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Published in final edited form as: Exp Parasitol. 2018 Dec 15;196:55–62. doi: 10.1016/j.exppara.2018.12.003

Systematic Review and Meta-Analysis of Variation in Toxoplasma gondii Cyst Burden in the Murine Model

Gabrielle F Watson 1, Paul H Davis 1,+
PMCID: PMC6447088  NIHMSID: NIHMS1007035  PMID: 30562481

Abstract

Toxoplasma gondii is an obligate intracellular protozoan parasite that infects approximately 30% of the population of the United States, with worldwide distribution. The chronic (latent) infection, mediated by the bradyzoite parasite life stage, has attracted attention due to possible links to host behavioral alteration and psychomotor effects. Mice are a common model organism for studying the chronic stage, as they are natural hosts of infection. Notably, published studies demonstrate vast ranges of measured cyst burden within the murine brain tissue. The inconsistency of measured cyst burden within and between experiments makes interpretation of statistical significance difficult, potentially confounding studies of experimental anti-parasitic approaches. This review analyzes variation in measured cyst burden in a wide array of experimental mouse infections across published literature. Factors such as parasite infection strain, mouse strain, mode of infection, and infectious dose were all examined. The lowest variation in measured cyst burden occurred with the commonly available Balb/c and CBA mice undergoing infection by the ME49 strain of T. gondii. A summary of cyst variation and average cyst counts in T. gondii mouse models is presented, which may be useful for designing future experiments.

Keywords: Toxoplasma gondii, Murine Models, Cyst Variability, Method of Infection, Chronic Infection

1. INTRODUCTION

Toxoplasma gondii (T. gondii) is an intracellular protozoan parasite capable of causing infection worldwide in nearly all warm-blooded vertebrates, typically through oral transmission of bradyzoite cysts or oocysts [Su, 2003]. Members of the Felidae family are considered the definitive host where T. gondii can sexually reproduce, resulting in oocyst production expelled through defecation. Prevalence in humans varies by age of individuals and geographic region due to differences in exposure to cat droppings containing T. gondii oocysts in contaminated food or water; undercooked meats bearing bradyzoite tissue cysts also serve as a significant reservoir [Gontijo da Silva, 2015].

Once the [oo]cysts are ingested, the parasites transition to the tachyzoite stage in the intestines and disseminate through the hosts’ tissue and bloodstream, eventually forming chronic infection in the muscle and brain tissue, characterized by the parasite’s bradyzoite stage [Randall, 2011]. Tissue cysts begin forming as early as a week after infection in the murine brain tissue and persist for the remainder of the host’s life. Cysts are known to increase diameter during the latent stage of infection in the brains of mice over time, possibly rupturing and reforming cysts in an immunocompetent host [Watts, 2015]. Immunocompromised individuals are at high risk of recrudescent disease due to intracellular brain cysts which can reactivate and potentially cause lethal destruction of neural tissue [Montoya, 2004].

The latent stage is an important parasitic stage for study, as it has been correlated with reduced response time of the host [Flegr, 2007]. Moreover, in rodent species, the fear response has been shown to be attenuated when the host has acquired an infection with T. gondii, leading to an increased probability of predation by cats, where the parasite can sexually reproduce [Gatkowska, 2012; Vyas, 2007]. T. gondii tissue cysts are commonly studied in the murine model, due to the animal’s natural infection in the wild with the majority of tissue cysts present in the brain [Dubey, 1998]. However, current experimental studies attempting to model or reduce cyst burden via treatment in mice frequently report significant variation of cyst counts between infected hosts. Such variability can be a chief limiting factor when interpreting the efficacy of experimental anti-parasitic interventions such as drug or vaccine candidates.

Methods for inducing bradyzoite stage infection in mice also vary considerably between reports in the literature. However, there has not been a body of work which categorizes the various infection models of T. gondii and correlates them to cyst load variability. We have compiled and analyzed literature describing latent infection in murine models and noted the corresponding cyst levels and variance. Key variables include: parasite stage during infection, mouse and parasite strain, and method of infection. Such data may prove valuable to future experimental designs to establish greater statistical significance for experimental treatments or neurological evaluations.

2. Materials and Methods

A comprehensive review of literature in PubMed identified papers pertaining to Toxoplasma gondii variability of cyst levels in a murine model using keywords: [Toxoplasma gondii cyst (variability) and (levels) and (burden) in mice] (October 2017). This search criteria returned 150 papers where 62 met criteria for study inclusion: reported mean cyst burden with standard deviation and written in English. The majority of these papers focus on the ability of drugs or drug-like molecules to reduce cyst burden or the behavioral effects in relation to cyst burden. Literature detailing parasite burden measured by qPCR was not considered, as the focus of this review is cyst number, and not total number of parasites. Literature providing cyst variability with standard error of mean or undefined or non-present error bars were also excluded. In cases where data was organized non-quantitatively (i.e. figure only) ImageJ (1.8.0_112) measurement tool was used with axial figure units to extrapolate the data of interest (mean and standard deviation). In the case of experimental group sample sizes that were reported in a range, the lowest number was used for analysis. Exotic parasite strains that were not tested in multiple strains of mice were not analyzed in figures but instead only listed in the charts. The coefficient of variation (CV) (standard deviation/mean *100) provides the ratio of standard deviation to mean.

