Field efficacy and safety of a combination of moxidectin and imidacloprid for the prevention of feline heartworm (Dirofilaria immitis) infection

Throughout the end of March to beginning of May 2006, 212 owned cats and 608 owned dogs from a heavy endemic area for canine heartworm (HW) disease in northern Italy have been examined to assess HW infection prevalence. Both cats and dogs were clinically examined and blood samples were taken from each animal to be examined for HW antibody (Ab). Ab-positive cats were further examined for circulating microfilariae, HW antigens (Ag) and by echocardiography (ECHO) to assess the presence of adult worms. Dogs were clinically examined and blood samples taken from each animal were examined for circulating microfilariae and for HW Ag. Ten cats (4.7%) were found Ab positive. Of these, 6 cats were Ag positive (2.6%) and in 4 (1.8%) the worms were visualized by ECHO. HW prevalence in dogs was 36% (221/608). One hundred and seventy-six (29%) were both microfilaraemic and Ag positive, 40 (7%) had occult infections (no circulating microfilariae) and 7 (1%) were microfilaraemic but Ag negative. Upon owners’ consent, 132 cats (including cats Ab and/or Ag and ECHO positive) were prophylactically treated against HW disease with an imidacloprid/moxidectin spot-on combination (10% imidacloprid/1% moxidectin) monthly administered for 6 months. Cats were re-examined for HW infection in November, 1 month after the last drug administration, and in May–June 2007, 7–8 months after the last treatment. All 122 cats found HW negative before treatment, were found negative at the two examinations at the end of study. The 4 cats Ab positive, 2 cats Ab and Ag positive and 1 Ab, Ag and ECHO positive at the beginning of treatment were found negative. Throughout the treatment, transitory hypersalivation and generic signs of annoyance were reported by owners in 6 cats (4.5%). All signs regressed spontaneously.     

Feline heartworm (Dirofilaria immitis) infection: a statistical elaboration of the duration of the infection and life expectancy in asymptomatic cats

A study was conducted to assess the duration and the outcome (self-cure or death) of feline heartworm infection and the life expectancy of infected cats. To be included in the study, cats had to be positive for heartworm antibody (Ab) and heartworm antigen (Ag) and had to demonstrate the presence of worms by echocardiography. Self-cure was defined as (1) negative for heartworm Ag and (2) no further visualization of worms by echocardiography. Of the 1962 eligible cats, 364 (18.5%) were positive for heartworm Ag and 131 were positive for heartworm Ag and for echocardiography diagnosis (prevalence 6.7%). None of the cats was microfilaremic. Of 43 asymptomatic cats included into the follow-up study with owners' consent, 34 (79%) self-cured and nine (21%) died. Eleven (26%) cats remained asymptomatic and self cured within 21-48 months, 23 (53%) showed symptoms but self-cured within 18-49 months, 6 (14%) died within 8-41 months of follow-up and 3 (7%) suddenly died after 38-40 months, which was related to heartworm infection. The probability for death or sudden death increased significantly with age at diagnosis, but no difference was detected by gender, survival time after diagnosis, or the presence or absence of symptoms. The presence/absence of symptoms showed significant interaction with the age at diagnosis (i.e., symptomatic cats showed increasing duration of heartworm infection along with age at diagnosis compared to that for asymptomatic cats. Heartworm-infected cats survived significantly longer than heartworm-negative cats affected by hypertrophic cardiomyopathy, chronic renal failure, or neoplastic diseases.     

