A Literature Review of Neospora caninum Infection in Humans

Background: Neosporosis is a parasitic disease caused by Neospora caninum. This parasite is an obligate intracellular coccidia similar to Toxoplasma gondii with a global distribution. With regard to the experimental studies, vertical transmission of the parasite in the monkey (non-human primates) has increased the concern about the zoonotic potential of this disease. The principal aim of the current research was to perform a minireview on investigations regarding the Neospora infection in humans on a global scale for the first time. Methods: All peer-reviewed articles (published until April 2021) on the Neospora infection in humans were searched in English databases such as Google Scholar, ScienceDirect, Scopus, PubMed, and ProQuest. Results: Based on data in the available articles, the presence of antibodies against the Neospora infection was between 0 and 37.7% in people from different countries. The seroprevalence rate of this infection in HIVpositive individuals was higher (26.6% and 37.7%) compared to other cases. Finally, the genomic DNA of Neospora was detected up to 1% using molecular biology techniques. Conclusions: Overall, the detection of anti-Neospora antibodies in humans indicated that people have been exposed to the parasite. Comprehensive research studies are essential for clarifying the risk factors associated with the Neospora infection in humans. This report provides the baseline information for future researchers. Molecular investigations and genotypic works on N. caninum isolates are highly recommended as well.


Background
Neosporosis caused by Neospora caninum, a Toxoplasmalike parasite belonging to obligate intracellular coccidian from the Toxoplasmatidae family, is a common parasitic disease in animals around the world (1). The Neospora infection was first distinguished in six Norwegian Boxer puppies in 1984. Neuromuscular problems such as encephalitis and myositis were the predominant clinical findings in all sick dogs (2). This parasite was misdiagnosed by Toxoplasma gondii before its introduction (3).
Meanwhile, a wide range of animals and birds are definitive and intermediate hosts for N. caninum. Domestic dogs and dairy cattle are commonly definitive and intermediate hosts in the life cycle of this parasite, respectively (3). Interestingly, dogs may simultaneously play a role in both final and intermediate hosts (4). The infection has been accounted for various species of warm-blooded vertebrates, some of which may serve as intermediate hosts in domestic and sylvatic cycles (3,4).
Neosporosis is a serious disease in animals. The significant role of the disease in abortion and other reproductive failures in cattle is clear (4,5). The annual economic losses related to the Neospora infection have been estimated at more than US$1.3 billion on a global scale (6). The detection of this infection is possible by different laboratory methods such as serology and molecular biology in animals and humans (1,4).
Currently, there is no report on clinical neosporosis in humans (7). Regarding the close phylogenetic relationship between N. caninum and T. gondii, as well as a wide range of intermediate hosts, the possibility of the Neospora infection in humans is undeniable (8), and the pathology, immunology, and epidemiology aspects of the infection in humans must be further studied accordingly (9). N. caninum is an important cause of fatal infections through experimentally transferring to pregnant cases with the lesions closely resembling those caused by congenital toxoplasmosis (10). A different level of antibodies to the Neospora infection have been detected in humans' sera (3,4). N. caninum was successfully cultured in various cell lines of humans. Furthermore, Rhesus monkeys (Macaca mulatta) were experimentally infected with N. caninum (11,12). Barr et al (11) demonstrated the vertical transmission of this parasite in monkeys, reinforcing the concern about the zoonotic potential of the disease. In this regard, the present study mainly aimed to first review the global investigations on the Neospora infection in humans.