3. CONDITIONS AFFECTING INFECTION

3.1. Parasite Strain

Classical Type II and III T. gondii strains are well known to form latent infections and constitute the majority of published works. T. gondii strains commonly seen in Europe and North America are either type I, II, or III and will carry genetic variation up to a maximum of 1% [Su, 2012; Saeji, 2005]. T. gondii type II strains are most commonly seen in immunocompromised patients in the first world and remain relatively non-lethal most likely due to the production of a less severe parasite burden on the individual compared to Type I/III strains [Saeji, 2005; Xiao, 2015]. Strain virulence is not a constant factor: it is known that continuous passage in animals can increase virulence compared to cell passage, but little research has established the significance or consistency of these changes [Albert, 1941].

3.2. Mouse Strain

It is known that the major histocompatibility complex (MHC) genes capable of triggering an effective Th1 response are primarily responsible for resistance to infection, relative cyst load, and the severity of lethal encephalitis [Resende, 2008; Miller, 2009]. Thus, differing mouse strains are known to present diverse symptoms and severity of disease, regardless of infection administration. Mouse strain responses are briefly summarized below.

Balb/c mice (an inbred strain) are commonly used to study toxoplasmosis due in part to their resistance against the latent infection, suggested to be a result of their MHC haplotype (D) [Resende, 2008; Affymetrix Inc.]. Balb/c are resistant to acute infection through oral inoculation and susceptible via the intraperitoneal route with cysts of the ME49 strain [Subauste, 2012]. However, they generally show resistance to chronic infection, ultimately leading to reduced cyst load [Subauste, 2012]. For instance, in Suzuki et al. (1993), Balb/c mice formed fewer cysts (213 ± 151) in the brain during latent infection compared to CBA/Ca (3,015 ± 1,704) when inoculated with 10 cysts IP.

The inbred stain C3H with MHC haplotype Hk, appear to be more resistant to acute challenge when challenged IP as compared to orally with 10 cysts, with nearly all mice surviving up to 21 days post inoculation [Johnson, 1984; Affymetrix Inc]. In mice receiving the IP injections consisting of cysts, C3H mice were only slightly more resistant than Balb/c and significantly more resistant than Swiss Webster (SW) [Johnson, 1984].

C57BL/6, inbred mice with a Hb haplotype, are susceptible to acute infection via the oral route with ME49 cysts but resistant when administered IP [Subauste, 2012]. Unlike Balb/c, these mice are susceptible to chronic infection leading to an increase in cyst burden [Subauste, 2012]. They are more susceptible to acute toxoplasmosis and weight loss than Balb/c, with a majority expiring after 60 days following acute infection of 5 DX T. gondii cysts [Gatkowska, 2006; Affymetrix Inc]. This mouse strain is proven to be more susceptible to lethal acute disease via oral infection compared to IP of 10 cysts, with oral infection causing around a 70% mortality [Johnson, 1984].

The inbred strain CBA mice (Hk) typically are sensitive to T. gondii infection and die several months after infection from T. gondii ME49 cysts through encephalitis [Suzuki, 1993; Affymetrix Inc], potentially due to their relatively high average of 3,000 cysts per brain from IP infection of 20 ME49 cysts [Subauste, 2012].

No significant difference was seen in outbred Swiss Webster (SW) mice infected orally or IP with the same infective dose (10 cysts), although a slightly higher mortality rate was seen with IP infection [Johnson, 1984]. Regarding oral acute infection, SW have greater survival than C3H and C57BL strains [Johnson, 1984].

The Kunming strain of outbred mice are exclusively used in Asia and present with a susceptibility to the acute stage of T. gondii [Wang, 2017]. When Kunming mice were infected with 1,000 or more RH strain (hyper-virulent) tachyzoites they typically died within a week [Li, 2015]. The mouse strain’s susceptibility to chronic infection is not clearly stated, mainly due to lack of articles studying Kunming mice.

