The feeding behaviour of Psoroptes spp. mites on rabbits and sheep

A scanning electron microscope (SEM) study of the mouthparts of Psoroptes cuniculi from rabbits and P. ovis from sheep established that they are identical in morphology and are adapted for surface feeding rather than piercing the epidermis. Haemoglobin was found in both P. cuniculi and P. ovis on rabbits but not in P. ovis on sheep and it is concluded that the haemoglobin is derived from small haemorrhages on the surface of inflamed rabbit skin. This inflammation is caused by the deposition of antigenic material on the skin which is abraded by the mite mouthparts. The mites feed on the resulting exudate and on other surface material. An immunofluorescent assay revealed that the 2 mite species are antigenically similar.     

Characterisation of aminopeptidase activity in scab mites (Psoroptes spp.)

Soluble and membrane-bound aminopeptidase activities were demonstrated in extracts of P. cuniculi (Delafond). Leucine aminopeptidase (LAP) activity in the soluble fraction of P. cuniculi extracts displayed substrate preference for amino acid derivatives with terminal leucine and methionine over those with acidic, basic or heterocyclic groups. P. cuniculi LAP was inhibited by leucinethiol (IC(50) = 1.4 +/- 0.4 nM), bestatin (IC(50) = 3.9 +/- 1.7 microM), Arphamenine A (IC(50) = 0.37 +/- 0.03 mM) the chelating agent 1,10-phenanthroline (IC(50) = 2.3 +/- 0.5 mM), Zn(2+), Cu(2+) Ni(2+), and Co(2+), and activated by Mn(2+) and Mg(2+). The LAP activity was visualised as a single major band after electrophoresis on native gels and eluted from a size exclusion column as a single major peak representing a molecular mass range of 85-116 kDa. Degenerate oligonucleotide primers were used to amplify short fragments of genomic DNA containing nucleotide sequence coding for the cation-binding motifs of the co-catalytic Zn(2+) binding domains of dizinc leucine aminopeptidases in both P. cuniculi and P.ovis (Hering). The major soluble aminopeptidase from these mites therefore displays most of the characteristics associated with typical cytosolic leucine aminopeptidases belonging to the M17 family of metalloproteinases.     

Hydrolytic enzymes of Psoroptes cuniculi (Delafond)

Phosphatases, C4 and C8 esterases, leucine and valine aminopeptidases, N-acetyl-beta-glucosaminidase, beta-glucosidase, beta-galactosidase and beta-glucuronidase were detected in extracts of the parasitic mite Psoroptes cuniculi. Lipase, trypsin-like and chymotrypsin-like activities were not present. Haemoglobin was hydrolysed by a detergent-soluble fraction of the mite extracts with a maximum hydrolysis between pH 3 and 5. Acid proteinase activity was greater against haemoglobin than bovine serum albumin. Inhibitors of cysteine, serine and metallo-proteinases failed to inhibit the hydrolysis of H-Pro-Thr-Glu-Phe-Phe(NO2)-Arg-Leu-OH while pepstatin A inhibited its hydrolysis in a dose-dependent manner (IC50 8.02 x 10(-11) M (+/- 0.30 x 10(-11). Thermal inactivation of the proteolytic activity followed an exponential decay pattern. Typical K(m) and Vmax values were 7.2 x 10(-5) (+/- 0.7 x 10(-5) M-1 and 1.13 x 10(-3) (+/- 0.05 x 10(-3) OD unit-1 min-1 respectively. Acid proteinase activity eluted from a size exclusion column in a single, major peak representing a molecular weight range of 21-24.5 kDa. The major endoproteinase of P. cuniculi therefore appears to be a cathepsin D-like aspartic proteinase.     

