Virulence of six strains of duck plague virus in eight waterfowl species.

Susceptibility of New World waterfowl to the Lake Andes strain of duck plague virus (DPV) was assessed by intramuscular inoculation of adult muscovies (Cairina moschata), mallards (Anas platyrhynchos), Canada geese (Branta canadensis), wood ducks (Aix sponsa), redheads (Aythya americana), gadwalls (Anas strepera), blue-winged teal (Anas discors), and pintails (Anas acuta). The relative virulence of DPV strains isolated from five United States and one Canadian location was established in muscovies, mallards, and Canada geese. Differences in DPV strain virulence were detected by formation of plaques in cell culture. Two strains that consistently formed plaques killed adult mallards while non-plaque forming strains killed hatchling but not adult mallards. Based on mortality after exposure to the Lake Andes strain, blue-winged teal, then wood ducks and redheads were highly susceptible, muscovies and gadwalls moderately susceptible, mallards and Canada geese less susceptible, and pintails the least susceptible. Mean death times were significantly (P < 0.01) different between adult muscovies (4.5 days) versus mallards and Canada geese (5.8 days each). Mean death time of the virulent Lake Andes and Minnesota strains were shorter (P < 0.05) than for the other four, less virulent DPV strains. Four of the less virulent strains killed hatchling but not adult mallards. Susceptibility to mortality was dependent upon age and route of inoculation. The intramuscular route of inoculation required the least amount of virus to kill mallard and muscovy ducks, the intranasal and conjunctival routes required more virus, and the oral route the most virus. This study was conducted from 1974 to 1977 between the months of September and April, with the exception of two titrations conducted in early May at the University of Wisconsin Department of Veterinary Science and the Charmany research facility of the University of Wisconsin-Madison.

Intrauterine transmission of Borrelia burgdorferi in dogs.

To determine whether intrauterine transmission of Borrelia burgdorferi could exist in dogs, 10 female Beagles were inoculated intradermally with approximately 1,000 B burgdorferi on day 1 of proestrus; inoculation was repeated every 2 weeks during the gestation period. Ten female control Beagles were similarly inoculated with phosphate-buffered saline solution. Prior to the start of the study, all females and 3 males used for breeding were seronegative for B burgdorferi on the basis of results of the indirect fluorescent antibody test and immunoblot (western analysis. Similarly, results of culture of blood for B burgdorferi were negative. All 20 of the females were bred naturally. Blood samples were collected weekly for serologic testing and culture. Blood samples were obtained from live pups on day 1 of life, then weekly until pups were 6 weeks old when they were euthanatized. Tissues were obtained for culture and testing by use of polymerase chain reaction (PCR). Of 10 spirochete-inoculated (SI) females, 8 became infected with B burgdorferi as evidenced by spirochete culture results and/or PCR-detected B burgdorferi DNA in the tissues of females or their pups. Of the 10 SI females, 8 delivered litters (3 to 7 pups) that had at least 1 neonatal or 6-week-old pup with B burgdorferi DNA-positive tissues (by PCR), and spirochetes were cultured from tissues from pups of 2 litters.(ABSTRACT TRUNCATED AT 250 WORDS)

Borrelia burgdorferi infection in dairy cows, rodents, and birds from four Wisconsin dairy farms.

A combination of culture and subsequent spirochete identification with the polymerase chain reaction technique was used to identify cows, rodents, and birds infected with Borrelia burgdorferi. Animals were trapped on four Wisconsin dairy farms during the summer of 1990. Farms 1 and 2 were located in counties nonendemic for Lyme disease and Farms 3 and 4 were located in counties endemic for Lyme disease. The results of the rodent and bird samples were as follows given as the number yielding organisms number tested: Farm 1, 1/17 Mus musculus and 2/52 Peromyscus domesticus; Farm 2, 4/49 M. musculus, 1/2 P. maniculatus, 1/1 P. leucopus, and 1/35 P. domesticus; Farm 3, 0/27 M. musculus, 0/5 P. leucopus, 0/12 P. maniculatus and, 3/58 P. domesticus; and Farm 4, 1/24 M. musculus, 2/19 P. leucopus, 1/12 Microtus pennsylvanicus, and 0/17 P. domesticus. One P. leucopus and one M. musculus from Farm 2 were pregnant and fetal tissues from both were positive. Cow blood sample results were as follows: Farm 1, 7/47 in July, and 2/45 in August; Farm 2, 0/28 in August and 0/23 in October; Farm 3, 0/13 in July and 1/18 in August 29; and Farm 4, 3/45 in August. Ticks were found on rodents on Farm 4 and on one bird on Farm 3. Spirochetemic cows, rodents, and birds were found in non-Lyme endemic counties suggesting that alternate modes of transmission other than by ticks may be important. Transplacental transmission was shown in M. musculus and P. leucopus.

