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Natural prevalence of infection with Ehrlichia (Cytoecetes) phagocytophila of Ixodes ricinus ticks in Scotland.

Ixodes ricinus nymphs and adults were collected from vegetation and from sheep at four sites in Scotland typical of areas endemic for tick-borne fever in sheep caused by infection with Ehrlichia (Cytoecetes) phagocytophila (Rickettsiales). The great majority of ticks examined was from woodland sites adjacent to sheep farms where there was a high probability of them feeding on roe deer (Capreolus capreolus) in a non-domestic focus of infestation and infection. Ticks were examined for infection by five methods. Batches of ticks were examined either by feeding on susceptible sheep or by feeding on rabbits and then prepared as stabilate which was inoculated into susceptible sheep. The sheep were monitored for clinical signs of tick borne fever. Batches of ticks were examined by polymerase chain reaction for Ehrlichia phagocytophila. Salivary glands were dissected out and stained by the Feulgen method to detect Ehrlichia masses, and were examined by indirect fluorescent antibody test. Each of the methods detected infection in ticks and the prevalence of infection in nymphs with the various methods ranged from >0.25% to 2.0%. Small samples of adults examined by Feulgen staining of salivary glands indicated infection prevalences of 2.1% in males and 1.6% in females. It is considered that these low infection prevalences may be typical of natural foci of infection where deer could be a major host of ticks and E. phagocytophila.

Animals↗

A comparison of the occurrence of borreliae in nymphal and adult Ixodes ricinus ticks.

The prevalence of borreliae in 209 nymphal and 251 adult Ixodes ricinus was investigated in two areas of southern Moravia, Czechoslovakia, using the dark-field and the Giemsa stained-smear techniques. The proportions of infected ticks were 3.8% in nymphs and 10.6% in adults of area A, while they were 29.1% in nymphs and 35.9% in adults of area B. The mean number of borreliae per tick was about 3 to 10 times greater in adult than in nymphal I. ricinus. The results indicate a significant role of nymphal I. ricinus in the ecology of Borrelia burgdorferi and in the epidemiology and epizootiology of Lyme borreliosis.

Animals↗

Prevalence of Borrelia burgdorferi s.I. in Ixodes ricinus ticks from Styria (Austria) and species identification by PCR-RFLP analysis.

A total of 1163 I. ricinus ticks were collected in 3 different regions (15 localities) in Styria (Austria) in June 1997 and examined for the presence of spirochetes by dark field microscopy. The mean infection rate was 20.8%. Among 310 adults, 24.2% were positive and among 853 nymphs, 19.6% were positive. All 15 collection areas were shown to harbour infected nymphs with a positivity rate ranging from 5.8% (3/52) to 32.1% (18/56). Isolation attempts in BSKII medium resulted in 29 isolates. Species identification by PCR-RFLP analysis revealed 16 strains of B. garinii, 10 strains of B. afzelii and 2 strains of B. burgdorferi s. s. One isolate showed a mixed population of B. garinii and B. afzelii. In two collection areas, all three major Borrelia species were shown to be present in the tick population.

Animals↗

Feeding of Boophilus microplus larvae on a partially defined medium through thin slices of cattle skin.

Larvae of the cattle tick Boophilus microplus will attach to thin (0-3--0-5 mm) slices of cattle skin and engorge on a partially defined medium at 35 degrees C. Forty-seven to 83 per cent of the larvae had engorged by 8 days, and 51--71 per cent of these moulted to nymphs. Tissue culture medium alone allowed little growth unless supplemented with dialysed, freeze dried bovine serum (7 per cent, w/v). This medium could be further difined by substituting purified bovine serum albumin (Cohn fraction V) for the dialysed bovine serum. In one experiment, nymphs of Haemaphysalis longicornis engorged and later moulted to adults. Skin slices were used fresh or after freeze-drying and storing at --25 degrees C. The possible uses of the culture method are discussed.

Animals↗

Population regulation in ticks: the role of acquired resistance in natural and unnatural hosts.

