Shared epitopes between the soluble proteins of Hypoderma lineatum and Hypoderma bovis first instars.

Cattle are known to acquire immunological resistance to hypodermyiasis by repeated exposure to both species of cattle grubs, Hypoderma lineatum (Villers) and Hypoderma bovis (L.). Vaccination of cattle with purified proteins of H. lineatum, particularly hypodermin A, is known to protect cattle against hypodermyiasis by this species. The development of a protective recombinant vaccine against both species using hypodermin A isolated from H. lineatum would require that immunological epitopes be shared by complementary proteins in H. bovis. The purpose of this study was to investigate the soluble proteins of H. bovis first-instars for shared epitopes with H. lineatum. Soluble H. lineatum and H. bovis first-instar larval proteins were resolved by nondenaturing polyacrylamide electrophoresis, blotted onto nitrocellulose paper, and probed with selected polyclonal cow, polyclonal rabbit, and mouse monoclonal antisera. Considerable cross-reactivity was demonstrated by antibodies in the serum of an H. lineatum-infested cow as 6 of 10 resolved H. bovis proteins were bound by the antibodies. The most common shared epitope(s) was associated with hypodermin C, a collagenolytic protease. Hypodermin A shared epitope(s) were noted on 1 prominent H. bovis band (HB1-2). Hypodermin B, a prominent protein in H. lineatum, did not appear to share epitopes with H. bovis proteins. Shared epitopes between H. bovis proteins and hypodermins A and C of H. lineatum would suggest that cross-protection of cattle against H. bovis can be expected by vaccination with recombinant proteins of H. lineatum.     

Acoustic estimation of infestations and population densities of white grubs (Coleoptera: Scarabaeidae) in turfgrass

Incidental sounds produced by Phyllophaga crinita (Burmeister) and Cyclocephala lurida (Bland) (Coleoptera: Scarabaeidae) white grubs were monitored with single- and multiple-sensor acoustic detection systems in turf fields and golf course fairways in Texas. The maximum detection range of an individual acoustic sensor was measured in a greenhouse as approximately the area enclosed in a 26.5-cm-diameter perimeter (552 cm2). A single-sensor acoustic system was used to rate the likelihood of white grub infestation at monitored sites, and a four-sensor array was used to count the numbers of white grubs at sites where infestations were identified. White grub population densities were acoustically estimated by dividing the estimated numbers of white grubs by the area of the detection range. For comparisons with acoustic monitoring methods, infestations were assessed also by examining 10-cm-diameter soil cores collected with a standard golf cup-cutter. Both acoustic and cup-cutter assessments of infestation and estimates of white grub population densities were verified by excavation and sifting of the soil around the sensors after each site was monitored. The single-sensor acoustic method was more successful in assessing infestations at a recording site than was the cup-cutter method, possibly because the detection range was larger than the area of the soil core. White grubs were recovered from >90% of monitored sites rated at medium or high likelihood of infestation. Infestations were successfully identified at 23 of the 24 sites where white grubs were recovered at densities >50/m2, the threshold for economic damage. The four-sensor array yielded the most accurate estimates of the numbers of white grubs in the detection range, enabling reliable, nondestructive estimation of white grub population densities. However, tests with the array took longer and were more difficult to perform than tests with the single sensor.     

A third species of Hypoderma (Diptera: Oestridae) affecting cattle and yaks in China: molecular and morphological evidence

Cattle and yak hypodermosis in China is caused by Hypoderma bovis and H. lineatum, with a prevalence reaching up to 98-100% of the animals and maximum intensities exceeding 400 warbles for each animal. A third species, H. sinense, is also considered by Chinese researchers to affect livestock. The molecular characterization of the most variable region of the mitochondrial cytochrome oxidase I gene and of the ribosomal 28S gene has been performed for the third-stage larvae collected from cattle and yaks in China and identified (on the basis of the spinulation on the ventral side of the 10th segment) as H. bovis, H. lineatum, and H. sinense. Amplicons were digested with the HinfI and BfaI restriction enzymes, which provided diagnostic profiles to simultaneously differentiate the 3 Hypoderma species. Third-stage larvae of H. sinense were also examined by scanning electron microscopy, which revealed proper morphological characteristics different from those of H. bovis and H. lineatum. The molecular and morphological evidence herein reported support the existence of a third species of Hypoderma affecting cattle and yaks in China, and the results provide new tools for unequivocal identification of this species and present key components for the evaluation of its endogenous cycle and pathogenicity in animals and humans.     

