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Effectiveness of Metarhizium anisopliae (Deuteromycetes) against Ixodes scapularis (Acari: Ixodidae) engorging on Peromnyscus leucopus.

With the incidence of Lyme disease increasing throughout the United States, reducing risk of exposure to the disease is of the utmost concern. In the northeastern U.S., the blacklegged tick, Ixodes scapularis, is the primary vector and the white-footed mouse, (Peromyscus leucopus), the primary reservoir for Borrelia burgdorteri, the bacterium causing Lyme disease. Targeting I. scapularis engorging on white-footed mice with an effective biological control agent, such as the fungus Metarhizium anisopliae, could be an effective and relatively safe control technique. In 2002-2003, we performed laboratory and field experiments to determine whether M anisopliae-treated nesting material could effectively control larval I. scapularis ticks engorging on white-footed mice, and therefore reduce the number of infected nymphal I. scapularis questing the following summer. Our laboratory experiment demonstrated a strong negative effect of M. anisopliae-treated nesting material on survival of I. scapularis larvae feeding on P. leucopus, with 75% versus 35% larval mortality in treatment versus control nests. Our field trials caused only modest, localized reductions in nymphal abundance and had no effect on the proportion of nymphal I. scapularis infected with B. burgdorferi. Field results probably could be improved by increasing the density of nestboxes to allow fungal delivery to a higher proportion of the mouse population and by deploying nestboxes in an area with lower mammalian diversity, such as a suburban landscape.

Animals↗

Lyme borreliosis: a relapsing fever-like disease?

To determine by xenodiagnosis length and concentrations of spirochetemias produced by Borrelia burgdorferi in white-footed mice (Peromyscus leucopus), laboratory reared mice were inoculated with either spirochete-containing tick suspensions or BSK II spirochete culture and were exposed for as long as three months to larval Ixodes dammini. Upon development to the nymphal stage, ticks were evaluated for spirochetal infections by direct immunofluorescence. All mice were found to circulate spirochetes for at least three months in concentrations sufficient to infect ticks. The percentage of infected ticks alternated from low to high, suggesting occurrence of episodes of mild and heavy spirochetemias. The results suggest that B. burgdorferi in its animal hosts and possibly also in humans causes prolonged spirochetemias characterized by episodes of alternating high and low concentrations of spirochetes as reflected by similar percentages of infected ticks. The long persistence of spirochetes in the peripheral blood stream and the cyclical form of Lyme borreliosis appear to be related, as in relapsing fevers, to the capacity of B. burgdorferi to undergo antigenic variations.

Animals↗

Factors influencing the antigenic reactivity of Borrelia burgdorferi, the Lyme disease spirochete.

The immunologic reactivity of some antigens of low passage Borrelia burgdorferi varied with the temperature at which it was cultivated in vitro, with long-term cultivation in vitro, and following passage in white-footed mice, Peromyscus leucopus. The low passaged uncloned strain Sh-2-82 had either more antigens or antigens which were more immunoreactive when grown at various temperatures from 28-39 degrees C. The influence of temperatures was less evident, however with a higher passage of the same strain. Immune sera from white-footed mice experimentally infected with the spirochete were more reactive with antigens of the inoculum than with antigens of spirochetes reisolated later. The antigenicity of the Lyme borreliosis spirochete is becoming more complex and this may relate to the complex life cycle that includes tick and mammalian hosts.

Animals↗

Differential detachment from resting hosts of replete larval and nymphal Ixodes ticks.

To determine whether replete subadult Ixodes ticks detach more frequently from resting than from active hosts, diverse rodents and lizards were caged in an apparatus designed to record the ticks' sites of detachment relative to the resting site of the host. Replete larval Ixodes ricinus and Ixodes dammini accumulated mainly beneath the resting places of the mice (Apodemus agrarius and Peromyscus leucopus) most frequently parasitized in nature. Although nymphal I. ricinus similarly detached where these mice rested, nymphal I. dammini detached more randomly. When lizards were used as hosts, both subadult stages of I. ricinus tended to detach away from their main resting sites; these ticks detached from squirrels more randomly. Detachment ratios for other rodent hosts, that are abundantly infested by the larvae of these ticks in nature (Apodemus flavicollis and Clethrionomys glareolus), could not be derived because nymphs generally failed to attach. Our observations are consistent with reports that both subadult stages of I. dammini, but not the adult, feed on the same kind of nest-dwelling hosts and that the host range of I. ricinus is less focused. Detachment of mouse-feeding larvae from resting mice promotes subsequent nymphal attachment to conspecific hosts, and the absence of such behavior among nymphs facilitates access of the resulting adults to deer.