The validity of some haematological and elisa methods for the diagnosis of canine heartworm disease

Examinations for heartworm (Dirofilaria immitis) were performed on 175 impounded dogs from a hyperendemic area of the Po Valley (Italy). Each blood sample was used wiht five haematological diagnostic methods (filtration, direct smear, modified Knott, clotted blood and capillary tube) and three commercial ELISA kits (PetChek, Diasystems, Uni-Tec). The results were compared with the true infection status obtained from post-mortem examination of the heart, pulmonary arteries, thoracic venae cavae and lungs. The prevalence of the infection by adult worms at necropsy was 63%. The sensitivity of the tests ranged from 60% (capillary tube) to 81% (Diasystems) and the specificity from 88% (filtration) to 98% (PetChek). The results of all the tests differed significantly (p<0.01) from those obtained at necropsy. The sensitivity of the tests was also assessed with respect to the differing numbers of worms in the hosts. A positive correlation between the worm burden and the sensitivity was observed in all the tests. It is apparent that the ELISA methods were better able to detect cases with a low number of worms than the haematological tests.Abbreviations AC accuracy - CB clotted blood - CT capillary tube - DI Diasystems - DS direct smear - ELISA enzyme-linked immunosorbent assay - FI filtration - MK modified Knott - NPV negative predictive value - PE PetChek - PPV positive predictive value - SE sensitivity - SP specificity - UN Uni-Tec     

Evaluation of ELISA coupled with Western blot as a surveillance tool for Trichinella infection in wild boar (Sus scrofa)

Trichinella surveillance in wildlife relies on muscle digestion of large samples which are logistically difficult to store and transport in remote and tropical regions as well as labour-intensive to process. Serological methods such as enzyme-linked immunosorbent assays (ELISAs) offer rapid, cost-effective alternatives for surveillance but should be paired with additional tests because of the high false-positive rates encountered in wildlife. We investigated the utility of ELISAs coupled with Western blot (WB) in providing evidence of Trichinella exposure or infection in wild boar. Serum samples were collected from 673 wild boar from a high- and low-risk region for Trichinella introduction within mainland Australia, which is considered Trichinella-free. Sera were examined using both an ‘in-house’ and a commercially available indirect-ELISA that used excretory–secretory (E/S) antigens. Cut-off values for positive results were determined using sera from the low-risk population. All wild boar from the high-risk region (352) and 139/321 (43.3%) of the wild boar from the low-risk region were tested by artificial digestion. Testing by Western blot using E/S antigens, and a Trichinella-specific real-time PCR was also carried out on all ELISA-positive samples. The two ELISAs correctly classified all positive controls as well as one naturally infected wild boar from Gabba Island in the Torres Strait. In both the high- and low-risk populations, the ELISA results showed substantial agreement (k-value = 0.66) that increased to very good (k-value = 0.82) when WB-positive only samples were compared. The results of testing sera collected from the Australian mainland showed the Trichinella seroprevalence was 3.5% (95% C.I. 0.0–8.0) and 2.3% (95% C.I. 0.0–5.6) using the in-house and commercial ELISA coupled with WB respectively. These estimates were significantly higher (P < 0.05) than the artificial digestion estimate of 0.0% (95% C.I. 0.0–1.1). Real-time PCR testing of muscle from seropositive animals did not detect Trichinella DNA in any mainland animals, but did reveal the presence of a second larvae-positive wild boar on Gabba Island, supporting its utility as an alternative, highly sensitive method in muscle examination. The serology results suggest Australian wildlife may have been exposed to Trichinella parasites. However, because of the possibility of non-specific reactions with other parasitic infections, more work using well-defined cohorts of positive and negative samples is required. Even if the specificity of the ELISAs is proven to be low, their ability to correctly classify the small number of true positive sera in this study indicates utility in screening wild boar populations for reactive sera which can be followed up with additional testing.     

