into gerbils caused difficulties in establishing this model. In order to increase the colonization ability and pathogenicity, the clinical HP87 isolate was selected for adaptation to the gerbil stomach by multiple in vivo passages through gerbils. Development of gastritis was examined histologically at 4-52 weeks after infection. The proportion of gerbils which tested positive for H. pylori by culture at four weeks after inoculation gradually increased from 11.1% of gerbils inoculated with HP87 without prior in vivo passage (P0) to 100% of gerbils inoculated with HP87 with seven in vivo passages (P7). In addition, adaptation of HP87 resulted in more severe histopathological changes. Gerbils infected with adapted HP87 (P7) exhibited severe infiltration by monomorphonuclear and polymorphonuclear leukocytes in the mucosa, submucosa, and subserosa of the gastric antrum, as well as epithelial changes consisting of hyperplasia, erosion, and ulceration. Histopathological changes increased in severity from four to 52 weeks after infection. Adaptation of HP87 during its passages through gerbils could be due to genetic changes in bacterial colonization factors. Identification of these changes might be useful to understand the underlying mechanism of gastric adaptation and pathogenesis of H. pylori.     

Prevalence of Baylisascaris procyonis in habitat associated with Allegheny woodrat (Neotoma magister) populations in Indiana

Allegheny woodrat (Neotoma magister) populations have been in decline across their range since the late 1970s. Hypotheses proposed to explain these declines include habitat fragmentation and loss, decreased food availability, and increased mortality from infection with Baylisascaris procyonis. We investigated the prevalence of B. procyonis at raccoon (Procyon lotor) latrines in woodrat cliff habitats (n = 18) along the Ohio River in southern Indiana in 1995. We located 275 latrines (mean = 15.3/site; range, 6-34) and found B. procyonis in 13 (4.7%) latrines across all sites. When present at a site, B. procyonis occurred, on average, at 11.1%of latrines (range, 3-36%). Woodrat abundance, determined through a concurrent live-trapping program, was significantly higher (χ(2) = 5.12, df = 1, P = 0.024) at sites where B. procyonis was not found (9.5 ± 2.52) than at sites with B. procyonis (3.7 ± 2.2). Our analyses support the hypothesis that this parasite could contribute to declines in woodrat abundance. Because woodrats cache nonfood items, including raccoon feces, and are highly susceptible to the parasite, they are at increased risk for B. procyonis infection, which could be deleterious, especially to small populations.     

Survey of raccoons on Key Largo, Florida, USA, for Baylisascaris procyonis

Numbers of the endangered Key Largo woodrat (KLWR; Neotoma floridana smalli) have been declining for at least 25 yr. The raccoon (Procyon lotor) roundworm, Baylisascaris procyonis, has been found to have an adverse effect on the survival of Alleghany woodrats (N. magister). High densities of raccoons can exacerbate this problem by increasing the amount of feces containing viable eggs of B. procyonis available to woodrats. In 2002, 64 fecal samples were collected and examined for eggs of B. procyonis from >32 raccoons within the KLWR's known range on Key Largo, Florida, USA. All samples were negative for eggs of B. procyonis. Raccoon density in this area was approximately 0.62 raccoons/ha. Despite this high density of raccoons, B. procyonis does not appear to be a threat to the KLWR population.     

Severe and fatal central nervous system disease in humans caused by Baylisascaris procyonis, the common roundworm of raccoons: a review of current literature

Baylisascaris procyonis, a parasitic infection of raccoons, causes severe neurologic disease in humans when infective eggs from raccoon feces are ingested. Definitive diagnosis is challenging, but can be made by isolation of larvae in brain biopsy or exclusion of other potential causes of eosinophilic meningoencephalitis. Prevention efforts are critical due to the lack of effective treatment.     

