Crossreacting IgG antibodies against fox mite antigens in human scabies.

Scabies continues to be an important parasitic disease of mammals. There remain, however, major gaps in the understanding of the human host immune response, and a simple diagnostic test is lacking. In contrast to human mites, red fox mites (Sarcoptes scabiei var. vulpis) can be collected easily and have been used, due to crossreactivity, for enzyme-linked immunosorbent assay (ELISA) studies in dogs and pigs. We wanted to investigate the possibility that crossreactivity might also exist for the human mite, and determined titers against fox mite antigens by ELISA in 41 patients with scabies. Specific IgG was significantly higher in patients with scabies than in healthy controls (P=0.01). The sensitivity was, however, only 48%, although it increased slightly during treatment (P=0.86). A positive correlation was also noted between disease duration and severity of infestation (r=0.5), with specific IgG titers increasing in parallel with severity of symptoms (P=0.01). Patients with symptomatic scabies for more than 4 weeks had furthermore significantly higher IgG titers than patients with a shorter duration of disease (P=0.007). In conclusion, these findings demonstrate IgG antibodies in human scabies that crossreact with fox mite antigens, thus encouraging the search for improved ELISAs with more specific mite antigens to produce a more sensitive detection system for scabies in humans.

Evaluation of the efficacy of emodepside+praziquantel topical solution against cestode (Dipylidium caninum, Taenia taeniaeformis, and Echinococcus multilocularis) infections in cats.

Emodepside+praziquantel topical solution was developed to provide broad-spectrum anthelmintic activity against gastrointestinal parasites in cats. Eight controlled studies were conducted to evaluate the efficacy of a topical solution of emodepside (3 mg/kg) and praziquantel (12 mg/kg) (Profender, BayerAG, Leverkusen, Germany) against feline infections with three species of cestodes. Studies featured naturally acquired infections of Dipylidium caninum or Taenia taeniaeformis, or experimental infections with Echinococcus multilocularis that were placebo-controlled, randomized and blinded. Cats were euthanatized and necropsied between 2 and 11 days after treatment, depending on the target parasite. The efficacy of emodepside+praziquantel topical solution was 100% against D. caninum and T. taeniaeformis, and 98.5- 100% against E. multilocularis. No significant systemic or local adverse reactions to treatment were noted in cats that received the combination. Topical treatment of cats with emodepside+praziquantel topical solution was safe and highly effective against cestode infections.

High dose folic acid treatment of fragile (X) males.

We conducted an experimental trial of high-dose folic acid given to five males, ages 8 to 26 years, with the fra(X) syndrome. In this double blind study, each subject received 250 mg per day of folic acid for 3 months, followed by placebo for 3 months, and folic acid again for an additional three months. Based on IQ tests, behavior ratings, the Autistic Descriptors Checklist, and parental ratings, there was little evidence to suggest any positive effects seen during the administration of high-dose folic acid. Therefore, this study has provided little support for a hypothesis of benefit of high-dose folic acid in the treatment of the fra(X) syndrome.

Fragile X and autism: a multicenter survey.

We screened 183 autistic males for the fra(X) and found 24 (13.1%) to be positive. Adding the subjects of this study to those of 11 other surveys, of which 6 were positive and 5 were negative, a total of 614 autistic males have been screened. Overall 47 (7.7%) were positive. Based on this estimate and the prevalence of autism and fra(X), we estimate that 12.3% of fra(X) males are autistic. We have found that 17.3% of our fra(X) males were autistic and overall a 21.2% frequency has been reported, these higher figures are most likely due to biases in age and ascertainment. With an overall 7.7% frequency of fra(X) among autistic males and an estimated 12.3% of autism among fra(X) males, we conclude there is likely to be a significant association of fra(X) with autism. Because fra(X) appears to be the single most common cause of the condition, chromosomal testing is recommended for any autistic person with undiagnosed etiology.

Detection and differentiation of Theileria annulata and Theileria parva using macroschizont-derived DNA probes.

A lamda gt11 expression library based on T.annulata-infected cells was screened with an antiserum raised in rabbits against partially purified schizonts of T.annulata. Two clones were detected, sequenced and designated as SA288 and SB288 (Shayan et al., submitted for publication). From the sequences of these two genes oligonucleotide primers were designed for specific amplification of parasite DNA by polymerase chain reaction (PCR). We could show that these genes are of parasitic origin and do occur in all T.annulata stocks tested in the present study. In addition, a target sequence for SA288 could also be identified in T.parva-schizonts. None of them reacted with genomic DNA of different Babesia spp. A third primer pair was designed from the DNA-sequence of a gene encoding for the T.parva-specific casein kinase II-alpha subunit. Using this primer pair, a target sequence could only be detected in T.parva. Taken together, the primers described here can be used as molecular tools in PCR for the detection of Theileria parasites and to distinguish T.annulata from T.parva.