Comparative scanning electron microscopy of third-instar Hypoderma spp. (Diptera: Oestridae)

Scanning electron microscope study of third-instar larvae of four species of Hypoderma revealed differences among species in the pattern of spination, spine morphology and morphology of the spiracular plates. These observations identify characters that enable the differentiation of Hypoderma actaeon and H. diana, parasitizing red deer ( Cervus elaphus ) in Europe, and provide additional characters for differentiating H. bovis and H. lineatum parasitizing cattle.     

Comparison of tall fescue (Cyperales: Gramineae) to other cool-season turfgrasses for tolerance to European chafer (Coleoptera: Scarabaeidae)

Three cultivars of tall fescue, Festuca arundinacea Schreb., were compared with three cultivars each of fine fescue (Festuca spp.), Kentucky bluegrass (Poa pratensis L.), and perennial ryegrass (Lolium perenne L.) to evaluate tolerance to root-feeding by European chafer grubs, Rhizotrogus majalis (Razoumowsky). Potted turfgrasses were infested with initial densities equivalent to 33 or 66 grubs per 0.1 m2 on 19 August 2000. More grubs were added in late September and October, bringing the total to 66 or 143 grubs per 0.1 m2. Plant growth, root loss, weight gain, and survival of grubs were measured. The experiment was repeated in fall of 2001 with an initial density of 66 grubs per 0.1 m2. The proportion of root mass lost as a result of grub feeding was a function of turf species, root growth, grub survival, and grub growth during the test. Grubs gained the most weight and consumed the most roots when feeding on fine fescue. Fine fescue suffered the greatest percentage of root loss in 2000, despite having the most rapid root growth and largest mass in control pots. Cultivars of tall fescue appeared to be the most tolerant of grub feeding, having the smallest reduction in root mass in both years. Data from fine fescue, Kentucky bluegrass, and perennial ryegrass cultivars were not as consistent as tall fescue, because for some cultivars root growth and grub survival were different between years. We also found that grubs increased in mass by 20% when the mass of available roots was doubled.     

Myiasis caused by Oestridae: serological and molecular diagnosis

Myiasis-causing Oestridae (bot flies) infect several animal species world-wide, from palaearctic to subtropical/tropical areas. Oestrids affect livestock production causing abortion, reduced milk production, losses in weight and fertility, poor hide quality and an impairment of the host's immune system. In the last few years much research has been carried out on the immunology of these infestations, in order to acquire efficient and reliable diagnostic serological tools; the genome of the different species of Oestridae has been studied to further their molecular identification, taxonomy and phylogenesis. The immunodiagnostic methods for many myiasis causing Oestrids have proven to be a viable alternative to the clinical parasitological examination or the post-mortem examination. Numerous serological tests have been developed for the diagnosis of bovine hypodermosis caused by Hypoderma bovis and Hypoderma lineatum, and ELISAs using larval hypodermin C as the antigen are currently used on serum, individual and pooled milk samples to detect the presence of circulating anti-Hypoderma antibodies. In Italy the best period to sample the animals is November-January, since it is in this period that the antibody kinetics of the animals reaches a peak. Recently the efficacy of the ELISA test on pasteurized milk samples has been demonstrated, allowing the diagnosis of bovine hypodermosis also in areas where there is no information on the presence of the disease and the sampling of the animals is laborious. The cross-reactivity between Przhevalskiana silenus antigens and anti-Hypoderma antibodies led to assessing the usefulness of a simple and cost-effective ELISA for the diagnosis of goat warble fly infection. In particular, it has been demonstrated that infected goats display an antibody peak in November-December in blood and milk, thus making this period suitable for sampling. Although no extensive data is available on the immunology of sheep and goat oestrosis caused by Oestrus ovis, the efficacy of ELISA has been demonstrated by correlating serological results with clinical post-mortem examinations. No immunological techniques are currently used to diagnose gasterophilosis of equids and only one study reports the efficacy of ELISA for detecting anti-Gasterophilus antibodies in infected equids. Several studies have been conducted into the molecular characterization of the mitochondrial DNA (mtDNA)--in particular of the gene encoding for the cytochrome oxidase I (COI)--for many free-living and parasitic arthropods for diagnostic, taxonomic and phylogenetic purposes. As regards Oestridae causing myiasis, the first study features a PCR-RFLP assay of the most common Italian species (i.e. H. bovis, H. lineatum, Gasterophilus intestinalis, P. silenus, O. ovis), which showed clear genetic differences among the genera examined, but no inter-specific variation between the two species of Hypoderma considered. The molecular characterization of the most variable region of the COI gene (encoding for the region from E4 to the terminal COOH) was able to clearly differentiate H. bovis and H. lineatum. The E4-COOH region of the COI gene has been characterized for 18 oestrid species and from a taxonomical point of view, molecular data confirm the morphological classification, with the examined species divided into four subfamilies. New insights have also been gained on the molecular differentiation of the most common species of Hypoderma (i.e. H. bovis, H. lineatum, Hypoderma actaeon, Hypoderma diana and Hypoderma tarandi) and, in particular, the restriction enzyme BfaI, provides a diagnostic profile that can be used to simultaneously differentiate all the species examined. The characterization of the E4-COOH COI gene and the hypervariable region of the gene encoding for the ribosomal Isu revealed the identity of Hypoderma sinense as a new species, infecting cattle and yaks in China. Finally, the molecular analysis of the same mitochondrial and ribosomal regions showed that P. silenus, Przhevalskiana aegagri and Przhevalskiana crossii are morphotypes of the same species.     