Application of recombinant Sjc26GST for serodiagnosis of Schistosoma japonicum infection in water buffalo (Bos buffelus)

Schistosomiasis japonica is currently the most serious parasitic disease in mainland China and it is estimated that several million people are infected. Furthermore, it is also responsible for the deaths of many domestic animals. In order to establish an effective diagnostic method, the gene encoding Sjc26GST was cloned and expressed in Escherichia coli as a fusion protein with His-tag. The purified reSjc26GST was used as an antigen for an enzyme-linked immunosorbent assay (ELISA) and for immunoblotting detection of Schistosoma japonicum antibodies in water buffaloes. Our results showed that mean OD values of specific serum IgG antibodies from egg-positive buffaloes were 3.37-fold higher than what was found in egg-negative buffaloes from non-endemic areas. The data also showed the OD value of the endemic egg-negative group reached as high as 1.69 times as that found in non-endemic areas. The positivity rate of egg-positive buffaloes was 100%, but was 30.3% in the endemic egg-negative group. Infected bovine antisera also recognized reSjc26GST, a 27 kDa protein as determined by Western blot. These results suggest that the recombinant GST expressed in E. coli should be an effective diagnostic reagent for detection of antibody against S. japonicum in buffaloes. Due to straightforward production, excellent sensitivity and high specificity, the reSjc26GST described in this study can be considered as a candidate protein for immunological diagnosis of bovine schistosomiasis. Developing reSjc26GST, with its potential diagnostic values, will be useful for diagnosis and surveillance of schistosomiasis in controlling the spread of this parasitic disease in domestic animals.     

Utility of 2 Immunological Tests for Antemortem Diagnosis of Equine Protozoal Myeloencephalitis (Sarcocystis neurona Infection) in Naturally Occurring Cases

Background: Antemortem diagnosis of equine protozoal myeloencephalitis (EPM) is challenging. Limited information is available regarding a commercial test (surface antigen 1 [SAG‐1] ELISA). Performance of another commercial test (indirect fluorescent antibody test [IFAT]) using samples from an independent group has not been well described. Hypothesis/Objectives: The primary goal was to evaluate the SAG‐1 ELISA and IFAT using naturally occurring EPM cases. A secondary goal was to obtain more information regarding clinical presentation. Animals: Hospital cases were admitted over 20 months and classified into 4 groups. Confirmed positive cases (n = 9) had asymmetric or multifocal neurologic deficits or both and postmortem lesions consistent with EPM. Confirmed negative cases (n = 17) had variable clinical signs and postmortem lesions consistent with another neurologic disease (or no lesions). Suspected positive cases (n = 10) had asymmetric or multifocal deficits or both, marked improvement after treatment for EPM, and other likely diseases excluded. Suspected negative cases (n = 29) had orthopedic disease and no neurologic deficits. Methods: Results of immunological testing (SAG‐1 ELISA and IFAT on serum or cerebrospinal fluid [CSF] or both), neurologic examinations, CSF analyses, and postmortem examinations were analyzed retrospectively. Results: SAG‐1 ELISA sensitivity was 12.5% (95% CI, 1.6–38.4) and specificity was 97.1% (95% CI, 84.7–99.9) using serum. IFAT sensitivity was 94.4% (95% CI, 72.7–99.9) and specificity was 85.2% (95% CI, 66.3–95.8) using serum; sensitivity was 92.3% (95% CI, 64.0–99.8) and specificity was 89.7% (95% CI, 72.7–97.8) using CSF. Conclusions and Clinical Importance: Low sensitivity of the SAG‐1 ELISA limited its usefulness for antemortem diagnosis of EPM in this patient population.     

Purification and analyses of the specificity of two putative diagnostic antigens for larval cyathostomin infection in horses

Cyathostomins are important equine gastrointestinal parasites. Mass emergence of mucosal stage larvae causes a potentially fatal colitis. Mucosal stages are undetectable non-invasively. An assay that would estimate mucosal larval stage infection would greatly assist in treatment, control and prognosis. Previously, we identified two putative diagnostic antigens (20 and 25 kDa) in somatic larval preparations. Here, we describe their purification and antigen-specific IgG(T) responses to them. Western blots confirmed the purity of the antigens and showed that epitopes in the 20 kDa complex were specific to larval cyathostomins. No cross-reactive antigens appeared to be present in Parascaris equorum or Strongyloides westeri species. Low levels of cross-reactivity were observed in Strongylus edentatus and Strongylus vulgaris species. Use of purified antigens greatly reduced background binding in equine sera. These results indicate that both antigen complexes may be of use in a diagnostic assay.     