Animals↗

Experimental infection of the white-footed mouse with Borrelia burgdorferi.

Laboratory raised white-footed mice (Peromyscus leucopus) were inoculated experimentally with live spirochetes (Borrelia burgdorferi), the etiologic agent of Lyme disease (borreliosis). Prior to inoculation, mouse sera were tested with an indirect fluorescent antibody test, and all mice were seronegative. All inoculated mice seroconverted. In tick transmission studies, immature stages of Ixodes dammini and Dermacentor variabilis attached and fed to repletion on mice, but only I dammini transferred spirochetes to uninfected mice. Mice were susceptible to oral infection and transmitted infection to each other through direct contact. Infection did not affect reproduction or development of young born from infected dams, nor did spirochetes appear in the tissues of neonates. Borrelia burgdorferi spirochetes were identified in the kidneys, liver, and spleen of infected mice by the use of the modified Steiner silver stain and a tissue indirect fluorescent antibody test. Spirochetes also were isolated in culture of the heart blood of 1 mouse. Regardless of the source of infection, no mice developed clinical signs or had any pathologic change resulting from infection. Spirochetes were always observed extracellularly within interstitial spaces. Antibody titers persisted for over 4 months in some mice and spirochetes were found in the tissues of 1 mouse that had been infected 1 year earlier.

Animals↗

In vivo formation of a free radical metabolite of ethanol.

Free radical metabolism of ethanol has been suggested as a factor in its hepatotoxicity. Although evidence of lipid radical formation due to ethanol treatment in vivo has been reported, free radicals from ethanol itself have not been detected in living animals. However, by applying the EPR spectroscopy technique of spin trapping to the study of ethanol-treated alcohol dehydrogenase-deficient deermice (Peromyscus maniculatus), we have detected the alpha-(4-pyridyl-1-oxide)-N-t-butylnitrone (POBN)/alpha-hydroxyethyl radical adduct in bile from animals administered [1-13C]ethanol and the spin trap POBN. Hyperfine coupling constants were aN = 15.48, a beta H = 2.02, and a beta 13C = 4.61 G. In addition, an ethanol-dependent but 13C-invariant radical adduct, presumably lipid derived, was detected. Hyperfine coupling constants were aN = 15.38 and a beta H = 2.5 G. This report demonstrates, for the first time, the in vivo formation of the alpha-hydroxyethyl free radical metabolite of ethanol.

Animals↗

Molecular characterization of sylvatic isolates of Trichinella spiralis.

Genetic relationships of 20 Trichinella isolates from Indiana wildlife were assessed and compared to Trichinella isolated from an infected swine herd. Trichinella larvae were isolated from coyotes, mink, raccoons, and red foxes. The larvae were maintained and amplified in white mice (ICR) and wild mice (Peromyscus leucopus). Differences in phenotypic characters of sylvatic isolates in the 2 laboratory hosts included an approximately 10-30-fold increase in parasite fecundity in wild mice. DNA for each isolate was extracted from Trichinella larvae and analyzed by dot-blot hybridization using a repetitive DNA probe pBP2 that recognizes DNA sequences specific for swine Trichinella. The probe hybridized only to Trichinella from swine and a single coyote isolate. Restriction endonucleases were used to digest DNA and the resulting fragments were separated by gel electrophoresis. Based on the presence of repetitive DNA sequences in the Trichinella genome, distinctive banding patterns were seen among the isolates. Trichinella isolated from swine had a pattern distinct from all sylvatic isolates except 1 from a coyote. Because this coyote was from the same general locality as the swine Trichinella outbreak, it was concluded that the isolate represents transmission of swine trichinellosis to the wildlife population. Further analysis using the enzyme Cla I identified unique banding patterns for wild isolates, suggesting that the sylvatic group is a genetically heterogeneous complex.