Experimentally induced infection of cats with Borrelia burgdorferi.

To determine whether cats could be infected experimentally with Borrelia burgdorferi, 15 cats were inoculated with approximately 1,000 B burgdorferi. Seven cats were inoculated by the IV route, 2 by the oral route, 2 by the ocular route, and 4 by the oral-ocular route. Six control cats were inoculated with phosphate-buffered saline solution by the IV, oral, and ocular routes. Prior to the start of the study, all 21 cats were seronegative for B burgdorferi on the basis of results of the indirect fluorescent antibody (IFA) test, and their blood was B burgdorferi culture negative. All of the IV, orally, and ocularly inoculated cats developed IgG antibodies to B burgdorferi as detected by IFA testing. Of 4 oral-ocularly inoculated cats, 2 developed IFA-detectable antibodies and the remaining 2 cats developed low-titer response (1:128) on postinoculation (PI) day 10 only. All control cats remained seronegative. The organism was detected in blood smears from 2 of the IV inoculated cats on PI days 10 and 24 and from 2 oral-ocularly infected cats, 1 on PI days 17 and 24 and 1 on PI day 10. Spirochetes were not detected in the blood after PI day 24. The organism was isolated from tissues of only 1 cat (the lung of an ocularly inoculated cat necropsied at 7 months after inoculation). Spirochetes were not isolated from control cats. Neither clinical signs of infection nor gross or histologic abnormalities were found in any of the inoculated or control cats. Results indicate that cats are susceptible to infection with B burgdorferi, but clinically apparent disease may not be common.

Mortality from duck plague virus in immunosuppressed adult mallard ducks.

Environmental contaminants contain chemicals that, if ingested, could affect the immunological status of wild birds, and in particular, their resistance to infectious disease. Immunosuppression caused by environmental contaminants, could have a major impact on waterfowl populations, resulting in increased susceptibility to contagious disease agents. Duck plague virus has caused repeated outbreaks in waterfowl resulting in mortality. In this study, several doses of cyclophosphamide (CY), a known immunosuppressant, were administered to adult mallards (Anas platyrhynchos) to determine if a resultant decrease in resistance to a normally sub-lethal strain of duck plague virus would occur, and induce mortality in these birds. Death occurred in birds given CY only, and in birds given virus and CY, but not in those given virus only. There was significantly greater mortality and more rapid deaths in the duck plague virus-infected groups than in groups receiving only the immunosuppressant. A positively correlated dose-response effect was observed with CY mortalities, irrespective of virus exposure. A fuel oil and a crude oil, common environmental contaminants with immunosuppressive capabilities, were tested to determine if they could produce an effect similar to that of CY. Following 28 days of oral oil administration, the birds were challenged with a sub-lethal dose of duck plague virus. No alteration in resistance to the virus (as measured by mortality) was observed, except in the positive CY control group.

Systemic disease in Peromyscus leucopus associated with Borrelia burgdorferi infection.

Sixteen wild Peromyscus leucopus, trapped for the establishment of a breeding colony, developed signs of neurological damage (trembling, incoordination, circling, head tilt, and lameness of the rear legs) 2-47 days after capture in southern Wisconsin. Spirochetes were cultured from the brain of 5/11 mice, and Borrelia burgdorferi was cultured from 1 brain. A spirochete was isolated from the bladder of 1 mouse. The spirochete was identified by fluorescent antibody staining with the monoclonal antibody specific for B. burgdorferi, H5332. Serum antibodies to the spirochete were found in 14/15 mice. Negative results were obtained in all tests for viruses and bacteria, including Listeria (2/2), Mycoplasma (2/2), mouse hepatitis virus (10/10), Theilers's encephalomyelitis virus (GD VII) (8/8), REO 3 virus (2/2), and lymphocytic choriomeningitis virus (4/4). There was no bacterial growth from brains cultured on eosin methylene blue or blood agar (3/3). Histologic lesions included nonsuppurative cellular infiltrates in the brain, kidney, liver, and lung. Three outbred Swiss-Webster mice were inoculated orally with a suspension of the brain in BSKII medium, and 3 were inoculated with unpassed B. burgdorferi cultured from the brain of a P. leucopus with motor dysfunction. Five of the inoculated mice developed antibody titers of 1:128; one mouse was positive at 1:256. Motor signs of neurologic damage developed in 3/6 mice 2-24 weeks post-inoculation, and B. burgdorferi was detected in the brains of 2 mice by isolation and by fluorescent antibody.

Antibodies to Borrelia sp. in wild foxes and coyotes from Wisconsin and Minnesota.