Assessment of enzyme linked immunosorbent assay based diagnostic kits (Toxokit-G and Toxokit-M) for the detection of IgG and IgM antibodies to Toxoplasma gondii in human serum

An enzyme linked immunosorbent assay (ELISA) using penicillinase was developed in the form of diagnostic kits (Toxokit-G and Toxokit-M) for the detection of IgG and IgM antibodies to Toxoplasma gondii. The performance of both the kits was compared with commercially available diagnostic kits, i.e. Enzygnost-Toxoplasmosis/IgG (Behring Co., Germany), TOXOTEK-G (Flow Lab., U.K.) and Toxoplasma IgM Microassay (Diamedix Corp., U.S.A.) by testing toxoplasma-suspected human serum samples. The results indicate a good reliability between these diagnostic kits. Toxokit-G has 86.66 and 96.05% sensitivity and specificity respectively. The main advantage of Toxokit-G is that the end result can be assessed visually without using sophisticated instruments. Toxokit-M has 100% sensitivity and specificity and test results were not affected by the presence of antitoxoplasma IgG antibodies, rheumatoid factor or antinuclear antibodies.     

A competitive ELISA for the specific diagnosis of contagious bovine pleuropneumonia (CBPP)

A competitive ELISA, using a specific monoclonal antibody, was designed to detect antibodies to Mycoplasma mycoides subsp. mycoides SC, the agent of contagious bovine pleuropneumonia. One monoclonal antibody was found suitable for such a test, ‘117/5', it does not cross-react with any of the other mycoplasma species tested, furthermore, its binding is inhibited by positive sera. The cutoff, 50% of inhibition, was determined using a set of negative sera from CBPP-free areas. The sensitivity was controlled with sera from artificially infected animals as well as from sera from areas where CBPP is enzootic. In both cases, cELISA compared favorably with CFT. The precocity of detection was similar but cELISA detected more positives and the positive titers seemed to persist longer than in the case of CFT. Lysis of the antigen used to coat the ELISA plates reduced the variability of fixation and improved the repeatability of the test. A field evaluation is now in progress which will determine the true sensitivity and specificity of the test and also check if antibodies are detected after vaccination.     

Evaluation of a Western Blot and ELISA for the detection of anti-Trichinella-IgG in pig sera

Human trichinellosis is a foodborne disease caused by ingestion of infective Trichinella muscle larvae via pork or meat of other food animals which are susceptible to this zoonotic parasite. There are new approaches for a risk-oriented meat inspection for Trichinella in pigs which are accompanied by monitoring programmes on herd level to control freedom from this parasite. For this purpose, testing schemes utilizing serological tests with a high sensitivity and specificity are required.This study aimed at the evaluation of an ELISA and a Western Blot (WB) for the detection of anti-Trichinella-IgG in terms of sensitivity and specificity taking results of artificial digestion as gold standard. For this purpose, 144 field sera from pigs confirmed as Trichinella-free as well as 159 sera from pigs experimentally infected with T. spiralis (123), T. britovi (19) or T. pseudospiralis (17) were examined by ELISA (excretory–secretory antigen) and WB (crude worm extract). Sera from pigs experimentally infected with four other nematode species were included to investigate the cross-reactivity of the antigen used in the WB. For all Trichinella-positive pig sera, band pattern profiles were identified in the WB and results were analysed in relation to ELISA OD% values.Testing of pig sera revealed a sensitivity of 96.8% for the ELISA and 98.1% for the WB whereas the methods showed a specificity of 97.9 and 100%, respectively. WB analysis of Trichinella-positive pig sera revealed five specific band patterns of 43, 47, 61, 66, and 102 kDa of which the 43 kDa protein was identified as the predominant antigen. The frequency of the band pattern profile was irrespective of the dose and the period of infection as well as the Trichinella species investigated.In conclusion, monitoring in swine farms for Trichinella antibodies should be based on screening pig sera by means of ELISA followed by confirmatory testing through WB analysis.     