Natural regulation of white grub (Holotrichia sp: Scarabidae) by birds in agroecosystem

The white grub (Holotrichia sp: Scarabidae) is an important subterranean pest damaging root systems of several crops. Experiments conducted during I985 and I986 showed that at least I4 species of birds picked up the grubs exposed during ploughing operation. The important bird predators were mynasAcridotheres tristis (Linnaeus) andAcridotheres ginginianus (Latham), crowsCorvus splendens (Vieillot),Corvus macrorhynchos (Sykes), drongoDicrurus adsimilis (Hodgson) and cattle egretBubulcus ibis. The birds were found to reduce 45 to 65% grub population during 3 subsequent ploughings. The plant stand of second crop raised in bird exposed field was higher in experimental plot compared to the control. The number of birds attracted to the plough was not consistent with the density of grubs exposed but oh many extraneous factors. Factors affecting the extent of bird predation were presence of insectivorous birds in the surroundings, proximity to their breeding sites and timing of ploughing. White grub control by birds is economically cheaper and environmentally safe compared to the chemical control.     

在藏羚羊上发现的中国第四种皮蝇

对于从可可西里藏羚羊上采集到的藏羚羊皮蝇Hypoderma sp.3龄幼虫进行了形态学观察,发现其在伪头、第10腹节上的刺、气门板形态上与中国现有记录的3种皮蝇（牛皮蝇、纹皮蝇和中华皮蝇）有着明显的区别。对藏羚羊皮蝇的线粒体COI 基因种属特异性序列研究表明,其与牛皮蝇、纹皮蝇和中华皮蝇对应序列的相似性分别为88.1％,87.8％和88.8％。据此认为采自藏羚羊的此种皮蝇可能为中国境内又一皮蝇新种或新记录种,为中国第四种皮蝇。     

Field efficacy of entomopathogenic nematodes against the beetle Maladera matrida (Coleoptera: Scarabaeidae)