Infection of Psoroptes mites with the fungus Metarhizium anisopliae

The astigmatid mite, Psoroptes ovis (Hering) (Acari: Psoroptidae), is an obligatory ectoparasite that causes psoroptic mange in a range of domesticated animals, particularly sheep, where the clinical disease is known as sheep scab. A series of laboratory assays were used to assess the use of the fungus, Metarhizium anisopliae (Metchnikoff) (Deuteromycotina: Hyphomycetes) as a biocontrol agent for P. ovis derived from rabbits (syn. P. cuniculi). The immersion of mites in a suspension of conidia of M. anisopliae resulted in the acquisition of fatal infections. The number of mites which developed infections increased significantly with the increasing concentration of the conidial suspension to which they were exposed; 77% of mites developed infections when exposed to the highest concentration used (1 x 10(8) conidia ml(-1)). Controls developed no fungal infections. Mites allowed simply to walk across a surface which had been treated with a suspension of conidia also acquired fungal infections; the number infected was again related to the concentration of conidia present. After contact for 24 h with a surface treated with 1 x 10(8) conidia ml(-1), 73% of the mites became infected. To determine whether dead infected mites could act as sources of infection, infected cadavers were placed in chambers with live uninfected mites. The uninfected mites acquired fatal infections from the cadavers; a higher ratio of infected cadavers to uninfected mites resulted in greater transmission of infection. The time after death of the infected cadaver was also an important factor influencing the number infected, 5-day-old cadavers were the most infective and 18-day-old cadavers the least infective. The results indicate that M. anisopliae is a good candidate control agent for Psoroptes mites.     

Blackbuck antelope (Antilope cervicapra), a new host for Psoroptes cuniculi (Acari: Psoroptidae)

Information was requested on the cause of extensive lesions and death of a blackbuck antelope (Antilope cervicapra). Psoroptes cuniculi, the ear mite of domestic rabbits, was identified as the cause of the lesions. Death of the antelope was attributed to a secondary bacterial infection. This is the first report of P. cuniculi on blackbuck antelope. The potential impact of these mites on blackbuck antelope and other exotics in Texas is unknown.     

Pathogenicity and thermotolerance of entomopathogenic fungi for the control of the scab mite, Psoroptes ovis

Psoroptes ovis is responsible for a highly contagious skin condition, both in sheep and cattle. This parasite has a marked economical impact in the sheep and cattle industry. Biological control is considered as a realistic alternative to chemotherapeutic control. Laboratory experiments were carried out to evaluate the pathogenicity and the thermotolerance of twelve isolates of entomopathogenic fungi from four genera (Beauveria Vuillemin, Metarhizium Sorokin, Paecilomyces Bainier and Verticillium Nees). The pathogenicity was evaluated by the survival of P. ovis females after exposure to 10(6) to 10(8) conidia ml(-1) in humidity chambers. Results revealed intra- and interspecies differences. All isolates with the exception of B. bassiana IHEM3558 and V. lecanii MUCL8672 induced 50% mortality within 2 days at the highest concentration. At this concentration the entire mite population became infected with all isolates but B. bassiana IHEM3558; however, only four isolates gave rise to 100% infected cadavers at the lowest concentration. The thermotolerance of each isolate was evaluated by measuring its growth on an artificial medium kept between 25 and 37.5 degrees C. All isolates were able to grow up to 30 degrees C but only two, M. anisopliae IHEM18027 and Paecilomyces farinosus MUCL18885, tolerated temperatures up to 35 degrees C. These two isolates could be considered as good candidates for further use as biopesticide taking into account their virulence and thermotolerance. Other critical factors linked with the implementation of this type of biocontrol in P. ovis infected animals are discussed.     

Dexamethasone-induced bovine T-lymphocyte suppression and the effect upon susceptibility to sheep scab mite (Acari: Psoroptidae) infestation

The synthetic glucocorticoid dexamethasone was used to suppress bovine T-lymphocyte function. Dexamethasone-treated animals were challenged with Psoroptes ovis (Hering) to determine if the immunosuppressive treatment would normalize susceptibility to infestation. Dexamethasone suppressed T-lymphocyte function and anti-P. ovis antibody production, and delayed the onset of scab lesions. All animals were not equally susceptible to infestation as a result of dexamethasone treatment. These data further support the importance of anti-P. ovis antibody in colonization of cattle by P. ovis and the pathology associated with scab mite infestation.     