Attachment, engorgement and subsequent development of successive infestations of Ixodes trianguliceps larvae and nymphs on natural hosts, Apodemus sylvaticus, and unnatural hosts, laboratory mice, are compared. In laboratory mice, primary infestations above a threshold level of about 10 ticks elicit an immunological response which reduces, in a density-dependent manner, the rate of successful tick engorgement during subsequent infestations. In contrast, in A. sylvaticus successive infestations of larvae result in unchanged or slightly improved survival through to nymphs. The relevance of these results to the concept of host-parasite co-evolution and to tick population regulation is discussed.

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Preliminary characterization of the lipid and protein components of the protective surface membranes of a pentastomid Porocephalus crotali.

All instars of the pentastomid Porocephalus crotali, in the tissues of rat intermediate hosts and the lung of rattlesnake definitive hosts, are covered by a vesicular or stacked membranous secretory product which is synthesized in sub-parietal cells (SPC) and channelled to the cuticle via multitudinous chitin-lined ducts. In rats this enveloping foam of vesicles survives for the duration of a cuticle. We have purified and partially characterized the lipid and protein composition of lamellate droplets from SPC of infective nymphs. Lipids in the droplets comprised a mixture of neutral and polar lipid classes with cholesterol being the major neutral lipid and phosphatidylcholine the dominant polar lipid. In addition, Triton X-114 phase separation of the protein component of droplets partitioned these into an aqueous phase (with a major band at 60 kDa), a detergent phase with two hydrophobic polypeptides (24 and 16 kDa) and an insoluble pellet containing several minor proteins and major bands at 31 and 107 kDa. Western blotting, with rabbit anti-lamellate droplet antiserum, strongly label only the 60 kDa and the two hydrophobic proteins. Significantly no proteins at all label with serum from infected rats from 50-150 days post-infection, a finding endorsed by indirect fluorescent antibody tests on sectioned V-VII stage nymphs. Possible immunomodulatory functions of SPC-derived surface membranes are discussed and, in this regard, they are compared with pulmonary surfactant, a product of alveolar Type II cells which lines all vertebrate lungs.

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First report of Basidiolum fimbriatum since 1861, with comments on its development, occurrence, distribution and relationship with other fungi.

An obscure parasitic fungus, Basidiolum fimbriatum, was found on Amoebidium parasiticum (Amoebidiales) associated with Caenis sp. (mayfly) nymphs, during a survey of gut fungi (Trichomycetes) from a small stream in northeastern Kansas, USA. The hindguts of the nymphs harboured a species of Legeriomycetaceae and Paramoebidium sp. This is the first report of the ectocommensal protozoan, A. parasiticum, associated with the gills of Caenidae (Ephemeroptera), and of B. fimbriatum in the 142 years since its original documentation from Wiesbaden, Germany. B. fimbriatum is recorded from two midwestern USA states (Kansas and Iowa) and the morphological and developmental features of the parasite on its host are compared with Cienkowski's original observations and interpretation. B. fimbriatum is characterized as a parasitic fungus possessing merosporangia that from on a simple pyriform thallus that penetrates and consumes its host via a haustorial network. The hypothesis that B. fimbriatum is most closely related to members of the order Zoopagales sensu Benjamin (1979) is proposed. The importance of future collections and molecular-based phylogenetic approaches to place this parasitic fungus within a current system of classification are highlighted.

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Transmission of theileriosis in the traditional farming sector in the southern province of Zambia during 1995-1996.

The incidence of first contact with the protozoan Theileria parva was determined in three traditional cattle herds in the Southern Province of Zambia in 1995 and 1996. The majority of first contacts occurred during the dry season in June, July and August, at a time of nymphal activity and in the absence of Rhipicephalus appendiculatus adults, indicating that larva to nymph transmission plays a more prominent role than nymph to adult transmission under the prevailing conditions.

Animal Husbandry↗

Attraction of immature stages of the American dog tick (Dermacentor variabilis) to 2,6-dichlorophenol.