Single, double and triple releases of the entomopathogenic nematode Heterorhabditis bacteriophora Poinar, reduced the population of the beetle Maladera matrida Argaman, infesting peanuts (’Shulamit’ cv.) by 70, 75 and 93% respectively in microplot tests. Simultaneous and late (2 weeks after infestation) applications reduced beetle numbers by 63 and 79% respectively, in the microplots, while early application (2 weeks prior to infestation) did not reduce the beetle population. In a field trial, reductions in insect population and damage to the crop were achieved by early treatment with the nematode as well as by Heptachlor, leading to reductions in the insect population of 60 and 90% respectively, when recorded 4 weeks after nematode application. However, the nematode treatment did not maintain its effectiveness for a longer period and pest damage increased to the same level as the untreated control after 7 weeks. When the nematodes were applied at different concentrations (0.25–1.0 x 10⁶ infective juveniles (IJs) m^‐2) their effectiveness was not related to the concentration level. The only significant (P < 0.05) reduction in insect levels was recorded in the treatment with 0.5 X 10⁶ IJs m^‐2. In a second field trial, both H. bacteriophora and Steinernema glaseri reduced insect populations significantly (P < 0.05) by approximately 50% in comparison to the control. In the third trial, treatment with H. bacteriophora resulted in a decrease in insect population of 90% while treatment with S. carpocapsae reduced the grub numbers by 40% in comparison to the control. A differential susceptibility of various grub developmental stages was recorded in the field. The small grubs (I‐4 mm long, lst‐2nd larval stage) were not affected by the nematode treatments while the numbers of medium and large size grubs were reduced by 2‐ and 3‐fold respectively in the various tests. Nematodes were recovered by ‘nematode traps’ containing Galleria mellonella larvae from treated field plots 78 days after application. The implications of the results from the present studies on the use of entomopathogenic nematodes are discussed in relation to the development of an integrated pest management programme.     

A model for estimating abundance of cattle grub (Diptera: Oestridae) from the proportion of uninfested cattle as determined by serology*

A model is presented for determining the abundance of cattle grubs in the backs of calves from the proportion of uninfested calves in a herd. The distribution of grubs in calves' backs was compared with the negative binomial, but no relationships were found among the distribution parameters, suggesting that the negative binomial is an inappropriate choice for the basis of a sampling model. The relationship between the mean number of grubs per animal (mean), variance (delta 2), and proportion of uninfested calves (p0) in a herd was determined and used as the basis for the sampling model. The relationship between p0 and p0e [determined using serology (ELISA)] was evaluated. The variance of estimates of mean grubs per animal based on the regression model and uncertainty due to using p0e as an estimate of p0 was examined. A test of the model indicated that p0e could be used to obtain a reliable estimate of mean grubs per animal and that the method would be applicable for monitoring grub populations, assessing chemical control programmes, and determining release rates of sterile insects for control.     

Impact of four insecticides on Japanese beetle (Coleoptera: Scarabaeidae) egg predators and white grubs in turfgrass

Field experiments were conducted to measure the effects of four commonly used turfgrass insecticides (isofenphos, diazinon, imidacloprid, halofenozide) on white grubs (Coleoptera: Scarabaeidae) and ant predators of white grub eggs. Ant populations were measured over time with canned tuna, whereas predation by the ants was measured with artificially placed Japanese beetle, Popillia japonica Newman, eggs. The effectiveness of each insecticide at controlling Japanese beetle grubs, when applied at different times during the growing season, also was measured. Isofenphos and diazinon significantly reduced both ant numbers and white grub egg predation, whereas imidacloprid and one halofenozide treatment did not significantly impact either measurement. A second halofenozide treatment significantly reduced white grub egg predation. Isofenphos and diazinon were ineffective at controlling Japanese beetle grubs when applied in June but were highly efficacious when applied in August. Evidence of enhanced biodegradation was found in plots that received both June and August applications of diazinon. Both June and August applications of imidacloprid and halofenozide provided good control of white grubs.     

Cattle lice (Anoplura, Mallophaga) in New York: seasonal population changes, effects of housing type on infestations of calves, and sampling efficiency

A year-long survey was made of commercial dairy herds in New York for cattle lice (Anoplura and Mallophaga). All herds were infested with lice. The cattle bitting louse, Bovicola bovis (L.), accounted for about 90% of the observed infestations; infestations of mature animals were most common during the winter months, especially March (26% infested); and infestation rates of calves were high (30-45%) from January through June. Cows that were housed in stanchion barns were about twice as likely to be infested (24.7%) as were those in free stalls (11.1%). Calves housed in individual outdoor hutches had substantially lower infestation rates (4.5%) than calves maintained inside barns in collective stalls and pens (46.0%).     

Bovine babesiosis: failure to induce interferon gamma production in response to Babesia bovis antigens in cattle.

The immune response to Babesia bovis infection or vaccination was evaluated by measuring antibody and interferon gamma (IFN-gamma) production to protective recombinant and crude native B. bovis antigens. Cells from vaccinated or infected cattle failed to produce detectable IFN-gamma when stimulated with B. bovis antigens in vitro. In contrast, antibody was induced by protective recombinant B. bovis antigens. These findings are consistent with the argument that immunity to B. bovis infection is correlated most strongly with humoral rather than cell-mediated immune responses.     