Animals↗

Safety and immunogenicity of ERA strain of rabies virus propagated in a BHK-21 cell line.

The ERA strain of rabies virus was propagated in a baby hamster kidney cell line (BHK-21/C13). The viral titer was 10(1.8) tissue culture infective doses (TCID) higher than that of commercial ERA vaccine. The ERA/BHK-21 vaccine in baits retained titers of 10(6.3) to 10(6.4), TCID when subjected to daily temperature fluctuations from 9 degrees C to 24 degrees C for 21 days. This titer, according to a dose response in laboratory foxes, was still capable of immunizing up to 100% of foxes consuming a bait. The ERA/BHK-21 vaccine, when presented in baits, produced antibodies in 80 to 100% of dogs consuming more than one bait. Duration of immunity in foxes, from feeding the ERA strain rabies virus in baits, as determined by resistance to challenge with virulent virus, was at least 48 months. The vaccine strain retained some pathogenicity for nontarget species. In tests carried out on foxes, raccoons, dogs, cats and cattle, the vaccine did not cause vaccine-induced rabies. One of 14 skunks which consumed four baits developed vaccine-induced rabies, but virus could not be isolated from the salivary glands of this animal. The vaccine, when presented in baits, caused vaccine-induced rabies in 37% of laboratory mice, 3.4% of Microtus and 2.6% of Peromyscus species. Rabies virus could not be isolated from the salivary glands of rodents with vaccine-induced rabies. It was concluded that ERA virus propagated in BHK-21/C13 cells and incorporated in an acceptable bait produced a high titer, stable, immunogenic and safe vaccine for foxes.

Administration, Oral↗

Differential distribution of immature Ixodes dammini (Acari: Ixodidae) on rodent hosts.

Ectoparasites such as ixodid ticks that remain attached to hosts for several days while feeding on blood are able to overcome the inflammatory and immune responses of some hosts and not others. The immature stages of the deer tick Ixodes dammini are found more frequently on the white-footed mouse, Peromyscus leucopus, than on other rodents. We propose that P. leucopus is more tolerant to I. dammini than is a less common host, the meadow vole, Microtus pennsylvanicus. To test this hypothesis, the distribution patterns and engorgement indices were determined for larval and nymphal I. dammini collected from wild-caught P. leucopus and M. pennsylvanicus. There were more immature ticks, which were more fully engorged, on P. leucopus than on M. pennsylvanicus. There were more and better engorged ticks on male than on female hosts. Laboratory studies on the number and weights of larval I. dammini collected off naive and previously exposed P. leucopus and M. pennsylvanicus support the results of the field study. Fewer larval ticks were recovered from previously exposed M. pennsylvanicus than P. leucopus, and the ticks weighed less. Larval and nymphal ticks aggregated among hosts in the study grid, and higher densities per male P. leucopus were correlated with higher engorgement indices, suggesting that immature I. dammini feed better at higher densities. The feeding success of I. dammini on its preferred host species might be due to its adaptation to the immune and inflammatory reactions of the host.

Animals↗

Host genetics as a cause of overdispersion of parasites among hosts: how general a phenomenon?

In the white-footed mouse, Peromyscus leucopus, the tapeworm Hymenolepis citelli occurs at low (2-3%) prevalence in the field. We found that mature infections (i.e., with egg production) developed in up to 100% of hosts. In the laboratory, a majority of hosts lost their infection by 28 days postintubation. In wild mice infected in the laboratory and returned to the field, infections were more prolonged, with half of the mice still infected at 100 days postintubation. A majority of previously infected hosts resisted challenge infection. Our introduction of laboratory-infected mice into a natural population of hosts appeared to cause infections among previously uninfected mice, leading to an increase in the prevalence of tapeworm infection among mice not intubated. Although genetically based expulsion of tapeworms before maturity is important in causing low prevalence in a similar host-parasite system, such resistance cannot explain low prevalence in the present system. It appears that both heterogeneous distribution and rarity of intermediate hosts as well as short parasite lifespan contribute to low prevalence and overdispersion. Host-parasite dynamics of 2 very similar systems appear to differ markedly.

Animals↗

Vertical distribution of soil removed by four species of burrowing rodents in disturbed and undisturbed soils.