Comparison of various tests for the serological diagnosis of Trypanosoma equiperdum infection in the horse

Comparative tests such as FAT, ELISA, RIA, IEO and CF in the diagnosis of dourine in the horse have proved a satisfactory concordance ratio of the ELISA with CF, which seems to be the most reliable test. Discrepancies have been observed as to the other tests which appear less sensitive than CF test.     

Sarcocystis neurona infections in raccoons (Procyon lotor): evidence for natural infection with sarcocysts, transmission of infection to opossums (Didelphis virginiana), and experimental induction of neurologic disease in raccoons

Equine protozoal myeloencephalitis (EPM) is a serious neurologic disease of horses in the Americas and Sarcocystis neurona is the most common etiologic agent. The distribution of S. neurona infections follows the geographical distributions of its definitive hosts, opossums (Didelphis virginiana, Didelphis albiventris). Recently, cats and skunks were reported as experimental and armadillos as natural intermediate hosts of S. neurona. In the present report, raccoons (Procyon lotor) were identified as a natural intermediate host of S. neurona. Two laboratory-raised opossums were found to shed S. neurona-like sporocysts after ingesting tongues of naturally-infected raccoons. Interferon-gamma gene knockout (KO) mice fed raccoon-opossum-derived sporocysts developed neurologic signs. S. neurona was identified immunohistochemically in tissues of KO mice fed sporocysts and the parasite was isolated in cell cultures inoculated with infected KO mouse tissues. The DNA obtained from the tongue of a naturally-infected raccoon, brains of KO mice that had neurological signs, and from the organisms recovered in cell cultures inoculated with brains of neurologic KO mice, corresponded to that of S. neurona. Two raccoons fed mature S. neurona sarcocysts did not shed sporocysts in their feces, indicating raccoons are not likely to be its definitive host. Two raccoons fed sporocysts from opossum feces developed clinical illness and S. neurona-associated encephalomyelitis was found in raccoons killed 14 and 22 days after feeding sporocysts; schizonts and merozoites were seen in encephalitic lesions.     

Use of echocardiography for the diagnosis of heartworm disease in cats: 43 cases (1985-1997)

OBJECTIVE: To determine the usefulness of echocardiography in the diagnosis of heartworm disease in cats and to compare this modality with other tests. DESIGN: Retrospective study. ANIMALS: 43 cats with heartworm infection that had echocardiographic examinations at 2 veterinary teaching hospitals between 1985 and 1997. Twenty-two of these 43 cats also underwent radiography of the thorax and heartworm antibody and heartworm antigen testing. PROCEDURE: Cats were determined to be infected with Dirofilaria immitis infection on the basis of 1 or more of the following findings: positive modified Knott or antigen test result, echocardiographic evidence of heartworm disease, or confirmation of the disease on postmortem examination. The percentage of echocardiographs in which heartworms were evident was compared with the percentage of radiographs in which pulmonary artery enlargement was evident and results of antigen or antibody tests in cats in which all tests were performed. RESULTS: Overall, heartworms were detectable by use of echocardiography in 17 of 43 cats, most often in the pulmonary arteries. In the 22 cats in which all tests were performed, antibody test results were positive in 18, antigen test results were positive in 12, and pulmonary artery enlargement was evident radiographically and heartworms were identifiable echocardiographically in 14. Heartworm infection was diagnosed exclusively by use of echocardiography in 5 cats in which the antigen test result was negative. CONCLUSIONS AND CLINICAL RELEVANCE: Although echocardiography was less sensitive than antigen testing, it was a useful adjunctive test in cats that had negative antigen test results in which there was a suspicion of heartworm disease. The pulmonary arteries should be evaluated carefully to increase the likelihood of detection of heartworms echocardiographically.     