3.3. Method of Infection Variability

Most infections using mice are delivered with intraperitoneal or oral inoculation, both with tachyzoites grown in tissue culture or bradyzoite cysts harvested from infected tissue. The route of infection determines dissemination of the parasites and the initial symptoms within the murine model. Mouse strains that are relatively more susceptible to oral inoculation are not necessarily susceptible to IP infection; however, IP infection generally leads to a more potent and rapid response compared to peroral. Mice infected via IP injection with 106 type II tachyzoites showed that parasites were confined to the abdomen for the first week and seen in the brain by day 12, forming cysts in clustered locations within the brain [Di Cristina, 2008].There appears to be more inconsistency when mice are orally inoculated, although this may be obfuscated as only a limited number of studies report whether mice were gavaged or fed orally. This clarification is important considering oral gavage leads to inconsistent migration of the parasite compared to the more natural method of feeding: when mice were orally gavaged, within a week many parasites were seen through in vivo imaging in the chest cavity, while only a subset initially demonstrated parasites in the intestines. This is starkly contrasted to natural feeding, where all the mice had parasites in their intestines within a week followed by the brain, regardless of the strain [Boyle, 2007].

4. VARIABILITY

4.1. Parasite Strain Variability

The T. gondii strain ME49 produces the lowest brain cyst variability in Balb/c, C3H, and CBA mice compared to other Toxoplasma strains, either through oral or intraperitoneal inoculation of cysts, or orally with tachyzoites. Kunming, Balb/c, and Swiss Webster mice display realtively low cyst varaibility when infected with T. gondii PRU strain (Tables 1, 2, and 3). Of note, when Swiss Webster and Balb/c were infected with PruΔku80, mice formed substainely less cysts than when infected with wildtype PRU parasites (Tables 2 and 3). Specifically, Swiss Webster have an average coefficent of variance (CV) of 34.7% when inoculated IP with PRU tachyzoites, compared to PRU cysts infecting Kunming mice orally with a variance of 8.8%. Balb/c mice present with a low variance when infected orally with PRU cysts (15.2%) and IP with PRU tachyzoites (25.8%). In contrast, the highest cyst variance displayed in the CBA strain at 45.6% was caused from IP infection with PRU tachyzoites (Table 4). The 76K T. gondii strain was tested only in C57BL (Table 5) and CBA mice, exhibiting the lowest CV in C57BL mice at 23.4%. DX cysts were injected IP produced the highest variance in C3H (Table 6) and C57BL mice. An uncommonly used T. gondii strain, Beverly, adminstered orally with cyst established the largest variance in Balb/c and a relatively low variance (24.5%) in C57BL/6 mice.

Table 1.

Kunming Variability in Cyst Load.

Kunming
Cyst Count (+−SD) Coefficient of Variation (Percentage) Infection Method Parasite Strain Brain Isolation in Weeks Post Infection Sample Size Reference
18080 ± 642 4 Oral 5 cysts PRU 4 5 Li, 2015
3963 ± 37 1 Oral 10 cysts PRU 4 6 Yang, 2017
3876 ± 37 1 Oral 10 cysts PRU 4 6 Yang, 2017
3167 ± 672 21 Oral 10 cysts PRU 4 6 Xu, 2014
3232 ± 534 17 Oral 10 cysts PRU 4 6 Xu, 2014
3167 ± 259 8 Oral 10 cysts PRU 4 3 Zhang, 2013
3233 ± 197 6 Oral 10 cysts PRU 4 3 Zhang, 2013
1956 ± 44 2 Oral 10 cysts PRU 4 3 Chen, 2015
1933 ± 67 4 Oral 10 cysts PRU 4 3 Chen, 2015
3337 ± 183 6 Oral 10 cysts PRU 4 6 Zhang, 2015
3276 ± 231 7 Oral 10 cysts PRU 4 6 Zhang, 2015
4296 ± 687 16 Oral 20 cysts PRU 5 6 Wang, 2017
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Table 2.

Balb/c Strain Cyst Variability.