Incidence of turf-damaging white grubs (Coleoptera: Scarabaeidae) and associated pathogens and parasitoids on Kentucky golf courses

Root-feeding grubs (Coleoptera: Scarabaeidae) were sampled from damaged areas of 61 irrigated roughs on 32 Kentucky golf courses to determine species composition and natural enemy incidence, the first such survey in the United States' transitional turfgrass climatic zone. Masked chafers (Cyclocephala lurida Bland and C. borealis Arrow) and Japanese beetle (Popillia japonica Newman) accounted for ≈73 and 26% of grubs found in an autumn survey, with Cyclocephala spp. predominating at most sites, although mixed infestations were common. Only a few Phyllophaga spp., and no exotic species other than P. japonica were found. Cyclocephala spp. also predominated in seasonal and statewide surveys regardless of whether a course had cool- or warm-season grass fairways. Pathogenic bacteria, Paenibacillus and Serratia spp., and the autumn-active parasitoid Tiphia pygidialis Allen were the main enemies associated with Cyclocephala spp. Predominant enemies of P. japonica were Paenibacillus, Serratia, and Metarhizium spp. in autumn, and eugregarines, Stictospora sp. (probably S. villani Hays and Clopton) and Tiphia vernalis Rohwer in spring. Entomopathogenic nematodes and the microsporidian Ovavesicula popilliae Andreadis & Hanula were nearly absent in our samples. No predictive relationships were found between soil parameters and proportionate abundance of Cyclocephala or P. japonica, or with natural enemy incidence at particular sites. Although incidence of individual enemies was generally low (<20%; often <5%) in these point-in-time surveys, collectively and over their hosts' prolonged development they may take a significant toll on grub populations.     

Lack of interactions between fire ant control products and white grubs (Coleoptera: Scarabaeidae) in turfgrass

Insecticides are widely used to manage turfgrass pest such as white grubs (Coleoptera: Scarabaeidae). Red imported fire ants, Solenopsis invicta (Buren) are important predators and pests in managed turfgrass. We tested the susceptibility of white grub life stages (adults, egg, and larvae) to predation by S. invicta and determined if insecticides applied for control of S. invicta would result in locally greater white grub populations. Field trials over 2 yr evaluated bifenthrin, fipronil, and hydramethylnon applied to large and small scale turfgrass plots for impacts on fire ant foraging and white grub populations. Coincident with these trials, adults, larvae, and eggs of common scarab species were evaluated for susceptibility to predation by S. invicta under field conditions. Field trials with insecticides failed to show a significant increase in white grub populations resulting from treatment of turfgrass for fire ants. This, in part, may be because of a lack of predation of S. invicta on adult and larval scarabs. Egg predation was greatest at 70% but < 20% of adults and larvae were attacked in a 24 h test. Contrary to other studies, results presented here suggest that fire ants and fire ant control products applied to turfgrass have a minimal impact on white grub populations.     

Development and decline of serum antibody activity toPsoroptes ovis antigens in infested cattle in an endemic and nonendemic scabies area of texas

The serum antibody activity toPsoroptes ovis antigens, as determined by enzyme-linked immunosorbent assay (ELISA), was monitored during the development and decline phase of scabies in cattle herds under feedlot conditions in an endemic (Bushland) and a nonendemic (Kerrville) scabies area of Texas. The infestations were initiated by exposing six calves in each of the herds toP. ovis and allowing the infestations to spread to other members of the herd. The cattle in the endemic area developed more lesions and a higher incidence of the disease than did the cattle in the nonendemic area; also, there were more cattle with specific serum antibody activity in the Bushland herd than in the Kerrville herd at the peak of the infestations (late January-early February). As infestations declined because of summer latency, the number of calves with specific serum antibody activity declined in both herds. Specific serum antibody activity disappeared in the Kerrville cattle whenP. ovis was no longer detected. However, specific serum antibody activity was still detected in some of the Bushland cattle in late summer (August) and fall (October) examinations, when low numbers ofP. ovis were detected. These results indicate that the ELISA would be very useful in detection of specific serum antibody activity in lightly infested calves, especially during the developmental and decline phases of the disease.     