Serum samples from 93 red foxes (Vulpes vulpes) and nine gray foxes (Urocyon cinereoargenteus) trapped in Wisconsin and 23 coyotes (Canis latrans) trapped in Wisconsin and Minnesota were tested for antibodies to Borrelia sp. with an indirect fluorescent antibody test which used Borrelia burgdorferi as the whole-cell antigen. Seven red foxes (8%) and two coyotes (9%) had antibody titers greater than or equal to 1:64. All the positive samples were from areas known to be endemic for human Lyme disease. Implications for the epizootiology of Lyme borreliosis in wild canids are not well understood, but even if these species are not actual reservoirs of B. burgdorferi they could serve to increase the range of the vector and establish new endemic foci of the spirochete.

Borrelia sp. infection in coyotes, black-tailed jack rabbits and desert cottontails in southern Texas.

Coyotes (Canis latrans) from southern Texas were sampled for antibodies to Borrelia burgdorferi from 1980 to 1986; black-tailed jack rabbits (Lepus californicus) and desert cottontails (Sylvilagus audubonii) were sampled in 1986. Coyote fetuses, adult coyote kidneys, and black-tailed jack rabbit and desert cottontail kidneys were cultured for B. burgdorferi in 1986. Results of indirect immunofluorescent antibody (IFA) tests for B. burgdorferi in coyotes were as follows (number positive at a dilution of greater than or equal to 1:128/number tested): 1980 (0 of 30), 1981 (0 of 21), 1982 (0 of 53), 1983 (0 of 78), 1984 (47 of 97), 1985 (20 of 88), and 1986 (42 of 80). Eight of 26 black-tailed jack rabbits and two of seven desert cottontails tested in 1986 had IFA titers to B. burgdorferi of greater than or equal to 1:128. Borrelia burgdorferi was isolated from one of five coyote fetuses, three of 31 adult coyote kidneys, and two of 10 black-tailed jack rabbit kidneys in 1986. These results indicate that B. burgdorferi infection has been present in coyotes in Texas, at least since 1984 and that transplacental transmission occurs.

Experimental inoculation of mallard ducks (Anas platyrhynchos platyrhynchos) with Borrelia burgdorferi.

Birds have been incriminated as disseminaters of Borrelia burgdorferi and have the potential to spread the organism over a wide geographic range. Borrelia burgdorferi has been isolated from the liver and blood of passerine birds and from Ixodes dammini removed from passerines. The objective of this study was to determine if waterfowl, specifically mallards (Anas platyrhynchos platyrhynchos), were susceptible to infection with B. burgdorferi. Eight ducks were inoculated with B. burgdorferi; four orally and four intravenously (i.v.) and two ducks were inoculated with phosphate buffered saline as controls. All eight inoculated birds became infected and developed antibodies to B. burgdorferi. The spirochete was isolated from cloacal material from an orally infected duck on day 22 postinoculation (PI) and from an i.v. infected bird on day 29 PI, from the blood of an i.v. infected bird on day 7 PI, and from the kidney of an orally infected bird. Borrelia burgdorferi was detected by indirect immunofluorescence using the B. burgdorferi specific monoclonal antibody H5332 in kidneys of three orally infected birds and one i.v. infected bird and from the mesentery of one orally infected bird. These findings show that mallard ducks are susceptible to infection by B. burgdorferi and that they can be infected orally and shed the organism in the droppings. Thus, mallards could disseminate B. burgdorferi over long distances without the need of an arthropod vector.

Heterogeneity in immunoblot patterns obtained by using four strains of Borrelia burgdorferi and sera from naturally exposed dogs.

This study evaluated the differences in immunoblot patterns when serum specimens from seropositive dogs were reacted against four strains of Borrelia burgdorferi. Intense bands were consistently detected for the 83-, 66-, 61- to 60-, 41-, and 31-kilodalton regions for all four strains. Most variations were observed in the regions of 45 to 34 and 26 to 15 kilodaltons. Adsorption studies suggested that one reason for the observed variability was a difference in proteins among the organisms, rather than a difference in migration of proteins. Therefore, knowledge and consistency of the test antigen are essential when evaluating and comparing canine immunoblot patterns to B. burgdorferi, but for diagnostic purposes all of the serum samples would have been considered positive regardless of the strain used.

Renal lesions associated with Borrelia burgdorferi infection in a dog.

Borrelia burgdorferi infection was diagnosed serologically in a dog with lethargy, stiffness, and anorexia. Treatment with ampicillin and chloramphenicol did not alleviate the signs. Azotemia, proteinuria, cylindruria, pyuria, and hematuria developed over a 3-month period. Antibody titer for B burgdorferi remained high (1:8,192) during this time. Renal histopathologic findings included severe, chronic, diffuse, membranoproliferative glomerulonephritis and moderate chronic, multifocal, interstitial nephritis. Borrelia burgdorferi organisms were identified in renal tissue and in urine by results of immunofluorescent studies and bacteriologic culture, respectively.

