First detection and molecular identification of Neospora caninum from naturally infected cattle and sheep in North Africa

Neosporosis, caused by the protozoan Neospora caninum , is a major cause of reproductive failure in ruminants causing enormous economic losses. The objective of this study was to estimate the infection rate and molecular identification of N. caninum in Tunisian cattle and sheep. A total number of 348 meat samples were collected from 150 cows and 198 ewes slaughtered in the regional slaughterhouse of Béja (North‐west Tunisia) and tested for the presence of N. caninum ITS 1 gene using PCR followed by sequencing of some PCR products. A phylogenetic tree was then constructed to compare the partial sequences of the ITS 1 gene with GenBank sequences. The overall molecular infection prevalence of N. caninum was significantly higher in cattle than in sheep (22 and 10.6%, respectively, p  = .003). In sheep, the highest prevalence was observed in the northern Béja locality (31.2 ± 16.1), with the Noire de Thibar breed as the most infected sheep breed (31.7 ± 14.2%) (p  < .001). In cattle, there were no differences in the molecular prevalence of N. caninum according to breeds and localities. The association between age and N. caninum molecular prevalence was statistically significant in both species; the highest prevalence was observed in sheep of more than one year of age (19.4 ± 9.1%), and in cattle between two and eight years of age (28.8 ± 10.9%). Comparison of the partial sequences of the ITS 1 gene revealed 96%–100% similarity among our N. caninum amplicon and those deposited in GenBank. To our knowledge, this is the first detection and molecular identification of N. caninum in sheep and cattle in North Africa. This information is pertinent in designing control programmes that would reduce economic losses in the livestock industry.     

The development and evaluation of a nested PCR assay for detection of Neospora caninum and Hammondia heydorni in feral mouse tissues

The development of a novel nested polymerase chain reaction is described and used for detecting the presence of Neospora caninum and Hammondia heydorni DNA in DNA extracted from feral rodent tissues. A unique strategy was used for design of an assay that could be adapted for detecting DNA from more than one member of Toxoplasmatinae simultaneously with a minimal number of additional steps. The level of sensitivity described for this assay is comparable to real time-PCR and other nested PCR assays. Twenty-eight of 104 feral mice tested positive for N. caninum in at least one tissue (the brain, heart or liver) studied. In this study, eight instances are reported where the brain tested negative to N. caninum while at least one other tissue was positive. This suggests that prior studies, which screened only the brain, describe prevalence levels that are under-represented. None of 54 mouse brains tested positive for H. heydorni DNA. This suggests that mice are rarely infected by H. heydorni although this hypothesis needs to be explored further. Data obtained in the current study suggest that N. caninum is a common parasite of feral rodents which may be important in the epidemiology of the disease.     

Vaccination with the recombinant chimeric antigen recNcMIC3-1-R induces a non-protective Th2-type immune response in the pregnant mouse model for N. caninum infection

The major route of transmission of Neospora caninum in cattle is transplacentally from an infected cow to its progeny. Therefore, a vaccine should be able to prevent both the horizontal transmission from contaminated food or water and the vertical transmission. We have previously shown that a chimeric vaccine composed of predicted immunogenic epitopes of NcMIC3, NcMIC1 and NcROP2 (recNcMIC3-1-R) significantly reduced the cerebral infection in BALB/c mice. In this study, mice were first vaccinated, then mated and pregnant mice were challenged with 2 × 10⁶N. caninum tachyzoites at day 7–9 of pregnancy. Partial protection was only observed in the mice vaccinated with a tachyzoite crude protein extract but no protection against vertical transmission or cerebral infection in the dams was observed in the group vaccinated with recNcMIC3-1-R. Serological and cytokine analysis showed an overall lower cytokine level in sera associated with a dominant IL-4 expression and high IgG1 titers. Thus, the Th2-type immune response observed in the pregnant mice was not protective against experimental neosporosis, in contrary to the mixed Th1-/Th2-type immune response observed in the non-pregnant mouse model. These results demonstrate that the immunomodulation that occurs during pregnancy was not favorable for the protection against N. caninum infection conferred by vaccination with recNcMIC3-1-R.     

Immunity to experimental neosporosis in pregnant sheep

Neospora caninum is an important cause of fetal loss in cattle but has also infrequently been shown to cause disease in sheep and goats. Experimental infection of pregnant sheep with N. caninum causes clinical and pathological changes very similar to those of neosporosis in cattle. An experiment in sheep was undertaken to examine whether infection with N. caninum before pregnancy conferred immunity to subsequent challenge with the parasite during pregnancy. Primary inoculation of NC1 tachyzoites subcutaneously, either before or during pregnancy, caused a significant temperature response in ewes, while those given a secondary challenge at 90 days gestation (dg) did not show such a response. Primary infection of 12 ewes during pregnancy resulted in the loss of all fetuses while a further 12 ewes inoculated with NC1 tachyzoites before mating and subsequently challenged with the same dose at 90 dg produced nine live and seven dead lambs. There were no fetal deaths in ewes only infected with Neospora before mating although there was serological evidence of vertical transmission in four of their clinically normal offspring while Neospora DNA was detected in the cerebrospinal fluid of a fifth healthy lamb. Thus an experimental primary infection with N. caninum during pregnancy killed all the fetuses while inoculation before pregnancy did not cause any mortality but did provide a degree of protection against subsequent challenge with Neospora during pregnancy.     