To determine whether 2,6-dichlorophenol is solely a sex pheromone, the response to it by the various stages of the American dog tick, Dermacentor variabilis, were compared. In contrast to adults, 2,6-dichlorophenol was attractive to unfed nymphs and to unfed larvae. Use of this chemical also prompted the expression of a novel type of 'feeding' posture behavior in adults. The overlap in attraction to other substituted phenols plus the lack of functional value of this response for larvae and nymphs rules out the possibility that 2,6-dichlorophenol is a general attractant. However, 2,6-dichlorophenol likely plays a dual role as an attachment stimulant in the adult tick.

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Cold-hardiness of Dermacentor marginatus (Acari: Ixodidae).

The cold-hardiness of Dermacentor marginatus using laboratory-reared offspring of ticks collected in Germany was characterized. Investigations of unfed stages revealed that adult ticks suffered 50% mortality at -10 degrees C after 4-5 months, but larvae and nymphs suffered mortality within few days, whereas -15 degrees C was lethal for all stages within a very short period. Larval hatch and moulting of engorged larvae and nymphs did not occur at < or = 10 degrees C. Embryonic development of eggs with larval hatch was considerably reduced by exposure of eggs to < or = 10 degrees C. Engorged females did not lay eggs at < or = 10 degrees C, the oviposition capability, however, persisted over 6 months at 10 degrees C, 5 months at 5 degrees C, 3 months at 0 degree C and 2 months at -10 degrees C without substantial decrease of the oviposition capacity or reduction of viable eggs. These results present evidence that unfed adult ticks are the ecoepidemiologically most effective stages, which are capable to tolerate long and extremely cold winters without substantial impairment of the population density. It is also considered that engorged females interrupt their oviposition at low and subzero temperatures delaying it for months and so contribute in bypassing winter conditions. None of the stages survived supercooling indicating that D. marginatus is freeze intolerant. Mean supercooling point (SCP) ranged between -26 degrees C in eggs and -12, 6 degrees C in engorged females. Compared with eggs, the SCP of the other stages was significantly higher. In conclusion, the SCP is considered to have no predictive value in the context with cold-hardiness.

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Interstadial and infestation level-dependent variation in the transmission efficiency of Borrelia burgdorferi from mice to Ixodes ricinus ticks.

The efficiency with which the spirochaete Borrelia burgdorferi sensu stricto was transmitted from laboratory mice to larval and nymphal Ixodes ricinus ticks was assessed, using the polymerase chain reaction. The transmission efficiency to nymphs was significantly greater than to larvae when both fed together on the same host. Increased tick infestation levels of mice were correlated with significantly greater engorgement weights and higher B. burgdorferi transmission coefficients from mice to nymphs. These observations indicate that both the feeding success of ticks and the transmission coefficients from host to tick may be influenced by the tick infestation level of an infected host. The infestation level and the relative numbers of each life stage of the tick are factors which should be considered in the design of transmission experiments.

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The pupal specifier broad directs progressive morphogenesis in a direct-developing insect.

A key regulatory gene in metamorphosing (holometabolous) insect life histories is the transcription factor broad (br), which specifies pupal development. To determine the role of br in a direct-developing (hemimetabolous) insect that lacks a pupal stage, we cloned br from the milkweed bug, Oncopeltus fasciatus (Of'br). We find that, unlike metamorphosing insects, in which br expression is restricted to the larval-pupal transition, Of'br mRNA is expressed during embryonic development and is maintained at each nymphal molt but then disappears at the molt to the adult. Induction of a supernumerary nymphal stage with a juvenile hormone (JH) mimic prevented the disappearance of br mRNA. In contrast, induction of a precocious adult molt by application of precocene II to third-stage nymphs caused a loss of br mRNA at the precocious adult molt. Thus, JH is necessary to maintain br expression during the nymphal stages. Injection of Of'br dsRNA into either early third- or fourth-stage nymphs caused a repetition of stage-specific pigmentation patterns and prevented the normal anisometric growth of the wing pads without affecting isometric growth or molting. Therefore, br is necessary for the mutable (heteromorphic) changes that occur during hemimetabolous development. Our results suggest that metamorphosis in insects arose as expression of br, which conveys competence for change, became restricted to one postembryonic instar. After this shift in br expression, the progressive changes that occur within the nymphal series in basal insects became compressed to the one short period of morphogenesis seen in the larva-to-pupa transition of holometabolous insects.

