This study was conducted to determine seroprevalence of Neospora infection in cattle, sheep, horses, donkeys, and dogs in Hamedan province, Iran. Blood samples (n=2254) from the animals were collected randomly during 2009 to 2012. Sera were prepared from the collected blood samples, which were then examined for the presence of antibodies against Neospora using enzyme-linked immunosorbent assay (ELISA), Neospora modified direct agglutination test (N-MAT), and indirect fluorescent antibody test (IFAT). The seroprevalence rates of Neospora were found as 17.4% (n=245/1406) in cattle, 2.2% (n=8/358) in sheep, 40.8% (n=49/120) in horses, 52% (n=52/100) in donkeys, and 27% (n=73/270) in dogs. In this study, higher levels of Neospora infection were detected in cattle, horses, donkeys, and dogs. This is the first comprehensive study of Neospora infection in domestic animals in Iran. Further researches on molecular and bioassay studies and designing appropriate control strategies against neosporosis in Iran are necessary and strongly recommended.

Objective--To evaluate the immunologic response of a killed tachyzoite vaccine against Neospora caninum and its effectiveness in preventing fetal loss associated with experimentally induced neosporosis in sheep. Animals--30 Dorset ewes. Procedure--Ewes were randomly allocated to receive vaccination on days 1 and 60 of the study with a killed N caninum tachyzoite preparation in a commercially available adjuvant or a saline-adjuvant mixture. A ram was placed on pasture with the ewes from days 15 to 60. Blood was collected from ewes before primary and booster vaccinations and prior to experimental challenge with N caninum tachyzoite performed on day 90; sera were assessed via Neospora agglutination (NA) and immunofluorescence antibody (IFA) assays. Blood was collected from lambs before they suckled, and sera were tested for antibodies against N caninum. Results--Of the 14 vaccinated ewes that became pregnant, 12 gave birth to live-born lambs; in contrast, 5 of 11 pregnant control ewes gave birth to live-born lambs. Whereas vaccination improved fetal survival in pregnant ewes challenged with N caninum tachyzoites, it did not appear to have any appreciable effect on transmission of N caninum to offspring, as indicated by results of NA and IFA assays. Conclusions and Clinical Relevance--The N caninum tachyzoite vaccine used in this study appeared to provide protection against fetal loss associated with experimentally induced neosporosis in a high proportion of pregnant ewes.

Neospora caninum-induced abortion is a major production problem in the daily cattle industry in the United States and worldwide. Abortions attributable to naturally acquired N caninum infection also have been observed in pygmy goats. We studied experimentally induced infections with N caninum in pregnant pygmy does to determine whether abortions attributable to N caninum infection would occur after inoculation. Seven pregnant pygmy does (1 control doe and 6 inoculated with N caninum) were studied. The control doe remained clinically normal throughout the study and delivered 2 healthy kids. Abortion, fetal death, and stillbirths were observed in some pregnant does inoculated with N caninum. Two pregnant pygmy does inoculated with N caninum early in gestation (day 51) had fetuses that died and were aborted, or died and were reabsorbed. Neospora caninum tachyzoites and lesions were observed in the brain, spinal cord, and heart of aborted fetuses; parasites also were isolated from the placenta. Four additional pregnant pygmy does (2 inoculated at mid-gestation [day 85], and 2 at late gestation [day 127]) did not abort after inoculation. However, 1 doe inoculated during mid-gestation delivered a stillborn fetus that had died about 1 week prior to parturition. This kid was congenitally infected with N caninum. Neospora caninum was isolated from the placentas of all inoculated does examined. Neonatal neosporosis was not observed in live-born kids, nor were stages of N caninum isolated from any live-born kid. Does did not undergo abortion or have congenitally infected kids when they were rebred and evaluated for neosporosis.

Neospora caninum causes serious disease in dogs, and it, or a similar parasite, is a major cause of abortion in cattle. Little is known about the susceptibility of this protozoan to antimicrobial agents. We studied several antimicrobial agents to determine which classes might have activity against this parasite. We also determined whether activity of such agents was coccidiocidal or coccidiostatic. A 2-day of treatment, monoclonal antibody-based enzyme immunoassay and a 5-day of treatment, cell culture flask (CCF), lesion-based assay were developed to examine the ability of test agents to inhibit tachyzoite multiplication. Seven sulfonamides were examined, with the following activities observed: sulfathiazole greater than or equal to sulfamethoxazole > sulfadiazine > sulfaquinoxaline greater than or equal to sulfamethazine > sulfadimethoxine > sulfamerazine. Dapsone, a sulfone, had little activity. Six dihydrofolate reductase/thymidylate synthase inhibitors were examined, with the following activities observed: piritrexim > pyrimethamine > ormetoprim > trimethoprim = diaveridine > methotrexate. Six ionophorous antibiotics were examined; lasalocid, maduramicin, monensin, narasin, and salinomycin had equivalent activities, but alborixin was toxic for host cells at the lowest concentration examined. Three macrolide antibiotics--azithromycin, clarithromycin, and erythromycin--were examined and had equivalent activities. Two tetracycline antibiotics, doxycycline and minocycline, were examined and had equivalent activities. Three lincosamide antibiotics were examined, with the following activities observed: clindamycin hydrochloride > clindamycin phosphate > lincomycin hydrochloride. Pentamidine and 6 of its analogs were examined, and only hexamidine and 1,4-Di[4-(2-imidazolinyl)-2-methoxy-phenoxylbutane had activity. Eight miscellaneous antiprotozoal agents were examined for activity. Amprolium, metronidazole, paromomycin, and roxarsone had little activity. Arprinocid, diclazuril, nitrofurazone, and robenidine had good activity. Eleven agents were examined in both assays, whereas 32 agents were examined in the CCF assay only. The enzyme immunoassay and CCF assay provided similar results for agents that rapidly killed tachyzoites. However, agents that inhibited development, but were not rapidly fatal for tachyzoites, had better activity in the CCF assay. Of the classes of agents examined, the dihydrofolate reductase/thymidylate synthase inhibitors, 2 of the 6 pentamidine analogs, and the ionophores were coccidiocidal and the sulfonamides, macrolides, tetracyclines, and lincosamides were coccidiostatic. Of the miscellaneous agents examined, arprinocid, nitrofurazone, and robenidine were coccidiocidal and diclazuril was coccidiostatic.