Lymphocyte stimulatory capacity of Theileria annulata-infected ovine lymphoblastoid cells.

T. annulata, the causative agent of tropical theileriosis in cattle, is transmitted by ticks of the genus Hyalomma. Sporozoites of this parasite invade their target cells, where they differentiate to macroschizonts. T. annulata additionally invades and transforms ovine and caprine leukocytes. T. annulata infection in the ovine system is poorly studied, thus we used a mixed lymphocyte culture (MLC) to analyze the capacity of these cells to activate naïve uninfected ovine cells. The peak response was observed on day three or four and the response could not be induced by lysates of infected cells or their supernatants. The stimulated cells expressed IL-2 and secreted an IL-2-like growth factor.

Generation of cytotoxic T lymphocytes and cytostatic acting cells in T. annulata-immune cattle.

Cattle immunized against Theileria annulata with schizont containing autologous cell lines are immune to challenge with a homologous parasite strain. Two cell types have been detected in the peripheral blood of the immunized animals: cytotoxic T lymphocytes (CTL) and cytostatic acting cells (CAC). Killing the target cells by CTL is infection associated and is MHC class I restricted. Hence, no cytotoxicity was observed against target cells that were treated with the theilericidal drug buparvaquone or autologous Con A-blasts. The growth inhibition of CAC is MHC unrestricted, and not mediated by cytokine interferon gamma (IFN-gamma).

Electron microscopic studies on the development of kinetes of Theileria parva Theiler, 1904 in the gut of the vector ticks Rhipicephalus appendiculatus Neumann, 1901.

The development of motile stages, called kinetes, from a stationary stage (regarded as zygote) has been followed in Theileria parva by means of electron miscroscopy. This process started after moult of the tick nymphs which had sucked on highly infected calves, i.e. about 20 days after repletion (a.r.) of the ticks. The transformation took place by formation of a growing protrusion (= anlage) into an inner, enlarging vacuole. During this process the limiting membrane of the enlarging vacuole serves as the outer membrane of the developing motile stage, whereas the two inner ones as well as the subpellicular microtubules are newly formed. This transformation proceeds rapidly, so that on the 25th day a.r. most of the kinetes have already left the gut cells and started penetration into the salivary gland cells. On the way to the salivary glands nuclear divisions occurred within the kinetes. The steps of the transformation described were compared to those in T. annulata and to ookinete formation in haemosporidia.

Electron microscopical studies on the development of Babesia canis (Sporozoa) in the salivary glands of the vector tick Dermacentor reticulatus.

The formation of sporozoites of Babesia canis was studied by light- and electron microscopy in the salivary gland cells of adult female ticks from the 2nd day after attachment until 1 day after detachment. It was found that this process was initiated by the binary division of kinetes that had already entered or entered during the period examined. During division the kinetes (15 X 2.5 microns) lost their typical organelles, reduced their three-layered pellicle to a single membrane and became spherical. After nuclear division and a further time-lag cell division occurred, giving rise to two cells in which this process was repeated. After numerous binary divisions the parasites acquired more and more closely the shape of the later infectious, pyriform sporozoite. These sporozoites measured about 2.5 X 1.5 microns and also had a three-layered pellicle, with rhoptries and a few micronemes, but never contained "spherical bodies". The formation process needed about 2--3 days so that the transmission to the dog could be carried out while the tick engorges and this is probably the stimulation for the development. Finally the cytological features of this sporozoite formation were compared to those in the Theileria species studied by our group.

Borna disease virus-specific antigens. II. The soluble antigen is a protein complex.

Borna disease virus-specific soluble antigen from persistently infected rat brains was purified to homogeneity using preparative sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The soluble antigen is a complex of three proteins with apparent molecular weights of 35 kDa, 38 kDa and 24 kDa. The 35/38 kDa antigen double band was separated into its two components. The 24 kDa protein has no common epitopes with the 35/38 kDa protein.

Effect of temperature on transovarial transmission of Babesia bigemina (Smith and Kilborne, 1893) in Boophilus annulatus (Say, 1821).