Kinetics of T cell cytokine gene expression in gerbils after a primary subcutaneous Brugia pahangi infection

The majority of patients infected with lymphatic filariae are microfilaremic but tend to manifest little obvious pathology because of the infections. Data collected from the Mongolian gerbil-Brugia spp. model for human lymphatic filariasis suggest this experimental animal model system most closely represents this patient group and will be useful in studying immunological parameters associated with chronic infections. This article reports the quantitation of interleukin (IL)-4, IL-5, IL-10, IL-13, and interferon (IFN)-gamma messenger RNA (mRNA) in gerbils after a primary subcutaneous infection with Brugia pahangi. Chronically infected gerbils showed elevated IL-4 in all tissues, compared with earlier time points, linking this Th2 cytokine to the downregulation of responsiveness, which develops in gerbils and humans. Both IL-5 and IL-13 mRNA expression were transient in all tissues. The peak in IL-5 at 14-28 days postinfection reflects the peak of peripheral eosinophilia observed in B. pahangi-infected gerbils. Little IFN-gamma mRNA was reported from chronically infected gerbils. The data collected thus far suggest that the expression profile of many of the measured cytokines in B. pahangi-infected gerbils reflects what is seen in an important subset of humans infected with lymphatic filariae, the microfilaremic, asymptomatic patient.     

Ocular larva migrans caused by Toxocara cati in Mongolian gerbils and a comparison of ophthalmologic findings with those produced by T. canis

To elucidate the pathogenic potential of Toxocara cari, we observed the ophthalmologic changes of the fundi in Mongolian gerbils, Meriones unguiculatus, after oral inoculation of 17 embryonated eggs/g body weight. Ophthalmic conditions in 8 T. cati-infected gerbils were monitored using an ophthalmoscope from day 0 to day 156 and were compared with those of 57 T. canis-infected gerbils. The results showed that T. cati larvae migrated into the eye of the gerbil and then elicited ophthalmic changes, including retinal (25%) and vitreous (50%) hemorrhaging, vasculitis (37.5%), and exudative lesions (25%). Lesions were less prevalent, however, in T. cati-infected than in T. canis-infected gerbils. Unlike in T. canis-infected gerbils, the hemorrhagic lesions did not reappear in T. cati-infected gerbils after they were absorbed. These findings suggested that T. cati larvae are a potentially hazardous pathogen for ocular toxocariasis and that Mongolian gerbils infected with T. cati may be a useful model for the study of human ocular toxocariasis caused by T. cati. This is the first study to report that T. cati larvae can induce ophthalmic lesions in the retina of gerbils.     

Helminths of the raccoon (Procyon lotor) in western Kentucky

Seventy raccoons (Procyon lotor) from western Kentucky were examined for helminths from December 1985 through May 1986. Twenty-three species of helminths were collected including 10 species of Trematoda (Brachylaima virginiana, Euryhelmis squamula, Eurytrema procyonis, Fibricola cratera, Gyrosoma singulare, Maritreminoides nettae, Mesostephanus appendiculatoides, Metagonimoides oregonensis, Paragonimus kellicotti, Pharyngostomoides procyonis), 2 species of Cestoda (Atriotaenia procyonis, Mesocestoides variabilis), 10 species of Nematoda (Arthrocephalus lotoris, Baylisascaris procyonis, Capillaria putorii, C. plica, Crenosoma goblei, Dracunculus insignis, Gnathostoma procyonis, Molineus barbatus, Physaloptera rara, Trichinella spiralis), and 1 species of Acanthocephala (Macracanthorhynchus ingens). A mean of 6.4 (3-11) helminth species per host was recorded. Fibricola cratera, Atriotaenia procyonis, Mesocestoides variabilis, Arthrocephalus lotoris, Capillaria plica, Dracunculus insignis, Molineus barbatus, and Physaloptera rara were ubiquitous parasites of the raccoon, whereas specific nidi were observed for Eurytrema procyonis, Gyrosoma singulare, Paragonimus kellicotti, Baylisascaris procyonis, Trichinella spiralis, and Macracanthorhyncus ingens. With an overall prevalence of 10% or higher, 15 of the 23 helminth species were considered common parasites of the raccoon in western Kentucky. When the 10% prevalence rate was applied within geographical quadrants to correct for the presence of nidi it was found that 18 of the 23 helminth species were common and 5 were regarded as rare parasites of the raccoon. Two species of nematodes, T. spiralis and B. procyonis, displayed a markedly higher prevalence in male raccoons.     