A murine monoclonal antibody (MAB) 6G7 generated against tachyzoites of Neospora caninum recognized 8 major and several minor antigens, as observed by western blot analysis. Relative rate of migration of the 8 major antigens ranged from 31 to 97.4 kd. In addition, MAB 6G7 recognized a Toxoplasma gondii tachyzoite antigen with a relative rate of migration of 107 kd. Immunogold labeling of N caninum tachyzoites grown in human foreskin fibroblast cells indicated that MAB 6G7 binds to micronemes, dense granules, basal portions of rhoptries, and intravacuolar tubules within the parasitophorous vacuole. Monoclonal antibody 6G7 also bound to micronemes and basal portions of rhoptries within tachyzoites of T gondii. Monoclonal antibody 6G7 did not significantly inhibit development of tachyzoites in vitro.

Neospora caninum is considered the main cause of bovine abortion worldwide. The correct laboratorial diagnose is very important to identify the infected animals and to apply control measure. The objetive of this study was to show the persistence period of colostral antibodies in calves. Eight newborn Holstein Friesan calves, males, were selected from N. caninum soronegative dams. Pre-colostral blood samples were collected of these calves and all of them were seronegative to N. caninum. They were fed with two liters of pooled colostrum from seropositive cows within two hours after birth. Blood samples were collected and tested weekly until the animals turned negative. Serum samples were tested for antibodies to N. caninum using indirect fluorescence antibody test at 1:50; 1: 100 and 1:200 dilutions. Antibodies were not detected from three out of eight calves and they were excluded from the study. The remaining 5 calves seroconverted in all dilutions at the fifth day after colostrums ingestion. At 1:50 dilution, one calf remained positive for 21 weeks, two for 20 weeks and one for 13 weeks. At 1:100, one calf was positive for 15 weeks and the remaining 4 calves for 13 weeks. At 1:200, each calf was positive for 1, 7, 12, 12 and 13 weeks, respectively. These results demonstrate that the colostral antibody to N. caninum may persist until 21 weeks after birth in calves and it's very important to exclud the calves at the first month of age in the seroprevalence studies to avoid the false-positive results. | Neospora caninum é considerado a principal causa de aborto bovino mundial. O diagnóstico laboratorial correto é muito importante para identificar os animais infectados e para aplicar medidas de controle. O objetivo deste trabalho foi mostrar o declínio de anticorpos colostral em bezerros. Este estudo empregou oito bezerros holandeses, recém-nascidos, machos, descendentes de vacas soronegativas para N caninum. Amostra de sangue pré-colostral foram colhidas destes bezerros e todos estavam soronegativos pra N. caninum. Estes bezerros foram alimentados com dois litros de um pool de colostro de vacas soropositivas dentro de duas horas após o nascimento. Amostras de sangue dos bezerros foram colhidas semanalmente até os animais soroconverterem negativo. As amostras foram testadas para anticorpos de N. caninum usando teste de imunofluorescência indireta nos títulos de 1:50; 1: 100 e 1:200. Os resultados mostraram que 3 dos 8 bezerros não soroconverteram e foram excluídos do estudo. Os restantes cinco bezerros soroconverteram em todos os títulos no quinto dia após a inoculação. No título 1:50, um bezerro permaneceu positivo por 21 semanas, dois por 20 semanas e um por 13 semanas. No título 1:100, um bezerro foi positivo por 15 semanas e o restante quatro bezerros por 13 semanas. No título 1:200, cada bezerro foi positivo por 1; 7; 12; 12 e 13 semanas, respectivamente. Estes resultados demonstram que o anticorpo colostral para N. caninum pode permanecer até 21 semanas após o nascimento nos bezerros e é muito importante excluir os bezerros até quatro meses de idade nos estudos de soroprevalência para impedir os resultados falso-positivos.