Balb/c
Cyst Count (+−SD) Coefficient of Variation (Percentage) Infection Method Parasite Strain Brain Isolation in Weeks Post Infection Sample Size Reference
193 ± 58 30 Oral 20 cysts Beverly 8 N.D. Roberts, 1992
122 ± 79 65 10/20 cysts oral/IP Beverly 4 10 Couper, 2003
192 ± 67 35 IP 5 cysts DX 4 10 Dziadek, 2012
63.5 ± 6.758 11 IP 15 cysts Kenya chx 2 4 Mokua, 2017
115.8 ± 12.53 11 IP 15 cysts Kenya chx 3 4 Mokua, 2017
192.8 ± 47.23 25 IP 15 cysts Kenya chx 4 4 Mokua, 2017
345 ± 56.42 16 IP 15 cysts Kenya chx 5 4 Mokua, 2017
416.3 ± 32.2 8 IP 15 cysts Kenya chx 6 4 Mokua, 2017
216 ± 8 4 IP 20 cysts ME49 4 6 Lee, 2016
250 ± 107 43 Oral 20 cysts ME49 6 N.D. Resende, 2008
237 ± 106 45 Oral 4 cysts ME49 6 N.D. Resende, 2008
1198 ± 153 13 Oral 1000 tachys ME49 4 9 Chew, 2012
885 ± 70 8 Oral 1000 tachys ME49 2 10 Chew, 2011
1489 ± 35 2 Oral 1000 tachys ME49 4 10 Chew, 2011
1587 ± 46 3 Oral 1000 tachys ME49 8 10 Chew, 2011
1621 ± 63 4 Oral 1000 tachys ME49 16 10 Chew, 2011
640 ± 72 11 IP 10^3 tachys PruΔ Ku80 N.D. 4 Abdelbaset, 2017
231 ± 116 50 IP 10^4 tachys PruΔ Ku80 N.D. 6 Abdelbaset, 2017
169 ± 27 16 IP 10^6 tachy PruΔ Ku80 5 4 Abdelbaset, 2017
260 ± 124 48 N.D. 5×10^2 tachys PruΔ Ku80 4 9 Mouveaux, 2014
721 ± 257 36 Oral 10 cysts PRU 4 6 Wang, 2016
1340 ± 225 17 Oral 20 cysts PRU 4 12 Lu, 2017
1860 ± 60 3 Oral 20 cysts PRU 4 6 Wang, 2017
1747 ± 133 8 Oral 20 cysts PRU 4 6 Wang, 2017
1924 ± 82 4 Oral 20 cysts PRU 4 6 Wang, 2017
2050 ± 57 3 Oral 20 cysts PRU 4 6 Wang, 2017
1955 ± 120 6 Oral 20 cysts PRU 4 6 Wang, 2017
1283 ± 193 15 Oral 20 cysts PRU 4 10 Han, 2017
2350 ±757 32 Oral 20 cysts PRU 6 6 Luo, 2017
1735 ± 495 29 Oral 20 cysts PRU 4 6 Yin, 2015
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Table 3.

Cyst Variability in Swiss Webster Mice.

Swiss Webster
Cyst Count (+−SD) Coefficient of Variation (Percentage) Infection Method Parasite Strain Brain Isolation in Weeks Post Infection Sample Size Reference
560 ± 57 10 IP 10 cyst ME49 4 10 El-Sayad, 2016
9914 ± 6209 63 IP 20 cysts ME49 4 7 Martins-Duarte, 2010
13611 ± 21260 156 IP 20 cysts ME49 4 7 Martins-Duarte, 2010
4966 ± 4095 82 IP 20 cysts ME49 4 11 Martins-Duarte, 2010
14090 ± 19767 140 IP 20 cysts ME49 4 9 Martins-Duarte, 2010
2000 ± 500 25 IP 20 cysts ME49 14 5 Araujo, 1992
750 ± 100 13 IP 20 cysts ME49 16 5 Araujo, 1992
750 ± 100 13 IP 20 cysts ME49 26 5 Araujo, 1992
450 ± 100 22 IP 20 cysts ME49 28 5 Araujo, 1992
14.4 ± 2.9 20 Oral 10 cysts ME49 8 20 Eissa, 2015
12.48 ± 5.72 46 Oral 10 cysts ME49 8 10 El-Zawawy, 2015
11.37 ± 6.24 55 Oral 10 cysts ME49 8 10 El-Zawawy, 2015
1289 ± 1579 123 Oral 10 cysts ME49 4 15 Djurković-Djaković, 2002
1315 ± 1315 100 Oral 20 cysts ME49 7 15 Djurković-Djaković, 2002
1782 ± 1695 95 Oral 20 cysts ME49 7 15 Djurković-Djaković, 2002
95 ±33 35 IP 500 tachys PruΔ Ku80 3 4 Singh, 2002
1083 ± 609 56 Oral N.D. PRU 8 4 Aldebert, 2011
1143 ± 553 48 Oral N.D. PRU 8 5 Aldebert, 2011
912 ± 371 41 IP 30 cysts Weiss 6 30 Witting, 1979
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Table 4.

Cyst Variability in the CBA Strain.