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Effects of acorn production and mouse abundance on abundance and Borrelia burgdorferi infection prevalence of nymphal Ixodes scapularis ticks.

Risk of exposure to Lyme disease is a function of the local abundance of nymphal Ixodes ticks that are infected with the etiological agent, the spirochete Borrelia burgdorferi. We monitored abundance of white-footed mice (the principal B. burgdorferi reservoir in the eastern and central United States) and acorns (a critical food resource for mice), and Ixodes scapularis ticks, as well as ambient temperature (cumulative growing degree days) and growing season precipitation, in a forested landscape of southeastern New York State from 1994 to 2000. We found that acorn production in autumn strongly influenced abundance of white-footed mice the following summer and that abundance of mice in summer, when larval ticks are active, influenced the abundance of infected nymphs the following year. Consequently, the abundance of infected nymphal ticks can be predicted from acorn production 1.75 years earlier. Monitoring of natural fluctuations in acorn production thus supports results of prior acorn addition experiments that were conducted at small spatial scales. Growing degree days and precipitation either had no significant effect on density of nymphs or marginally increased the explanatory power of models that included acorns or mouse density as independent variables. We conclude that, at our study site in New York, the risk of human exposure to Lyme disease is affected by mouse density in the prior year and by acorn production 2 years previously.

Animal Feed↗

Blood-meal analysis for the identification of reservoir hosts of tick-borne pathogens in Ireland.

The results of analysis of blood-meal remnants in unfed nymphs, despite relatively low detection levels (49.4%, n = 322), support the conclusion from an earlier study that small rodents are relatively unimportant as reservoir hosts of B. burgdorferi s.l. in this particular area, and suggest that songbirds (Passeriformes) are the most significant hosts in this respect. Tick (Ixodes ricinus) abundance was greater in the present study, but the overall Borrelia burgdorferi s.l.-infection prevalence of nymphal ticks was the same (12.2%), and the relative proportions of the various Borrelia burgdorferi s.l. genospecies were similar. B. garinii and B. valaisiana were the most frequent, B. burgdorferi s.s the least frequent, and B. afzelii of intermediate frequency. An unusually high proportion of nymphs (39%) with multiple infections of different B. burgdorferi genospecies was detected, and Borrelia spp. related to relapsing-fever spirochetes were detected in Ireland for the first time. The results of the present study contribute to the validation of blood-meal analysis as a means of determining the host origin of certain pathogens in unfed questing ticks, and raise some questions concerning the extent of B. burgdorferi s.l. host specificity.

Animals↗

Toxicity of abamectin to cockroaches (Dictyoptera: Blattellidae, Blattidae).

Abamectin was fed to German cockroaches, Blattella germanica (L.), in non-choice tests. LT50s and LC50s were estimated by probit analysis. The LT50s for the German cockroach ranged from 4.4 to 1.7 d for males, from 9.0 to 2.4 d for females, and from 4.4 to 1.6 d for nymphs for bait concentrations of abamectin between 0.0025 and 0.0500%. The LC50s of abamectin were 0.0110 and 0.0040% from males, 0.0240 and 0.0090% for females, and 0.0200 and 0.0080% for nymphs at 3 and 6 d, respectively. The LT50 values of 0.0550% abamectin bait were 3.4, 3.4, 2.4, 7.5, 2.9, and 4.5 d for Periplaneta americana (L.), P. fuliginosa (Serville), P. brunnea Burmeister, P. australasiae (F.), Blatta orientalis L., and Supella longipalpa (Serville). Although the bait was effective against various cockroach species, time to death for the larger species was longer than for the German cockroach. In preference tests in which male German cockroaches were allowed to feed on rat chow or abamectin bait, all died within 5 d of exposure to abamectin bait. Abamectin bait consumption was not significantly lower than that of untreated rat chow. Arena tests with 0.0550% abamectin bait resulted in 31-75% mortality of German cockroaches after 9 d, with most control being achieved by treating harborages with the bait. The hydramethylnon standard resulted in 65% mortality after 9 d.