Discussion
Zoonotic diseases are a developing concern because of their novel and erratic nature, as well as their fast circulation and ability to emerge anywhere. To design the Lunch Control Programs of zoonosis, it is essential to obtain knowledge on the risk factors and epidemiology of diseases in animals (22). Most clinical manifestations of neosporosis in animals are similar to those of toxoplasmosis (1). The tachyzoite form of the parasite may be disseminated in various tissues (i.e. blood, placenta, and amniotic fluid). Although no convincing evidence exists indicating that N. caninum effectively infects humans, there is still a concern and ambiguity for transmitting the infection (23). Serologic findings confirmed humans' exposure to this parasite, especially in immunodeficiency people (Table 1) although complementary works are essential to determine the extent and significance of humans' exposure (15). Neosporosis is yet an uncertain issue in medical infectious diseases (4,7). Regarding the high frequency of the Neospora infection in the transplacental transmission mode of cows (up to 90%), and the close similarity with the T. gondii infection, the possibility of Neospora posing a risk for pregnant women should receive special attention (24).
In an experimental work by Carvalho et al (25), the transplacental transmission and teratogenic lesions of the N. caninum infection were found in non-human primates parallel to histopathological lesions caused by T. gondii. Additionally, humans' cervical cells and trophoblasts were successfully infected by the tachyzoite form of N. caninum in vitro. The differences in susceptibility to the infection, cytokine production (type and rate), and cell viability were calculated in this evaluation (25). Some studies also focused on humans' neosporosis in the world. There is a limit of knowledge on epidemiology, pathology and the zoonotic aspect of Neospora-infection. The overall prevalence of the Neospora infection was estimated 17.14% (95% CI: 15.25-19.10%), 20% (95% CI: 18-21%), and 48.4% (95% CI: 47.5-49.3%) in dogs, cattle, and buffaloes, respectively, and 13.46% (95% CI: 10.26-17.42%) in horses and donkeys in different countries (26)(27)(28)(29).
Nam et al (13) first reported the Neospora infection in humans from Korea. In this work, the seroprevalence rate was 4.6% (13/282) in blood donors. In Brazil, antibodies to the Neospora infection were detected between 26.6% and 37.7% in HIV-positive people, as well as 18%, 10.5%, and 5% in patients with neurological signs, farmworkers, and newborn children, respectively (10,16,17). Moreover, no antibodies to the Neospora infection were found in women with a history of abortion and genitally failures in serological studies by Petersen et al (8) and Trees and Williams (18). On the other hand, the genomic DNA of Neospora was not detected in cases (n = 600) with clinical manifestations in favor of toxoplasmosis and negative for the T. gondii infection in molecular evaluations. According to their opinions, there was an unlikely opportunistic zoonotic agent (21).
The seroprevalence rate of infection to N. caninum and T. gondii in pregnant women was 24.3% and 26.8% using the immunofluorescence antibody test (IFAT), respectively (9). Two samples of the cord blood from Neospora-seropositive humans were positive using the molecular biology assay. Direct sequencing showed 98-99% homology compared to the reported strains from other countries. Based on the findings, tissue cysts and/or inflammatory infiltrate lesions related to the Neospora infection were not observed in histopathology examinations. There was a statistically significant association between seropositivity to the Neospora infection and the presence of domestic animals (P = 0.039), as well as dogs (P = 0.038) in the studied regions. This research acquired significant findings in terms of the Neospora infection in humans based on both serology and molecular biology tests (9).
With regard to the close biologic similarity between N. caninum and T. gondii, along with the possible presence of the parasite in immunocompromised individuals, it has been speculated that the Neospora infection can be transmitted to humans (8,10). According to Graham et al (14), the presence of N. caninum in the cotyledonary villi of the bovine placenta can be a risk option and a possible source for humans' infection, especially in farmers and veterinarians. The infected dogs easily contaminate the environment life of humans by excreting the parasite oocytes through their feces (5). Due to the close contact of humans with dogs (pet dogs and sheepdogs are common in urban and rural regions, respectively), the chance of humans' exposure to the Neospora infection is extremely high (15). The assessment of tissue liquids in individuals with immune deficiencies and fetuses suspected with toxoplasmosis during pregnancy could confirm that a subpopulation of the patients were infected with N. caninum (15). A significant association was detected between the seroprevalence of N. caninum and T. gondii infections in HIV-positive people and those with Neosporosis in Humans neurological problems (10). In a report by Graham et al (14), the Neospora-seropositive sera samples were negative for the T. gondii infection, which was not responsible for cross-reactivity. No cross-reactivity was found between N. caninum and T. gondii infections in serology examinations with IFAT (4). However, using laboratory methods with high sensitivity and specificity is suggested for avoiding false-positive and false-negative reactions.
Different options such as the main purpose for designing the research, sampling and sample size, and laboratory diagnostic methods, as well as climatic and environmental factors of the studied location are crucial in the reported results in Table 1 (24). The sporulation (time and rate) and the survival of the oocysts in the environment with different temperature and air humidity rates are of various types (3).
A wide range of serological diagnostic methods with different sensitivity and specificity rates have been used in previous studies (8,10,12,13). An investigation is recently conducted based on the molecular biology technique (21). The sensitivity of enzyme-linked immunosorbent assay and specificity of IFAT are higher compared to other serologic methods (3,4). Toxoplasma and Neospora were classified in the same family (Toxoplasmatidae). Thus, the rate of infections and clinical manifestations should be high in immunosuppressed individuals. For this reason, the seroprevalence rate of the Neospora infection was high (26.6% and 37.7%) in HIV-positive humans in Brazil (10,17).

Conclusions
This literature review is the first one to focus on the Neospora infection in humans. It was found that the presence of antibodies to N. caninum is due to humans' exposure to the parasite. A comprehensive and systematic research study is essential for identifying the risk options related to the Neospora infection in humans. In addition, the figure of the Neospora infection and its complications must be properly identified during pregnancy. The findings may contribute to the implementation of diagnostic tests in routine prenatal screening, especially in people with impaired immune systems. The results of this research can be used as the baseline information for designing and expanding future studies. Therefore, further research using molecular biology tools should be conducted to detect the genomic DNA of Neospora, as well as the genotypic diversity of the isolates.