CBA
Cyst Count (+−SD) Coefficient of Variation (Percentage) Infection Method Parasite Strain Brain Isolation in Weeks Post Infection Sample Size Reference
10250 ± 3140 31 Oral 70 cysts 76K 4 10 Bonenfant, 2001
2309 ± 825 36 Oral 100 cysts 76K 4 40 Velge-Roussel, 1997
1451 ± 16 1 IP 10 cysts ME49 5 3 Araujo, 1992
1373 ± 26 2 IP 10 cysts ME49 6 3 Araujo, 1992
1342 ± 37 3 IP 10 cysts ME49 7 3 Araujo, 1992
1353 ± 68 5 IP 10 cysts ME49 9 3 Araujo, 1992
1451 ± 41 3 IP 10 cysts ME49 11 3 Araujo, 1992
1306 ± 42 3 IP 10 cysts ME49 13 3 Araujo, 1992
1368 ± 42 3 IP 10 cysts ME49 15 3 Araujo, 1992
1150 ± 52 5 IP 10 cysts ME49 17 3 Araujo, 1992
1400 ± 737 53 IP 18 cysts ME49 7 7 Schultz, 2014
1673 ± 854 51 IP 18 cysts ME49 7 6 Schultz, 2014
1251 ± 419 34 IP 18 cysts ME49 9 10 Schultz, 2014
2523 ± 894 35 IP 18 cysts ME49 7 19 Doggett, 2012
3888 ± 3268 84 IP 18–20 cysts ME49 3 29 Watts, 2015
3072 ± 2170 71 IP 18–20 cysts ME49 4 29 Watts, 2015
3342 ± 1499 45 IP 18–20 cysts ME49 5 12 Watts, 2015
3086 ± 1829 59 IP 18–20 cysts ME49 6 15 Watts, 2015
2540 ± 1567 62 IP 18–20 cysts ME49 8 14 Watts, 2015
112 ± 73 65 IP 20 cysts ME49 6 6 Brinkmann, 1987
29.5 ± 10 34 IP 20 cysts ME49 6 7 Brinkmann, 1987
14 ± 13 93 IP 10^4 tachys Mic1–3KO 6 3 Moiré, 2009
5 ± 7 140 IP 10^3 tachys Mic1–3KO 6 4 Moiré, 2009
2 ± 1 50 IP 10^2 tachys Mic1–3KO 6 8 Moiré, 2009
369 ± 224 61 IP 200 tachys PruΔ Ku80 4 15 Jones, 2017
164 ± 63 38 IP 2.5×10^4 tachys PruΔ Ku80 4 3 Buchholz, 2013
435 ± 230 53 IP 2.5×10^4 tachys PruΔ Ku80 8 6 Buchholz, 2013
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Table 5.

Cyst Variability in C57BL Mice.

C57BL/6
Cyst Count (+−SD) Coefficient of Variation (Percentage) Infection Method Parasite Strain Brain Isolation in Weeks Post Infection Sample Size Reference
2448 ± 573 23 Oral 10 cysts 76K 4 20 Velge-Roussel, 1997
13433 ± 3306 25 Oral 85 cysts Beverly 5 N.D. El-Malky, 2005
13295 ± 3237 24 Oral 85 cysts Beverly 5 14 El-Malky, 2014
2273 ± 1361 60 IP 5 cysts DX 4 7 Dziadek, 2011
1142 ± 320 28 Oral 10 cysts Fukaya 6 3 Makino, 2011
1100 ± 260 24 Oral 10 cysts Fukaya 6 3 Makino, 2011
391 ± 188 48 IP 40 cysts ME49 2 7 Kim, 2017
3193 ± 412 13 Oral 10 cysts ME49 3 6 Bhadra, 2013
5609 ± 693 12 Oral 10 cysts ME49 5 6 Bhadra, 2013
1083 ± 402 37 Oral 10 cysts ME49 7 6 Bhadra, 2013
3471± 1559 45 Oral 10 cysts ME49 10 6 Bhadra, 2013
1002 ± 111 11 Oral 40 cysts ME49 4 N.D. Pinzan, 2015
500 ± 100 20 IP 20,000 tachys PLK 6 2 Zhang, 2007
210 ± 91 43 IP 10,000 tachys PruΔ Ku80 7 4 Sugi, 2016
248 ± 63 25 IP 200 tachys PruΔ Ku80 5 5 Walker, 2013
142 ± 92 65 IP 200 tachys PruΔ Ku80 4 18 Jones, 2017
139 ± 91 66 IP 200 tachys PruΔ Ku80 4 18 Jones, 2017
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Table 6.

C3H Cyst Variability.

C3H
Cyst Count (+−SD) Coefficient of Variation (Percentage) Infection Method Parasite Strain Brain Isolation in Weeks Post Infection Sample Size Reference
2055 ± 723 35 IP 5 cysts DX 4 7 Dziadek, 2011
5793 ± 347 6 IP 40 cysts IPB-G 6 4 Scorza, 2003
25 ± 8 32 IP 20 cysts ME49 6 5 Brinkmann, 1987
4550 ± 1400 31 Oral 20 cysts ME49 4 5 Clemente, 2005
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Coefficient of Variation utilized the mean and standard deviation seen from cyst variability to determine the ratio of variability, as a percent. Kunming mice were consistently infected with the T. gondii PRU strain via oral cysts, with typically low variation. Most variables were kept constant; isolation of brain at four weeks post infection and a three to seven sample size.