Susceptibility of the gray wolf (Canis lupus) to infection with the Lyme disease agent, Borrelia burgdorferi.

Four juvenile gray wolves (Canis lupus) were inoculated with live Borrelia burgdorferi. One received an intravenous inoculum, a second was inoculated subcutaneously, and two more were fed Peromyscus maniculatus sucklings which had earlier been inoculated with B. burgdorferi. The intravenously inoculated wolf developed a generalized lymphadenopathy and a persistent serum antibody titer to the spirochete which peaked at 1:512. Borrelia burgdorferi was visualized in liver sections of this wolf using direct immunofluorescent staining. The subcutaneously inoculated wolf showed a low and transient antibody response which peaked at 1:64, and manifested no clinical or postmortem abnormalities. The wolves which were fed inoculated mice showed no detectable antibody response. They were clinically normal throughout the project, and there were no detectable lesions at necropsy. Two control wolves were inoculated intravenously with formalin killed B. burgdorferi. Serum antibody titers of these controls peaked at 1:64 and 1:32, respectively, and fell to 1:16 by day 48 postinoculation. A survey of serum samples from 78 wild-trapped wolves from Wisconsin and Minnesota revealed that one was positive and another was suspect for B. burgdorferi infection based on presence of antibody to the spirochete. We conclude that the wolf is susceptible to infection by B. burgdorferi and that wolves are being infected in the wild.

Prevalence of ocular microorganisms in hospitalized and stabled horses.

Microorganisms from normal eyes of hospitalized and stabled horses were identified, and the frequency of isolation was compared between the 2 groups. Using standard techniques, swab specimens from both eyes of 22 hospitalized horses and both eyes of 18 stabled horses were cultured for aerobic bacteria and fungi. Ninety-six aerobic bacteria and 57 fungi were isolated. The predominant bacterial isolates were gram-positive organisms, most of which belonged to the genera Corynebacterium, Bacillus, Staphylococcus, and Streptomyces. Gram-negative organisms comprised less than one-fourth of the bacterial isolates, with the genera Neisseria, Moraxella, and Acinetobacter being the most commonly isolated. Environmental fungi Cladosporium and Alternaria accounted for half of all fungal isolates. In only 5 horses were fungi isolated without accompanying isolation of bacteria. The frequency of isolation of fungi was higher (P less than 0.01) in stabled horses. For bacteria, the frequency of isolation was higher (P less than 0.08) in male horses. Results of susceptibility testing were recorded as the percentage of all isolates susceptible to a given antimicrobic drug. Bacterial isolates were highly susceptible (greater than or equal to 90%) to neomycin, polymixin B, gentamicin, and chloramphenicol. Overall, filamentous fungi had highest susceptibility to natamycin (97%). Miconazole was highly efficacious (100% susceptibility) against Fusarium and Aspergillus.

Borreliosis in free-ranging black bears from Wisconsin.

Blood, kidney and tick samples were obtained from 18 hunter-killed black bears (Ursus americanus) from three sites in northern Wisconsin. A Borrelia sp., morphologically and antigenically similar to Borrelia burgdorferi, was isolated from the blood of two of the animals, and from the kidney of a third. Ixodes dammini and Dermacentor variabilis were found on the bears. This is the first report of borreliosis in the Ursidae, and of the primary vector of Lyme disease, I. dammini, from this host.

Immunoblot analysis of immunoglobulin G response to the Lyme disease agent (Borrelia burgdorferi) in experimentally and naturally exposed dogs.

Immunoblots were used to study the immunoglobulin G response to Borrelia burgdorferi in experimentally and naturally exposed dogs. Adsorption studies confirmed that the antibodies were specific for B. burgdorferi. Experimentally exposed dogs were asymptomatic. Naturally exposed dogs included both asymptomatic animals and animals showing signs compatible with Lyme disease. Naturally exposed dogs were from four geographic regions of the country. No differences were detected between immunoblot patterns of naturally exposed symptomatic or asymptomatic dogs from different areas of the country. The immunoblot patterns obtained with sera from experimentally exposed dogs were different from those obtained with sera from naturally exposed dogs and were characterized by reactivity to fewer and different protein bands. Immunoblot analysis using an OspA-protein-producing Escherichia coli recombinant showed that experimentally exposed dogs produced antibodies to OspA, whereas naturally exposed dogs did not. Modifications of the immune response over time, different routes of antigen presentation, and strain variation are factors postulated to account for the observed differences.