The effect of fasting time on plasma total cholesterol concentration in Mongolian gerbils

This study investigated the effect of length of fasting time on plasma total cholesterol response of male Mongolian gerbils (Meriones unguiculatus). Plasma cholesterol levels from fed and fasted gerbils were also compared with those reported for humans under similar metabolic states. Plasma total cholesterol response showed a significant quadratic relationship with time over a 15-hour period. Between 6 and 9 hours of fasting (the time during which plasma triglyceride concentration became relatively constant), the average plasma total cholesterol concentration was 178 mg/dl, compared with a zero hour (fed) cholesterol level of 265 mg/dl. The difference in plasma cholesterol levels observed in fed and fasted gerbils is unlike what has been reported for humans. Results from most human studies show no differences in plasma total cholesterol concentrations for fed and fasted subjects. Failure to consider species differences in metabolic responses may have implications when results from animal experiments are extrapolated to humans.     

Detection of DNA from the zoonotic raccoon roundworm Baylisascaris procyonis in a French wolf

Baylisascaris procyonis is a zoonotic nematode whose main definitive host is the raccoon, an invasive carnivore in Europe introduced from the United States. B. procyonis causes larva migrans with poor prognosis in humans. This parasite was unexpectedly detected in France for the first time upon molecular screening of wolf faecal samples. Because no patent infection was found, the wolf cannot be considered as a definitive host. This discovery of B. procyonis in France nonetheless raises questions about the parasite status of the expanding raccoon populations in the country, which will be investigated in the future.     

Enterovirus 71-Induced Neurological Disorders in Young Gerbils,Meriones unguiculatus: Development and Application of a Neurological Disease Model

A reliable disease model mimicking Enterovirus 71 (EV71) infection in humans is essential for understanding pathogenesis and for developing a safe and effective vaccine. Commonly used rodent models including mouse or rat models are not suitable for vaccine evaluation because the rodents are resistant to EV71 infection after they reach the age of 6 days. In this study, 21-day-old gerbils inoculated intraperitoneally (IP) with a non mouse-adapted EV71 strain developed neurological lesion-related signs including hind limb paralysis, slowness, ataxia and lethargy similar to those of central nervous system (CNS) infection of EV71 in humans. The infected gerbils eventually died of the neurological lesions and EV71 could be isolated from lung, liver, spleen, kidney, heart, spinal cord, brain cortex, brainstem and skeletal muscle. Significantly high virus replication was detected in spinal cord, brainstem and skeletal muscle by cellular analysis, real-time quantitative PCR (RT-PCR) and immunohistochemical staining. Histopathologic changes such as neuronal degeneration, neuronal loss and neuronophagia were observed in spinal cord, brain cortex, brainstem, and skeletal muscle along with necrotizing myositis and splenic atrophy. Gerbils that received two doses of inactive whole-virus vaccine showed no EV71-specific symptoms after challenged with EV71. In contrast, gerbils that received mock vaccination died of EV71-induced neuropathology after challenged with EV71. The result indicates that gerbils can serve as a reliable disease model for evaluating safety and efficacy of EV71 vaccine.     

Changes in transmission of Baylisascaris procyonis to intermediate hosts as a function of spatial scale

Physical changes in landscapes alter the abundance and distribution of species. Higher-order effects can occur when changes in ecological processes result in altered interspecific interactions and subsequent changes in a species' abundance or persistence. Baylisascaris procyonis , a roundworm parasite of raccoons ( Procyon lotor ), is pathogenic to numerous small vertebrates that serve as intermediate hosts, including white-footed mice ( Peromyscus leucopus ). Raccoons have responded positively to agriculturally induced changes in landscapes, with potential consequences for intermediate hosts of B . procyonis . We examined white-footed mice from a homogeneous, predominantly forested landscape in south-central Indiana and a heterogeneous, predominantly agricultural landscape in northwestern Indiana for presence of larval B . procyonis . We compared prevalence of infection, intensity of infection, and average number of larvae per mouse between the landscapes, and among forest patches within the highly fragmented agricultural landscape. Prevalence, intensity of infection, and average number of larvae were significantly higher in the highly fragmented landscape. Within the agricultural landscape, regression models predicted probability of infection, intensity of infection, and average number of larvae per mouse per patch as functions of forest patch area and isolation. We conclude that positive responses of raccoons to agriculturally induced fragmentation of forests have resulted in increased encounter rates between white-footed mice and infective stages of B . procyonis between and within landscapes, resulting in increased transmission of the parasite to intermediate hosts.