The coefficient of variation compared the ratio of the mean and standard deviation, from the cyst variability, and calculated a percentage. A majority of papers infected Swiss Webster mice with ME49 cysts and isolated brains at various weeks post infection.

Studies were ordered based upon parasite strain followed by method of infection. Cyst variability (mean ± standard deviation) determined the coefficient of variation, shown as a percent. A majority of papers administered ME49 cysts intraperitoneally, with lower variability occurring in smaller inoculation doses.

ME49 is the most common T. gondii strain inoculated in C57BL when testing for cyst burden (mean ± standard deviation). The ratio of standard deviation to the mean is represented in the coefficient of variation values, as a percentage. A few sources did not mention to the sample size, denoted as N.D.

A small portion of papers reviewed tested cyst burden in C3H mice utilizing three strains of T. gondii. Cyst variability is listed in terms of mean ± standard deviation, from which the coefficient of variation ratio is determined, as a percentage. Most of the variation values are similar except when C3H mice were infected with IPB-G, generating a large mean with relatively low standard deviation.

Cyst burden in the Balb/c model is ordered by parasite strain followed by method of inoculation, identifying the number and route of infection. The Coefficient of Variation is determined by the ratio of cyst variability (mean ± average) and is represented as a percent. Balb/c displayed the lowest cyst variance particularly when infected with ME49 tachyzoites orally and intraperitoneally, along with oral PRU cysts in one study.

4.2. Mouse Strain Variability & Cyst Burden

The lowest cyst variation identified occurred in CBA mice infected IP with 10 ME49 cysts, producing a CV less than 2 (Figure 1). However, when the dose was increased a larger range of variation, 33% - 84%, was observed. In contrast, Kunming mice produced overall the lowest cyst variability compared to other mouse strains studied, generating an average CV value of 8% (Table 7), while also producing the highest cyst burdens. Notably, the Kunming strain is not utilized or available in most countries. Balb/c mice produced the next overall lowest cyst variability compared to other mouse strains studied, generating a CV value of 20% (Figure 2). This value decreases to 15% when mice are infected with the T. gondii strain ME49, specially orally with 1,000 tachyzoites (6%) and 20 cysts IP (4%). Swiss Webster mice, an outbred strain, produce the highest variability in CV value at 60%. Mouse strains not commonly utilized are detailed in Table 8.

Figure 1. CBA Cyst Variability.

Figure 1.

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As an illustration, cyst burden is displayed in CBA mice and the coefficient of variation was determined for each parasite strain and mode of inoculation tested. Sample size is denoted as the thickness of the circles. Mean coefficient of variation values are denoted beside each variable, with larger values expressing a greater variance of standard deviation from the mean. CV values for each sample set were averaged to form representative means for each variable (e.g. parasite strain, mode of inoculation). In the CBA strain high variance was seen in all conditions even when a large sample size infected ME49 cysts intraperitoneal.

Table 7.

Cyst Variability Summary

Average Median Range Maximum Minimum
Balb/c
Cyst Count 913 681 2287 2350 64
CV 20 14 62 65 2
C3H
Cyst Count 3106 3303 5768 5793 25
CV 26 31 29 35 6
C57BL/6
Cyst Count 2922 1100 13294 13433 139
CV 34 25 55 66 11
CBA
Cyst Count 1750 1368 10248 10250 2
CV 41 38 139 140 1
Kunming
Cyst Count 4460 3255 16147 18080 1933
CV 8 6 20 21 1
Swiss Webster
Cyst Count 2881 1083 14079 14090 11
CV 60 48 146 156 10
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Cyst count and CV numbers were analyzed for their average, median, range, maximum, and minimum values.

Figure 2. Balb/c Cyst Variability.

Figure 2.

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As an illustration, cyst burden is displayed in Balb/c mice and the coefficient of variation was determined for each parasite strain and mode of inoculation tested. Sample size is denoted as the thickness of the circles. Mean coefficient of variation values are denoted beside each variable, with larger values expressing a greater variance of standard deviation from the mean. CV values for each sample set were averaged to form representative means for each variable (e.g. parasite strain, mode of inoculation). In the Balb/c strain high variance was seen in all conditions except when mice were infected with ME49 cysts intraperitoneal or orally with ME49 tachyzoites.

Table 8.