Burrow volumes were determined in disturbed and undisturbed soils for four species of rodents in southeastern Idaho. Comparisons were made between soil types for the average volume and the proportion of the total volume of soil excavated from 10-cm increments for each species, and the relative number of burrows and proportion of total soil removed from beneath the minimum thickness of soil covers over buried low-level radioactive wastes. Burrows of montane voles (Microtus montanus) and deer mice (Peromyscus maniculatus) rarely extended below 50 cm and neither volumes nor depths were influenced by soil disturbance. Townsend's ground squirrels (Spermophilus townsendii) had the deepest and most voluminous burrows that, along with Ord's kangaroo rat (Dipodomys ordii) burrows, were more prevalent beneath 50 cm in disturbed soils.

Animals↗

Safety and immunogenicity of a vaccine bait containing ERA strain of attenuated rabies virus.

Ninety percent of foxes fed commercial ERA vaccine in a specially designed bait developed rabies serum neutralizing antibodies. The vaccine bait did not cause clinical signs of rabies when consumed by foxes, raccoons, skunks, dogs, cats, cattle and monkeys. When presented, in the laboratory, to wild rodents of the species Microtus, Mus musculus and Peromyscus, the vaccine baits caused vaccine-induced rabies only in Mus musculus. Laboratory mice of the CD-1 and CLL strain were susceptible to vaccine-induced rabies; however, studies showed that transmission of virus to other animals did not occur. These studies suggest that the vaccine bait described could be useful in a rabies control program in areas where foxes and wild dogs are the principal vectors.

Animals↗

A comparison of three geographical isolates of Trichinella spiralis from the mid-Atlantic United States.

Three isolates of Trichinella spiralis (black bear: 41o50'N, 79o00'W, 1982; grey fox: 41o00'N, 76o00'W, 1982; domestic pig: 39o80'N, 75o30'W, 1983) from the mid-Atlantic United States were characterized in Crl: COBS CFW (SW) mice during 10 successive 40-day generations. Over 10 generations, the isolates differed as follows: worm position of the ursine isolate was significantly more posteriad compared to the porcine isolate; sex ratio of the vulpine isolate was significantly lower compared to the ursine and porcine isolates; females of the porcine isolate produced significantly more newborn larvae in vitro than the sylvatic isolates; both the larvae per gram (LPG) and reproductive capacity index (RCI) were significantly higher for the porcine isolate; and male worms of the vulpine isolate were significantly smaller than those of the porcine or ursine isolates. No differences were observed among isolates for percentage of inoculum recovered, length of female worms, or uterine length. The fecundity of the isolates in hamsters, gerbils, multimammate rats, Sprague-Dawley rats, and Peromyscus sp. was examined also. Based upon RCI and LPG, the porcine isolate was the most fecund in all hosts except gerbils. This higher fecundity was often manifest as increased morbidity and mortality in hosts infected with this isolate. The relationship between the virulent nature of the porcine isolate and the enhanced transmission of T. spiralis are discussed.

Animals↗

Genomic 5-methyldeoxycytidine decreases with age.

Significant losses of DNA 5-methyldeoxycytidine residues in old age could disrupt cellular gene expression and contribute to the physiological decline of the animal. Thus, the 5-methyldeoxycytidine content of DNAs, isolated from the tissues of two rodent species of various ages, were determined. Mus musculus lost DNA methylation sites at a rate of about 4.7 X 10(4) (approximately 0.012% of the newborn level)/month. Peromyscus leucopus lost DNA 5-methyldeoxycytidine residues at a rate of only 2.3 X 10(4) (approximately 0.006% of the newborn level)/month. Since P. leucopus generally live twice as long as M. musculus, the rate of loss of DNA 5-methyldeoxycytidine residues appears to be inversely related to life span. Similar losses in genomic 5-methyldeoxycytidine content were also observed to correlate with donor age in cultured normal human bronchial epithelial cells.

Aging↗

Chronic and acute trypanosomiasis in mice: a study by in vitro cloning of lymphohaematopoietic progenitors.