A physiological and biochemical model for digestion in the ectoparasitic mite,Psoroptes ovis (Acari: Psoroptidae)

Mites are an important group of arthropod pests affecting crops, animals and humans. Despite this, detailed physiological studies on these organisms remain sparse due largely to their small size. Unifying models are required to draw together the diverse information from studies on different groups and species. This paper describes a model for digestion in the parasitic mite, Psoroptes ovis, the causative agent of psoroptic mange or sheep scab disease. The limited information about this species is supplemented with data from other acarines, especially house dust mites and ticks. We review the range of enzymes and allergens found in mites and consider their possible roles in digestion in mites, generally and in particular, P. ovis. Histological studies, enzyme biochemistry and molecular biology and experimental evidence suggest that P. ovis utilises a digestive system reliant upon acid peptidases functioning in a largely intracellular environment. The actions of the digestive enzymes are supplemented by the involvement of bacteria as potential direct and indirect sources of nutrition. It is possible that some extra-corporeal digestion also takes place. The interaction of bacteria and digestive enzymes on the skin surface of the sheep may be responsible for the excessive pathological reactions evident in clinical sheep scab.     

Tactic responses of the parasitic mite, Psoroptes ovis, to light and temperature

The astigmatid mite, Psoroptes ovis (Hering) (Acari: Psoroptidae), is an obligate, non-burrowing ectoparasite of vertebrates, of particular economic importance in domestic sheep flocks where it causes clinical psoroptic mange. To help understand the behaviour which facilitates transmission via the environment, the responses of P. ovis derived from rabbits (syn. Psoroptes cuniculi) to temperature and light were examined in the laboratory. On a vertical surface of uniform temperature, the presence and direction of illumination had a significant effect on the distance and direction moved by the mites. In darkness or with illumination from both above and below, the mites moved relatively little, but this movement was upwards. In contrast, with illumination from above only, mites moved downwards. When the direction of the illumination was reversed so that it came from below only, the mites moved upwards. On a vertical surface with a temperature gradient, in darkness or with illumination from both above and below, the mites moved up or down towards the area of highest temperature, depending on whether this was above or below, respectively. However, the movement of the mites in response to the temperature gradient was strongly displaced up or down by the presence of unidirectional illumination from above or below, respectively. The results indicate that the movement of these mites is strongly directed towards areas of high temperature but away from higher light intensity. These behaviours might be expected to maintain the position of the mites on a host animal and help them locate the skin surface of a new host when displaced into the environment.     

A note on hydrolysis of tributyrin by Branhamella and Neisseria

Sixty-three strains of Branhamella and Neisseria were tested by two methods for their ability to hydrolyse glycerol tributyrate. After the conventional plate test, gas liquid chromatographical (GLC) analysis of the agar medium was carried out to detect the hydrolysis product, butyric acid, and other volatile fatty acids. All strains of Branhamella catarrhalis, Neisseria caviae, N. cuniculi and N. ovis but no other Neisseria spp. gave positive results with the conventional test. With GLC, however, most strains of Branhamella and Neisseria were shown to liberate butyric acid. In addition, some strains liberated acetic and isovaleric acids. Greater amounts of butyric acid were produced by clinical strains, in particular B. catarrhalis , compared with reference strains. It was concluded that the conventional plate test for tributyrin hydrolysis differentiates B. catarrhalis, N. caviae, N. cuniculi and N. ovis from other Neisseria.     

A note on hydrolysis of tributyrin by Branhamella and Neisseria

Sixty-three strains of Branhamella and Neisseria were tested by two methods for their ability to hydrolyse glycerol tributyrate. After the conventional plate test, gas liquid chromatographical (GLC) analysis of the agar medium was carried out to detect the hydrolysis product, butyric acid, and other volatile fatty acids. All strains of Branhamella catarrhalis, Neisseria caviae, N. cuniculi and N. ovis but no other Neisseria spp. gave positive results with the conventional test. With GLC, however, most strains of Branhamella and Neisseria were shown to liberate butyric acid. In addition, some strains liberated acetic and isovaleric acids. Greater amounts of butyric acid were produced by clinical strains, in particular B. catarrhalis, compared with reference strains. It was concluded that the conventional plate test for tributyrin hydrolysis differentiates B. catarrhalis, N. caviae, N. cuniculi and N. ovis from other Neisseria.     