Various mouse stain cyst variability

Mouse Strain Cyst Count (+−SD) Coefficient of Variation (Percentage) Infection Method Parasite Strain Brain Isolation in Weeks Post Infection Sample Size Reference
Balb/k 3120 ± 766 25 oral 20 cysts Beverly 8 N.D. Roberts (1992)
CD-1 111 ± 72 65 IP 500 tachys GT1 20 11 Xiao (2016)
CD-1 611 ± 402 66 oral 10 cysts HIF 18 7 Berenreiterová (2011)
CD-1 883 ± 938 106 oral 10 cysts HIF 18 7 Berenreiterová (2011)
F1 cross 100 ± 66 66 oral 1 cyst 01529/38 3 6 Hrdá (2000)
F1 cross 124 ± 42 34 oral 1 cyst 01529/38 6 10 Hrdá (2000)
F1 cross 97 ± 39 40 oral 1 cyst 01529/38 12 7 Hrdá (2000)
Gal3+/+ 840 ± 145 17 oral 20 cysts ME49 2 5 Bemardes (2006)
Gal3−/− 4408 ± 1266 29 oral 20 cysts ME49 2 5 Bemardes (2006)
IL-6 +/+ 2080 ± 960 46 oral 20 cyst Beverly 4 4 Jebbari (1998)
IL-6 −/− 10079 ± 6079 60 oral 20 cyst Beverly 4 4 Jebbari (1998)
NIH 3181 ± 89 3 oral 20 cysts ME49 8 21 Martínez-Gómez (2009)
TR 122 ± 50 45 oral 50 cysts ME49 12 6 Dias (2014)
TR 258 ± 55 21 oral 100 cysts ME49 12 6 Dias (2014)
TS 249 ± 59 24 oral 50 cysts ME49 12 6 Dias (2014)
TS 449 ± 72 16 oral 100 cysts ME49 12 6 Dias (2014)
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Several mouse strains were not commonly mentioned throughout literature and are seen in this table. The coefficient of variation took the mean and standard deviation, seen under cyst variability, to produce a ratio indicating percent variability in cyst burden.

4.3. Method of Infection Variability

The method of infection does not appear to be a primary determinant in cyst variability. Infecting orally with tachyzoites, a method typically limited to Balb/c, produced the lowest variance in this review. Inbred mouse strains generally experience reduced varaibility when infected with cysts, either orally or intraperitoneally. The opposite can be seen in the outbred Swiss Webster: tachyzoites injected intraperitoneally exhibited lower variance then cysts administered orally or intraperitoneally.

5. Conclusion

Based on the collected data, our results indicate excessive variation seen in the Swiss Webster mouse strain (as expected with an outbred strain) in contrast to reduced relative variability in the Kunming, CBA, and Balb/c in-bred strains. The coefficient of variation (a ratio of standard deviation to the mean) was lowest in the Kunming strain, although this strain is used almost exclusively in Asia currently. CBA exhibited low variance when infected with 10 cysts of the ME49 strain, but as the dose increased, so also did the CV value. The more widely used mouse strain Balb/c produced low variation when ME49 cysts were infected intraperitoneally (IP). Along with mouse strain variance, parasite strain was also analyzed and ME49 produced the lowest fluctuation in half the mouse strains. This information will aid researchers to construct more optimized protocols with the T. gondii-murine model of the latent infection; which will allow improved investigation of interventions to reduce cyst burden or study behavioral changes.

Highlights.

  • Toxoplasma gondii chronic infection displays varied cyst numbers between mouse strains.

  • The parasite strain, ME49, produces the lowest cyst variation in 3 of the 6 mouse strains evaluated.

  • No method of infection was identified to substantially reduce the diverse cyst load amount in the brain.

  • The Kunming mouse strain established the most consistent cyst burden but is rarely used outside Asia.

Acknowledgements

Funding Statement: We recognize the U.S. National Institutes of Health [P20GM103427], Nebraska Research Initiative, and the GRACA and FUSE programs of the University of Nebraska at Omaha for support of this work. None of these agencies participated in study design, data collection, analysis, or the decision for publication.