The effect of the temperature on the transovarial passage of Babesia bigemina has been studied in female Boophilus annulatus during their oviposition. Kinetes of Babesia were present in eggs laid at the temperatures of 16, 20, 26, 28, 30 and 35 degrees C. The rate of infection was temperature dependent. It reached at least 50%, even at low temperatures. The average infection level at 26 and 30 degrees C was 270 kinetes per egg. However, between 20 and 35 degrees C the eggs laid during the first 3 days were parasite free. At 16 degrees C, no kinetes were detected during the first 13 days of oviposition.

Application of chemi- and bioluminescence in studies of immunological reactions against protozoa.

The opsonization and lysis of different protozoa by antibodies and/or complement was followed using luminol-dependent chemiluminescence and bioluminescence. The addition of immune serum to variable antigen type populations of Trypanosoma evansi led to the specific opsonization of trypanosomes resulting in an intense metabolic activation and chemiluminescence response of phagocytic cells. In comparison to those of uninfected control mice, the phagocytosis of coccidia merozoites by spleen cells from mice infected with Eimeria falciformis was enhanced during the acute stage of a primary infection. Opsonizing activity was demonstrated in phosphate-buffered saline extracts of gut contents of mice infected for 10 days. The incubation of E. falciformis merozoites together with guinea-pig complement resulted in slow lysis of the cells. The addition of mouse serum collected greater than 6 days after an infection led to an accelerated lysis of the merozoites, indicating the appearance of complement-fixing antibodies in the serum. Heat-inactivated immune serum alone had no lysing activity on merozoites. In the presence of complement, bovine lymphoblastoid cells infected with Theileria annulata were lysed by anti-lymphoblastoid cell serum raised in mice but not by serum from cattle which had developed immunity to Theileria annulata.

Detection of a new pathogenic Babesia microti-like species in dogs.

Small babesiae in dogs are generally considered to belong to Babesia gibsoni. Here we describe the genotypic characterisation of small piroplasms found in the blood of a dog which suffered from clinical babesiosis. Pairwise identities as well as distance, parsimony and maximum likelihood analyses of the 18S rDNA clearly demonstrated that this isolate was only distantly related to the other canine piroplasms characterised genetically so far, including B. gibsoni. It was more closely related to B. microti, B. rodhaini, and Theileria equi. It is concluded that the small canine piroplasms described in this study represent a hitherto unknown species and that the fauna of piroplasms occurring in dogs is more diverse than assumed so far.

Detection of antibodies in sera of weaned pigs after contact infection with Sarcoptes scabiei var. suis and after treatment with an antiparasitic agent by three different indirect ELISAs.

Three commercial enzyme-linked immunosorbent assays (ELISAs) were compared for the detection of antibodies to Sarcoptes scabiei var. suis using experimental sera of six 8-week-old pigs after contact infection with Sarcoptes scabiei var. suis. Six non-infected pigs were monitored as a control group. Blood sera were taken once a week from all animals. After successful infection the pigs were treated with an antiparasitic agent (12 weeks post infection (p.i.)) and the antibody titres were monitored until they were negative. The antibody levels of the experimental pigs reached the cut-off level 5 weeks after introduction of an infected animal to the group and were positive by both the Sarcoptes-ELISA 2001 PIG and the Acar-Test P-ELISA. Four weeks after treatment mean results showed optical densities (% OD) below the cut-off level in the Sarcoptes-ELISA 2001 and 8 weeks after treatment in the Acar-Test P-ELISA. In the Chekit Sarcoptest pigs had elevated antibody levels in comparison to control animals, but ODs remained below the given cut-off level at all times. In a second examination with Chekit Sarcoptest (different lot) and at a lower cut-off level, the sera of most of the piglets tested positive. Eight weeks after treatment, four from six pigs still had positive OD values. Therefore this investigation showed a higher sensitivity for the Sarcoptes-ELISA 2001 and the Acar-Test P-ELISA than for the Chekit Sarcoptest. Different test sensitivities must be considered when serologic methods are used for the diagnosis of swine sarcoptic mange, especially for monitoring and controlling eradication programs.

Genital myiasis (Wohlfahrtiosis) in camel herds of Mongolia.