Structure and function of the gnathosoma of the mange mite,Psoroptes ovis

Mites of the genus Psoroptes (Acari: Psoroptidae) are obligate, non-burrowing, astigmatid ectoparasites of mammals. A detailed understanding of the morphology of the gnathosoma is an important step towards elucidation of the feeding behaviour of this mite and, hence, the pathology of psoroptic mange. Scanning and transmission electron microscopy were used to examine Psoroptes ovis (Hering) (syn. P. cuniculi). The gnathosoma is composed of an infracapitulum (hypostome), which forms the floor of an extended U-shaped pre-oral trough, bounded by laterally compressed palps and enclosing paired chelicerae. Distally, each palp terminates in a flap-like process and four tapered terminal sensillae. The floor of the infracapitulum is interrupted along its length by a dorsally projecting ridge. At the distal end of the infracapitulum are paired, grooved pseudorutellae and, between them, paired, ungrooved lateral lips. Between the lateral lips is a grooved, elongate labium. Medially, a pair of finger-like projections emanates from the paraxial walls of the palps and project anteriorly. Each chelicera is made up of fixed and mobile digits, with a cheliceral membranous fold articulating the two. The distal ends of both digits are chelated. For much of its length, the mobile cheliceral digit appears to lie within a fold formed by the infracapitular ridge, thereby creating a central channel between the chelicerae. It is suggested that this arrangement of elements may facilitate a two-way flow of liquid, where saliva flows down a central salivary canal and spills out onto the host's skin over the labium at the point where the chelated tips of the chelicerae meet and abrade the skin. Liquid food may move along the grooves of the pseudorutellae and then be sucked up a food canal formed by the paraxial walls of the palps and the outer wall of the chelicerae.     

Results of exposure to synthetic insect development inhibitors in Psoroptes cuniculi (Psoroptidae) mites

Results of the effect of synthetic analogs of juvenile hormones of insects, dimiline, altozide and altozare, on the mite P. cuniculi have been shown. It has been noted that repeated coverings of mites in Petri dishes and rabbits' skin with 0.005--5% emulsions of altozide, 0.005--10% emulsions of altozare and 0.25--8% suspensions of dimiline cause no changes in their morphology and metamorphosis. 1--4% emulsions of altozide and altozare and 2--8% suspensions of dimiline have been found to have an acaricide effect and cause 100 per cent mortality of mites within 48 hours.     

Serodiagnostic antibody responses to Psoroptes sp. infestations in bighorn sheep

The antibody responses of bighorn sheep (Ovis canadensis) infected with Psoroptes sp. mites were investigated by enzyme linked immunosorbent assay on western blots of P. cuniculi antigens. Serum from 20 Psoroptes sp.-infested bighorn sheep (O. canadensis mexicana, O. canadensis nelsoni, O. canadensis canadensis) from New Mexico, Nevada, California, and Idaho reacted strongly with mite antigens ranging from 12 to 34 kd. Serum from 35 Psoroptes sp.-free bighorn sheep of unknown tick infestation status and from three Psoroptes sp.-free bighorn sheep infested with Dermacentor hunteri ticks did not react with these antigens. Psoroptes sp.-specific antibody responses were present throughout a 16 mo period in one infected bighorn sheep, but were not detectable 8 mo following successful treatment. These results demonstrate that specific serodiagnosis of Psoroptes sp. infestation is feasible in bighorn sheep and suggest that antibody responses are indicative of current or recent infestation.     

Cattle and small ruminant piroplasmosis in Macedonia, Greece

Macedonia is an endemic region of cattle as well as of sheep and goat piroplasmosis In cattle, Theileria orientalis (=T. buffeli?), the agent of Eurasian benign theileriosis, is the most common and widespread piroplasm species. T. annulata, the agent of tropical theileriosis, seems to be rare and limited to few foci, but causes very severe clinical cases, especially in imported pure-bred or cross-bred animals. Babesia bovis and B. bigemina are present in several localities, and often coexist. Cattle babesiosis cases are due to both piroplasms, but B. bovis is considered responsible for the severe and acute clinical cases. Ovine and caprine babesiosis is due to B. ovis and mainly affected imported animals as well as indigenous animals which have been transported from localities where the infection was absent. B. ovis is extremely widespread in both sheep and goats. During the last decades, no clinical cases of small ruminant theileriosis have been registered in this region. However, T. ovis, a non-pathogenic theileria, is common in sheep, but not in goats. Anaplasma ovis, a protobacterium of small ruminants, is also present in the region.     