犬钩虫第三期钩蚴免疫裸鼠再感染AcL3的组织病理学观察

目的 观察裸鼠经犬钩虫第三期钩蚴(AcL3)免疫后,其皮肤和肺内AcL3的形态变化及宿主的组织细胞反应,评价其作为疫苗筛选动物模型的可能性. 方法取BALB/c-nu/nu 小鼠,每两周由皮下免疫接种活的AcL3 500条,共3次,并丁末次免疫后1周由皮肤攻击感染AcL3 500条.用未免疫的感染AcL3 裸鼠作对照,攻击感染后不同时间取感染部位皮肤和肺脏,观察宿卡皮肤和肺内AcL3的组织病理学变化. 结果攻击感染后6、24、72 h及7 d,皮肤内的绝人部分虫体切面形态和组织结构与感染对照裸鼠皮肤内的相似,仪0.5%～2.2%的虫体切而示有变性、死亡,偶见有死虫肉芽肿,而肺组织内的AcL3及其周围的炎细胞反应对照组相比较,无明显差异. 结论用活的AcL3免疫裸鼠后,仅见对攻击感染的AcL3 有弱的免疫效应,裸鼠不适宜作为钩虫疫苗筛选的动物模型.     

Protecting babies: vaccine strategies to prevent foetopathy in Neospora caninum-infected cattle

Neospora caninum is an apicomplexan protozoan parasite that is a significant infectious abortifacient agent in cattle. Despite the fact that it is a member of a well described taxonomic group, it is a relatively newly discovered parasite and its biology is not yet fully understood. Cattle become infected either congenitally via transplacental transmission or post-natally by ingesting oocysts derived from the definitive host; dogs and coyotes are the only definitive hosts that have been described to date. It is not known which of these two forms of transmission occurs most frequently and which is the most likely to result in abortion; there are no drugs available to treat infected cattle, so current control strategies rely on prevention of infection by management methods and strict hygiene; an effective vaccine would be a great advantage in its control. Neospora caninum is an economically important veterinary pathogen, but we can also draw analogies between its foetopathic effects and those of human pathogens such as Toxoplasma gondii, Chlamydophila abortus and Plasmodium falciparum. Understanding the immune response and the materno-foetal relationship in N. caninum-infected cattle may help us to design vaccination strategies, not only for neosporosis but also for other foetopathic agents.     

GoPubMed在药物研发信息挖掘中的应用

采用GoPubMed数据库的在线分析软件进行科技查新,对国内外1983-2012年发表的钩虫抗凝肽研究文献信息进行数据挖掘,获得其研究文献主题、年代、地区分布、期刊、核心作者等相关信息,探索与药物研发密切相关的重组基因产物和药物作用机制.     

Seroprevalence of Neospora spp. in Horses in North East of Iran

Background

Neospora caninum, an obligate intracellular protozoan parasite, is recognized as a major cause of abortion in cattle, while limited information is presently available on the seroprevalence of Neospora antibodies in horses’ worldwide. The aim of the present study was to determine serologic prevalence of Neospora infection in horses in Iran.

Methods

Sera from 150 horses from Mashhad suburb in Razavi Khorasan Province, northeast Iran were examined for antibodies to Neospora spp. using Neospora modified direct agglutination test (N-MAT).

Results

Antibodies to this parasite were detected in 45 (30%) of the examined serum samples. Thirty four percent of the samples had titer of 1:40 while then reduced to 30% when 1:80 serum dilution was applied as significant cut off titer.

Conclusion

This study is the first investigation carried out on the Neospora in horses in Iran and indicates that horses in Iran are exposed to this parasite.     

First report of Neospora caninum infection in pigs in China

Neospora caninum is a protozoan parasite which can infect many mammals and birds with a worldwide distribution. However, no molecular data are available about the occurrence of N. caninum in pigs. In this study, the serological and molecular prevalence of N. caninum infection in farmed pigs were investigated in Hunan province, China, between January 2017 and December 2018. A total of 1,500 serum samples collected from 10 herds in Hunan province were evaluated using a competitive‐inhibition enzyme‐linked immunoassay assay (cELISA). The overall seroprevalence of N. caninum in the examined pigs was 1.9%. The seroprevalence of N. caninum ranged from 0.3% to 4.6% among different regions in Hunan province of China (p < .05). DNA was extracted from brain samples, and the Nc‐5 gene and ITS‐1 region were amplified and then sequenced. Three (0.5%) of the examined 600 brain tissues were found to contain N. caninum DNA. Our phylogenetic analyses indicated that N. caninum samples were classified into two distinct groups. Although the prevalence is low within the pig groups investigated, our results revealed the emergence of N. caninum infection in pigs in China. The finding of the present study provides molecular evidence that the pigs are the natural intermediate host of N. caninum and may have major epidemiological importance.     

婴儿感染犬复孔绦虫的鉴定和治疗

目的鉴定和治疗一婴儿感染犬复孔绦虫.方法根据虫体节片的形态特征进行虫种鉴定,并用特效药驱虫治疗.结果本例为我国自1923年以来报道的第16例犬复孔绦虫人体感染,并再次证实应用吡喹酮治疗有特效.结论犬复孔绦虫病是一种在家养动物(犬、猫)中常见、在人体少见的人兽共患寄生虫病.在我国,随着社会经济的发展,豢养宠物之风渐盛,应警惕此病发展的可能.