A semi-solid culture system was used to study the effects of trypanosome infection in two species of mice on the propagation of progenitor cells from the bone marrow and spleen. The deer mouse (Peromyscus maniculatus) survived infection with Trypanosoma (T.) equiperdum for more than 15 days. During the first 10 days there was inhibition of development of granulocyte-monocyte colonies from progenitor cells in the bone marrow. B-lymphocyte progenitors in the spleen showed increased activity, producing colonies 140-300% above normal control groups during the same period. - Conversely, all Balb/c mice infected with the trypanosomes died within 10 days with fulminating parasitemia and massive spleen enlargement. There was a general activation of progenitor cells; B-lymphocytes from the spleen and bone marrow and granulocyte-monocyte colonies from bone marrow, although this was not sustained more than 4 days after infection. - In chronically infected deer mice the pattern of response of the bone marrow and spleen progenitor cells was significantly different over successive weekly intervals. Periodicity of response in these organs was displayed by recurring waves of activation and depression of the progenitor cells. - Thus, there were significant differences in response patterns of deer mice and Balb/c mice to T. equiperdum infection which could be established by the behavior of host lymphohaematopoietic progenitor cells in culture. We therefore suggest that such in vitro cultures may be useful in assessment of the immune response to trypanosomiasis by the host and also for the study of the pathology of both chronic and acute trypanosomiasis.

Animals↗

Rodent models of the human isoniazid-acetylator polymorphism.

Inbred strains and subpopulations of rats, laboratory mice, and deer mice were examined for individual variation in the ability to metabolize several arylamines (p-aminobenzoic acid, sulfamethazine, aniline, alpha-naphthylamine, and aminofluorene) by N-acetylation. Individual differences within species were found to be dependent upon the tissue source of N-acetyltransferase activity and the acetyl acceptor employed. Long-Evans rats possessed about 2-fold more p-aminobenzoic acid N-acetyltransferase activity in blood and liver than Sprague-Dawley rats; no strain differences could be found with sulfamethazine. Nine strains of laboratory mice (Mus musculus) were found to have considerable liver p-aminobenzoic acid N-acetyltransferase activity but only slight activity towards sulfamethazine. No strain differences were apparent in regard to liver N-acetyltransferase activity. Blood p-aminobenzoic acid N-acetyltransferase activity was distinctly polymorphic in laboratory mice; of the nine strains tested, only A/J mice did not have this activity. Partially inbred deer mice (Peromyscus maniculatus) showed a narrower phenotypic range than random-bred stock from which they were obtained, which suggests the existence of distinct subpopulations with respect to N-acetylation capacity. Presumptive evidence for multiple forms of N-acetyltransferase in liver and blood was obtained through a study of substrate specificity.

Acetylation↗

Rodent housing units for field studies.

Rodent housing and nesting units were constructed from acrylic and provided with concrete tops. These hexagonal units, placed in a wooded field, were inhabited by deer mice (Peromyscus maniculatus). The use of the field housing units provided an effective method for observing free-ranging wild rodents.

Animal Population Groups↗

The IsTaR 1 serodeme of Trypanosoma brucei: development of a new serodeme.

An extensive serodeme of sequentially-isolated antigenic variants of African trypanosomes has been produced from both syringe-passaged and cyclically-transmitted Trypanosoma brucei of the IsTaR 1 clone derived from EATRO 164. The majority of the antigenic variants were isolated from chronically-infected deer mice (Peromyscus leucopus). The pattern of parasitemias during the course of infections initiated with syringe-passaged trypanosomes differed from those initiated with cyclically-transmitted trypanosomes. Trypanosome populations from syringe-passaged (192) and cyclically-transmitted (31) clones were each amplified by growth in lethally-irradiated mice and cryopreserved for retrospective analysis. Five clones derived from a single deer mouse during the first 44 days of infection, and 2 clones derived from an acutely-infected rat were established from these amplified populations. Homogeneous populations were grown in lethally-irradiated rats and mice for antigenic analysis purification of variant-specific glycoprotein. Six of the 7 clones were distinct variants by immunological criteria using antisera derived from whole cells or purified surface glycoproteins. Two clones, one derived from the acutely-infected rat, and the other from the first parasitemia in a chronic infection that was initiated with the former clone, were immunologically identical. Production of these clones established a well-defined serodeme that will allow detailed analysis of antigenic variation.

Animals↗