Kinetic ELISA for detection of antibodies to Psoroptes sp. (Acari: Psoroptidae) in bighorn sheep (Ovis canadensis)

A kinetic enzyme-linked immunosorbent assay (ELISA) was developed using antigenic extracts prepared from Psoroptes cuniculi mites and sera from 37 Psoroptes sp.-infested and 43 uninfested bighorn sheep (Ovis canadensis). Serial dilutions of these serum samples, representing 3 bighorn sheep subspecies and 9 geographic areas, gave parallel responses when plotted as log dilution versus log kinetic rate. Therefore, all 80 samples were run at a single dilution (1:100) and positive/negative cutoff values were established as the mean kinetic rate of all negative sera plus either 2, 3, or 4 standard deviations. The resulting ELISA was highly reproducible and accurate with sensitivities and specificities of 100% and 97.7%, 94.6% and 97.7%, and 94.6% and 100%, respectively. This immunoassay will be useful for prospective and retrospective studies assessing the distribution and prevalence of Psoroptes sp. infestations in bighorn sheep.     

Protein glycosylation in the spores of the microsporidia Paranosema (Antonospora) grylli

Long adaptation of microsporidia, a large group of fungi-related protozoa, to intracellular lifestyle has resulted in drastic minimization of a parasite cell. Thus, diversity of carbohydrates in microsporidia glycoproteins and proteoglycans is expected to be restricted by O-linked manno-oligosaccharides because three genes involved in O-mannosylation of proteins and no components of N-linked glycosylation machinery were found in genome of human pathogen Encephalitozoon cuniculi. In this study we investigated glycosylation of spore proteins of microsporidia Paranosema (Antonospora) grylli infecting crickets Gryllus bimaculatus. Using periodic acid-Shiff reagent staining we have demonstrated that some P. grylli spore proteins are highly-glycosylated. The major polar tube protein (PTP1) of 56 kDa was shown as the most intensively decorated band. The experiments with N-glycosidase F and WGA lectin did not reveal any N-glycosylated proteins in P. grylli spores. At the same time, incubation of major spore wall protein of 40 kDa (p40) with mannose specific lectin GNA resulted in specific binding that was reduced by pretreatment of the protein with mannosidases. Interestingly, in spite of PTP1 glycosylation, polar tube proteins extracted from P. grylli spores were not precipitated by GNA-agarose. Since P. grylli and E. cuniculi are distantly related, our data suggest that dramatic reduction of protein glycosylation machinery is a common feature of microsporidia.     

Aerobic and microaerophilic bacteria isolated from Psoroptes cuniculi

The bacterial flora of Psoroptes cuniculi removed from nine naturally infested rabbits was investigated. Mites were collected in sterile glass tubes; half of the mites were surface sterilised, the others were not. All mites were crushed using sterile glass pestles, placed in Buffered Peptone Broth, smeared on to several culture media, by glass rods, and incubated at 37 degrees C for 48 hours, aerobically and/or in 5% CO2. Representative colonies were removed and streaked on to several selective media. Different colour changes of the selective media used, macro and microscopic morphology, ability to grow aerobically, Gram staining, and several biochemical tests evaluated with API test strips, were used for bacterial identification. Staphylococcus aureus, Serratia marcescens and S. odorifera were the bacteria isolated from surface sterilised mites.     

Encephalitozoon cuniculi: quantitation of parasites and evaluation of viability

Two methods (manual and automated) for quantitation of viable versus dead Encephalitozoon cuniculi are reported. The manual method uses ethidium bromide and acridine orange to stain dead and viable organisms, respectively. The stained organisms are visually differentiated with the aid of a fluorescence microscope. The automated method uses propidium iodide to stain dead parasites, which are differentiated from viable unstained parasites with the aid of a flow cytometer. An automated cell counter (Coulter Counter) was used to count rapidly large numbers of samples and to improve the sensitivity of counting low concentrations of parasites. These methods will enhance investigators' abilities to conduct quantitative experiments on host defense mechanisms against E. cuniculi.