Genital infestations caused by Wohlfahrtia magnifica (Schiner, 1862) has been reported as being a common problem encountered in female camels in the Gobi desert. Thus, a field investigation to establish the prevalence of wohlfarthiosis in camel herds in the eastern Gobi district was carried out between May and July 1994. The objectives of the study were to establish the relevance of genital myiasis, and to describe the disease patterns, clinical course and pathomorphological changes associated with it. The field study was designed to suit the specific ecological, infrastructural and logistic conditions. Two cross-sectional samples were taken from 45 selected herds in six different areas of the Chatanbulag Sumon to obtain data on herd structure, age distribution, and the breeding history of infested female animals. Post partum events and previous occurrence of genital myiasis were recorded by use of a standardised questionnaire. Clinically infested animals were first detected visually. All camels to be investigated further were subsequently immobilised using ropes and the genitals were examined for tissue lesions and the presence of larvae. Examination of 1676 Bactrian camels from 45 selected herds in six different areas of the Chatanbulag Sumon in the Eastern Gobi district, Mongolia, led to an estimate of Wohlfahrtia magnifica infestation rates between 8-10%. Most myiasis cases were found in older females (> 4 yr), younger animals were infested at a lower rate. Highest prevalence rates were discovered during June and July, highest levels of infestation occurred in the Aman Us Chudak region with infestation rates up to 15%.

Conversational analyses of males with fragile X, Down syndrome, and autism: comparison of the emergence of deviant language.

Deviant, repetitive language of 33 males (9 with Down syndrome, 12 with fragile X (fra[X]) syndrome, and 12 with autism) was analyzed within three conversational contexts: direct responses, initiation of new material, and topic maintenance. Results indicated that males with fra(X) manifest deviant, repetitive language that is distinct from males with either Down syndrome or autism. Thus, the deviant repetitive language of males with fra(X) cannot be accounted for by either their level of adaptive functioning or autistic-like behaviors per se. Possible explanations for this etiologically specific language deviance were discussed.

Social gaze, social avoidance, and repetitive behavior in fragile X males: a controlled study.

Preference for social gaze as well as the percentage occurrence of social gaze, nonverbal social avoidance, and nonverbal repetitive behaviors were examined in autistic and nonautistic prepubertal males with the fragile X syndrome (fra[X]) during social interaction with a parent or stranger. Comparison groups were nonhandicapped, Down syndrome, atypical pervasive developmental disorder, and autistic males. The subjects with fra(X) and the nonhandicapped and Down syndrome control subjects discriminated parent from stranger as evidenced by their avoidance behavior. The overall percentage of avoidance was higher, however, for both parent and stranger, among the males with fra(X). Autistic and atypical groups without fra(X) failed to discriminate parent from stranger in their avoidance behavior. Possible explanations for these group differences in terms of language level or degree of language demand were ruled out. Implications for research concerning the relations among fra(X), autism, and mental retardation were discussed.

Light and electron microscopic study on developmental stages of Babesia canis within the gut of the tick Dermacentor reticulatus.

The initial developmental stages of Babesia canis within the gut of the vector tick, Dermacentor reticulatus, were studied by means of light and electron microscopy. Large quantities of varying "spiky-rayed" stages and few ovoid stages without any protrusions were observed after lysis of the engorged canine erythrocytes. The transformation of spherical erythrocytic stages into "spiky-rayed" forms was initiated inside the intact erythrocytes. The significance of all developmental stages in a hypothetical life cycle of Babesia species is discussed.

Fine structure of Babesia equi Laveran, 1901 within lymphocytes and erythrocytes of horses: an in vivo and in vitro study.

The development of Babesia equi (Piroplasmia) in the vertebrate host was studied by electron microscopy. The tick-borne sporozoites initiated an exoerythrocytic schizogony in lymphocytes in vivo and in vitro. In lymphocyte cultures the schizonts of B. equi lay as fissured bodies directly within the host cell forming numerous cytomeres. These cytomeres were bordered by a single membrane and contained polymorphous nuclei. Merozoite differentiation was initiated at several places by protrusions appearing at the periphery of the cytomeres. The cytoplasm of the schizont merged progressively into the young merozoites. The mature merozoites were pear-shaped (2.0 X 1.5 microns), bounded by a pellicle and had an apical complex without conoid. Numerous merozoites were observed entering erythrocytes in vitro. In vivo, lymphocytes containing B. equi schizonts were seen in capillaries within lymph nodes from the 12th day onwards after attachment of infected ticks. At the same time some erythrocytes were found that contained B. equi merozoites. Development of B. equi in the vertebrate host shows many similarities to the corresponding development of Theileria parasites. This makes a new discussion of the taxonomic position of B. equi necessary.