Use of ELISA employing Leishmania (Viannia) braziliensis and Leishmania (Leishmania) chagasi antigens for the detection of IgG and IgG1 and IgG2 subclasses in the diagnosis of American tegumentary leishmaniasis in dogs

American tegumentary leishmaniasis (ATL) shows a reduced humoral response in dogs and levels of specific antibodies may therefore not be detected by indirect immunofluorescence. Although the sensitivity of enzyme-linked immunosorbent assay (ELISA) is higher than that of indirect immunofluorescence, the best antigen for the diagnosis of ATL in dogs has not been defined. The detection of IgG subclasses represents an alternative to increase the efficiency of the serological diagnosis. In Rio de Janeiro, sporotrichosis is the main differential diagnosis of ATL in dogs, and a sensitive, specific and little invasive method that permits the discrimination of the two diseases is desired. In the present study, 69 serum samples, 34 obtained from dogs with ATL and 35 from dogs with sporotrichosis, all of them with a confirmed etiological diagnosis, were tested. The samples were analyzed by ELISA using Leishmania (Viannia) braziliensis and L. (L.) chagasi antigens for the detection of anti-Leishmania IgG, IgG1 and IgG2. The use of L. (V.) braziliensis antigens for the detection of IgG and IgG2 yielded the best results. Using L. (L.) chagasi antigen, the sensitivity and specificity for the detection of IgG were 82.4% and 100%, respectively, whereas both sensitivity and specificity were 97.1% with the L. (V.) braziliensis antigen. No improvement in the performance of the test was observed when IgG2 was analyzed separately. The IgG1 assays presented low accuracy, irrespective of the antigen used: sensitivity and specificity of 58.8% and 60% for L. (V.) braziliensis and of 64.7% and 77.1% for L. (L.) chagasi, respectively. The present results suggest that IgG ELISA using the L. (V.) braziliensis shows the best performance for the diagnosis of ATL, permitting the discrimination between cases of ATL and sporotrichosis in dogs.     

Two new records of helminth parasites of domestic cat from Uruguay: Alaria alata (Goeze, 1782) (Digenea, Diplostomidae) and Lagochilascaris major Leiper, 1910 (Nematoda, Ascarididae)

Eight helminth taxa were recovered from the necropsy of four stray domestic cats from Colonia Miguelete, county of Colonia, Uruguay. Two of them are recorded for the first time for domestic cats in that country: Alaria alata (Goeze, 1782) from the small intestine (which is also the first trematode species found in domestic cat in Uruguay), and Lagochilascaris major Leiper, 1910 from the pharynx. The remaining helminth species found were Toxocara mystax (Zeder, 1800) and Spirometra sp. from the small intestine, Trichuris serrata (von Linstow, 1879) from the caecum, Eucoleus aerophilus (Creplin, 1839) from the trachea, and Pearsonema feliscati (Diesing, 1851) from the urinary bladder. Moreover, four female specimens of an unidentified Spiruroidea were collected from the stomach and small intestine of one host.     

Validation of an ELISA method for the serological diagnosis of canine brucellosis due to Brucella canis

In the present study, the validation of an enzyme-linked immunosorbent assay (ELISA) for serodiagnosis of canine brucellosis is described. Two different antigenic extracts, obtained by heat or ultrasonic homogenization of microbial antigens from a wild isolate of Brucella canis bacteria, were compared by ELISA and Western blot (WB). A total of 145 canine sera were used to define sensitivity, specificity and accuracy of the ELISA as follows: (1) sera from 34 animals with natural B. canis infection, confirmed by blood culture and PCR, as well as 51 sera samples from healthy dogs with negative results by the agar–gel immunodiffusion (AGID) test for canine brucellosis, were used as the control panel for B. canis infection; and (2) to scrutinize the possibility of cross reactions with other common dog infections in the same geographical area in Brazil, 60 sera samples from dogs harboring known infections by Leptospira sp., Ehrlichia canis, canine distemper virus (CDV), Neospora caninum, Babesia canis and Leishmania chagasi (10 in each group) were included in the study. The ELISA using heat soluble bacterial extract (HE-antigen) as antigen showed the best values of sensitivity (91.18%), specificity (100%) and accuracy (96.47%). In the WB analyses, the HE-antigen showed no cross-reactivity with sera from dogs with different infections, while the B. canis sonicate had various protein bands identified by those sera. The performance of the ELISA standardized with the heat soluble B. canis antigen indicates that this assay can be used as a reliable and practical method to confirm infection by this microorganism, as well as a tool for seroepidemiological studies.     