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Diel activity cycles in nymphal stadia of the German cockroach (Dictyoptera: Blattellidae).

Computerized moving-image analysis was used to determine movement behavior of individual German cockroach, Blattella germanica (L.), nymphs (second and fifth instars) between food, water, and harborage resource sites. Both second and fifth instars (n = 10) were examined for the entire nymphal stadium, during which a 12:12 (L:D) photoperiod was maintained. In addition, corresponding resource consumption rates for these nymphal stages also were determined. Both nymphal stages exhibited a pattern of high activity for the first half of the nymphal stadium, especially during each scotophase. For the last third of the stadium, the nymphs remained continuously in the harborage, moved very little, and consumed little or no resources. We discuss implications of these findings regarding control, future research, and possible hormonal behavior regulation.

Animals↗

Nutrient uptake of rice roots in response to infestation of Nilaparvata lugens (Stål) (Homoptera: Delphacidae).

This study investigated the uptake of three macronutrients, including nitrogen (N), phosphorus (P), and potassium (K), by rice roots in response to different infestation levels of Nilaparvata lugens (Stål) (Homoptera: Delphacidae). Hydroponics experiments were conducted on the rice variety 'Zhendao 2' (moderately resistant to Tryporyza incertulas, Lepidoptera: Pyralidae) and the variety 'Xiushui 63' (susceptible to N. lugens). In 'Zhendao 2', N. lugens infestation significantly influenced the uptake of P and K but not N, with P and K uptake decreasing as the duration of N. lugens infestation increased. In 'Xiushui 63', N. lugens infestation influenced N, P, and K uptake to a different degree, depending on the infestation level, in which infestation for 2, 4, 6, and 8 d at a density of 60 nymphs did not affect N uptake, but such infestation levels significantly influenced the uptake of P and K. After the removal of N. lugens from rice plants, the N uptake recovered from infestation faster than that for P and K in the variety 'Zhendao 2', whereas the recovery rate of K uptake was faster than that for N and P in the variety 'Xiushui 63'. The recovery rate of the nutrient uptake was negatively correlated to the density and duration of infestation. The experimental results demonstrated that N, P, and K uptake of rice roots were largely not influenced by N. lugens infestation when the pest density was controlled below 15 nymphs per hill. This infestation level was in agreement with the proposed economic thresholds for control measures against the N. lugens infestation on rice plants.

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Hot water immersion for surface disinfestation of Maconellicoccus hirsutus (Homoptera: Pseudococcidae).

Mealybug Maconellicoccus hirsutus (Green) adults, nymphs, crawlers, and eggs were tested for their susceptibility to hot water immersion at 47, 48, and 49 degrees C. Eggs inside ovisacs were found most tolerant with prolonged survival compared with other stages at all temperatures. Ovisacs required an average of 1.38, 1.46, and 1.62 times longer treatment duration than adults, nymphs, and crawlers, respectively, for 99.9% predicted mortality at 47, 48, and 49 degrees C. Lethal time estimations were calculated from inverse predictions of regressions derived from logit-transformed data as well as those created using a kinetic model. LT 99.9 estimations were 47.0, 21.2, and 11.9 min at 47, 48, and 49 degrees C, respectively, by using regressions with logit transformations. The kinetic model predictions were 43.9, 19.6, and 11.1 min at 47, 48, and 49 degrees C, respectively. During the study no emergence from eggs inside ovisacs was found after treatments of 52, 24, and 14 min at 47, 48, and 49 degrees C, respectively. Results from this study provide efficacious temperature-time treatments.

Animals↗