EXPERIMENTS IN THE SUDAN GEZIRA ON CONTROL OF WILT OF DOLICHOS BEAN (DOLICHOS LABLAB) ASSOCIATED WITH ATTACK BY COCKCHAFER GRUBS (SCHIZONYCHA SP.)

Wilt of dolichos bean ( Dolichos lablab ) in the Sudan Gezira appeared to be due primarily to cockchafer grubs ( Schizonycha sp.) in the soil attacking the hypocotyls or roots of plants up to about 6 weeks after sowing. Many wilted plants also showed symptoms of ashy stem blight ( Macrophomina phaseoli ), but in these the fungus was probably a secondary invader rotting roots weakened or damaged by unfavourable soil conditions or insects. Wilt was often severe in dolichos sown on land cropped to sorghum ( Sorghum vulgare ) in the preceding season, sorghum roots in the soil harbouring the grubs. Under moderate grub attack seed dressings containing organomercurial and γ-benzene hexachloride (BHC) gave satisfactory-protection at 0.089%γ-BHC/seed but were inadequate when wilt was severe. Dieldrin and aldrin at 0.044 or 0.089%/seed gave excellent protection in all experiments, but their performance under exceptionally severe wilt conditions has yet to be tested, as also the relative efficiencies of seed treatment and soil treatment under such conditions. Fungicide-insecticide seed treatment also reduced preemergence rotting of germinating seeds due to grub attack.
Wilt rarely occurred in dolichos planted on land fallow in the preceding season, but, even in the apparent absence of wilt, seed treatment often appreciably improved emergence, plant populations, growth and yields. These effects possibly resulted from control of root damage by soil insects, such damage reducing growth and yields but not sufficiently severely to cause wilting. Pending further investigation a powder seed dressing containing organomercurial plus 20% dieldrin, and applied at 1:450 by weight to seed (about 5–6 g./acre of dieldrin), is recommended for dolichos bean in the Gezira.     

Immunological responses of rabbits artificially infested with the cattle grubs Hypoderma bovis (L.) and H. Lineatum (De Vill.) (Diptera: Oestridae)

Boulard Chantal and Weintraub J. 1973. Immunological responses of rabbits artificially infested with the cattle grubs Hypoderma bovis (L.) and H. lineatum (De Vill.) (Diptera : Oestridae). International Journal for Parasitology3: 379–386. Immunological responses against first-instar larvae of Hypoderma bovis (L.) and H. lineatum (De Vill.) were traced in artificially infested rabbits in which larvae grew normally in the first instar but did not survive to the second instar. Passive haemagglutination demonstrated a rapid rise in antibody titres during the first 2 months to maximum levels that persisted until about 200 days of the infestation. Immunoelectrophoretic analyses of the sera demonstrated that larval metabolic products were more important than breakdown products of dead larvae in stimulating the production of antibodies. Larval collagenase, which seemed to induce the most significant immune response, was inhibited by homologous antibodies in the serum of a hyperimmunized rabbit. The implications of these results for infestations in the normal bovine host were discussed, with emphasis on the need to identify antigenic fractions other than collagenase, which by itself had not produced total control of larvae in previous vaccination tests.     

Scanning electron microscopy of the posterior spiracles of cattle grubs Hypoderma bovis and Hypoderma lineatum

Posterior spiracles of newly hatched first instar larvae of Hypoderma bovis (L.) and H. lineatum (DeVill.) consist of two pairs of spiracular openings. Each pair is surrounded by a rima bearing three spines. Posterior spiracles of second instar larvae are composed of a pair of medial ecdysial scars bounded laterally by spiracular plates. H. bovis spiracular plates have twenty-nine to forty openings, each surrounded by a slightly raised rima. H. lineatum spiracular plates have eighteen to twenty-five openings. Spiracular openings lead to posterior felt chambers which are connected to a common anterior felt chamber filled with a meshlike network. In third instar H. bovis each medial ecdysial scar is surrounded by a strongly concave spiracular plate. Spiracular openings are surrounded by slightly raised rima. Most rimae bear a spine. Spiracular plates of H. lineatum are flat and rimae are without spines. Each spiracular opening leads to a posterior felt chamber, several of which are confluent with a larger anterior felt chamber. Anterior felt chambers open into the dorsal longitudinal tracheal trunk. Felt chambers in third instar larvae are also filled with a complex mesh.