Evaluation of an enzyme-linked immunosorbent assay (ELISA) for the serological diagnosis of canine sarcoptic mange

Abstract The purpose of this study was to evaluate a serodiagnostic test (enzyme-linked immunosorbent assay; ELISA) for sarcoptic mange in dogs and to characterize the assay antigen, based on the mite Sarcoptes scabiei var. vulpes. The ELISA, applied to sera from 359 dogs suspected of having sarcoptic mange, showed a sensitivity and specificity of 92 and 96%, respectively. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) of the antigen employed in the ELISA revealed polypeptide bands with molecular weights ranging between 14 and 164 kDa. In Western blot analyses antigens of molecular weights between 62 and 64 kDa dominated. Particularly dominant were antigens of 164 and 147 kDa. These were found to have isoelectrical points in the range of 5.7–6.9. Sera from dogs infected with Cheyletiella sp., Demodex canis, Linognathus setosus and Otodectes cynotis, as well as from dogs allergic to fleas, were negative in the ELISA. Résumé— Le but de cette étude est d'évaluer un test sérologique ELISA pour le diagnostic de la gale sarcoptique chez le chien et de caractériser l'antigène révélateur, extrait de l'acarien Sarcoptes scabiei var. vulpes. Le test ELISA, lors d'une étude conduite avec les sérums de 359 chiens suspects de gale sarcoptique a démontré une sensibilité et une spécificité de 92 et 96%, respectivement. L'électrophorèse en gel polyacrilamide dodécyl sulfate de sodium de l'antigène utilisé dans l'ELISA a révélé des bandes polypeptidiques de poids moléculaire compris entre 14 et 164 kDa. Dans l'analyse en Western blot, les antigènes de poids moléculaire compris entre 62 et 164 kDa étaient les plus abondants, notamment ceux de 164 et 147 kDa. Ces derniers ont des points isoélectriques compris entre 5.7 et 6.9. Les sérums de chiens infectés par des Cheyletiella sp. Demodex canis, Linognathus setosus et Otodectes cynotis, ou par des chiens allergiques aux puces, se sont révélés négatifs en ELISA. [Bornstein, S., Thebo, P., Zakrisson, G. Evaluation of an enzyme-linked immunosorbent assay (ELISA) for the serological diagnosis of canine sarcoptic mange (Evaluation d'un test ELISA pour le diagnostic sérologique de la gale sarcoptique canine). Veterinary Dermatology 1996; 7 : 21–28.] Resumen El objetivo de este estudio fue el de evaluar una pruba serodiagnóstica (prueba de inmunoadsorción ligada a enzima; ELISA) para la sarna sarcóptica en el perro y caracterizar el antigeno prueba, basado en el ácaro Sarcoptes scabei, var. vulpes. El ELISA, aplicado a sueros de 359 perros sospechosos de padecer sarna sarcóptica, mostró una sensibilidad y especificidad del 92 y 96%, respectivamente. La electroforesis en gel de poliacrilamida dodecil sulfato sódico (SDS-PAGE) del antigeno usado en el ELISA reveló bandas de polipétidos con peso molecular entre 14 y 164 kDa. En el análisis Western blot, predominaron los antigenos de pesos moleculares entre 62 y 164 kDa. Los antigenos entre 164 y 147 kDa fueron especialmente predominantes. Estos tuvieron puntos isoeléctricos entre 5.7 y 6.9. Los sueros de perros infectados por Cheyletiella sp., Demodex canis, Linognathus setosus y Otodectes cynotis, asi como el de perros alérgicos a las pulgas fueron negativos en el ELISA. [Bornstein, S., Thebo, P., Zakrisson, G. Evaluation of an enzyme-linked immunosorbent assay (ELISA) for the serological diagnosis of canine sarcoptic mange (Evaluation de una prueba de immunoadsorcion ligada a enzima (ELISA) para el diagnostico serologico de la sarna sarcóptica canina). Veterinary Dermatology 1996; 7 : 21–28.] Zusammenfassung— Ziel dieser Studie war, einen Serodiagnostiktest (Enzyme-Linked-Immunosorbent-Assay, ELISA) für Sarkoptesräude des Hundes zu überprüfen und das Testantigen zu charakterisieren, das auf der Milbe Sarcoptes scabiei var. vulpes basiert. Der ELISA-Test, der bei den Sera von 359 Hunden mit Sarkoptesverdacht angewendet wurde, zeigte eine Sensitivität von 92% bzw. 96%. Die Natriumdodecylsul-fatpolyacrylamid-Gelelektrophorese (SDS-PAGE) des Antigen, das im ELISA verwendet wurde, zeigte Polypeptid-Banden mit Molekulargewichten zwischen 14 und 164 kDa. In der Wester-blot-Analyse dominierten Antigene mit einem Molekulargewicht zwischen 62 und 164 kDa. Besonders dominierend waren Antigene von 164 und 147 kDa. Bei diesen stellte man isoelektrische Punkte im Bereich von 5,7 bis 6,9 fest. Die Sera von Hunden, die mit Cheyletiella sp., Demodex canis, Linognathus setosus und Otodectes cynotis infiziert waren, fielen ebenso wie die Hunde mit Allergie auf Flöhe im ELISA negativ aus. [Bornstein, S., Thebo, P., Zakrisson, G. Evaluation of an enzyme-linked immunosorbent assay (ELISA) for the serological diagnosis of canine sarcoptic mange (Die Auswertung eines Enzym-Linked-Immunosorbent-Assay (ELISA) für die serologische Diagnose der kaninen Sarkoptesräude). Veterinary Dermatology 1996; 7 : 21–28.]     

Flock-level factors associated with the risk of Mycobacterium avium subsp. paratuberculosis (MAP) infection in Greek dairy goat flocks

In this cross-sectional study we identified flock-level risk factors for Mycobacterium avium subsp. paratuberculosis (MAP) infection, in Greek dairy goat flocks. We collected 1599 milk samples from does that were at the last stage of lactation in 58 randomly selected dairy goat flocks, during May to September 2012. The collected samples were tested with a commercial milk ELISA (IdexxPourquier, Montpellier, France) and the results were interpreted at a cut-off that optimized the accuracy of the diagnostic process. For the analysis of the data we used Bayesian models that adjusted for the imperfect Se and Sp of the milk-ELISA. Flock was included as a random effect. Does in flocks that used common water troughs and communal grazing grounds had 4.6 [95% credible interval (CI): 1.5; 17.4] times higher odds of being MAP-infected compared to does in flocks that had no contact with other flocks. Does of flocks supplied with surface water from either streams or shallow wells had 3.7 (1.4; 10.4) times higher odds of being infected compared to those in flocks watered by underground and piped water sources. When kids were spending equal to or more than 10 h per day with their dams they had 2.6 (1.1; 6.4) times higher odds of being MAP infected compared to kids that were separated from their dams for less than 10 h per day. Finally, does in flocks that continuously used the same anti-parasitic compound had 2.2 (1.0; 4.6) times higher odds of MAP infection compared to those in flocks alternating anti-parasitic compounds. These results should be considered in the development of a nationwide future control program fοr caprine paratuberculosis in Greece.