Footnotes

Conflicts of interest: none

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References

  1. Abdelbaset AE, Fox BA, Karram MH, Abd Ellah MR, Bzik DJ, Igarashi M. Lactate dehydrogenase in Toxoplasma gondii controls virulence, bradyzoite differentiation, and chronic infection. PLoS One. 2017. March 21;12(3):e0173745. doi: 10.1371/journal.pone.0173745. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Affymetrix Inc. (n.d.). Mouse Haplotype Table. Retrieved from http://tools.thermofisher.com/content/sfs/brochures/Mouse_Haplotype_Table.pdf
  3. Sabin Albert B. Toxoplasmic Encephalitis in Children. JAMA. 1941;116(9):801–807. doi: 10.1001/jama.1941.02820090001001 [DOI] [Google Scholar]
  4. Aldebert D, Hypolite M, Cavailles P, Touquet B, Flori P, Loeuillet C, Cesbron-Delauw MF. Development of high-throughput methods to quantify cysts of Toxoplasma gondii. Cytometry A. 2011. November;79(11):952–8. doi: 10.1002/cyto.a.21138. [DOI] [PubMed] [Google Scholar]
  5. Araujo FG, Huskinson-Mark J, Gutteridge WE, Remington JS. In vitro and in vivo activities of the hydroxynaphthoquinone 566C80 against the cyst form of Toxoplasma gondii. Antimicrob Agents Chemother. 1992. February;36(2):326–30. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Berenreiterová M, Flegr J, Kuběna AA, Němec P. The distribution of Toxoplasma gondii cysts in the brain of a mouse with latent toxoplasmosis: implications for the behavioral manipulation hypothesis. PLoS One. 2011;6(12):e28925. doi: 10.1371/journal.pone.0028925. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Bernardes ES, Silva NM, Ruas LP, Mineo JR, Loyola AM, Hsu DK, Liu FT, Chammas R, Roque-Barreira MC. Toxoplasma gondii infection reveals a novel regulatory role for galectin-3 in the interface of innate and adaptive immunity. Am J Pathol. 2006. June;168(6):1910–20. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Bhadra R, Cobb DA, Khan IA. Donor CD8+ T cells prevent Toxoplasma gondii de-encystation but fail to rescue the exhausted endogenous CD8+ T cell population. Infect Immun. 2013. September;81(9):3414–25. doi: 10.1128/IAI.00784-12. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Bonenfant C, Dimier-Poisson I, Velge-Roussel F, Buzoni-Gatel D, Del Giudice G, Rappuoli R, Bout D. Intranasal immunization with SAG1 and nontoxic mutant heat-labile enterotoxins protects mice against Toxoplasma gondii. Infect Immun. 2001. March;69(3):1605–12. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Boyle JP, Saeij JP, Boothroyd JC. Toxoplasma gondii: inconsistent dissemination patterns following oral infection in mice. Exp Parasitol. 2007. July;116(3):302–5. [DOI] [PubMed] [Google Scholar]
  11. Brinkmann V, Remington JS, Sharma SD. Protective immunity in toxoplasmosis: correlation between antibody response, brain cyst formation, T-cell activation, and survival in normal and B-cell-deficient mice bearing the H-2k haplotype. Infect Immun. 1987. April;55(4):990–4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Buchholz KR, Bowyer PW, Boothroyd JC. Bradyzoite pseudokinase 1 is crucial for efficient oral infectivity of the Toxoplasma gondii tissue cyst. Eukaryot Cell. 2013. March;12(3):399–410. doi: 10.1128/EC.00343-12. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Chen J, Li ZY, Petersen E, Huang SY, Zhou DH, Zhu XQ. DNA vaccination with genes encoding Toxoplasma gondii antigens ROP5 and GRA15 induces protective immunity against toxoplasmosis in Kunming mice. Expert Rev Vaccines. 2015. April;14(4):617–24. doi: 10.1586/14760584.2015.1011133. [DOI] [PubMed] [Google Scholar]
  14. Chew WK, Segarra I, Ambu S, Mak JW. Significant reduction of brain cysts caused by Toxoplasma gondii after treatment with spiramycin coadministered with metronidazole in a mouse model of chronic toxoplasmosis. Antimicrob Agents Chemother. 2012. April;56(4):1762–8. doi: 10.1128/AAC.05183-11. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Chew WK, Wah MJ, Ambu S, Segarra I. Toxoplasma gondii: determination of the onset of chronic infection in mice and the in vitro reactivation of brain cysts. Exp Parasitol. 2011. January;130(1):22–5. doi: 10.1016/j.exppara.2011.10.004. [DOI] [PubMed] [Google Scholar]
  16. Clemente M, Curilovic R, Sassone A, Zelada A, Angel SO, Mentaberry AN. Production of the main surface antigen of Toxoplasma gondii in tobacco leaves and analysis of its antigenicity and immunogenicity. Mol Biotechnol. 2005. May;30(1):41–50. [DOI] [PubMed] [Google Scholar]
  17. Couper KN, Nielsen HV, Petersen E, Roberts F, Roberts CW, Alexander J. DNA vaccination with the immunodominant tachyzoite surface antigen (SAG-1) protects against adult acquired Toxoplasma gondii infection but does not prevent maternofoetal transmission. Vaccine. 2003. June 20;21(21–22):2813–20. [DOI] [PubMed] [Google Scholar]
  18. Di Cristina M, Marocco D, Galizi R, Proietti C, Spaccapelo R, Crisanti A. Temporal and spatial distribution of Toxoplasma gondii differentiation into Bradyzoites and tissue cyst formation in vivo. Infect Immun. 2008. August;76(8):3491–501. doi: 10.1128/IAI.00254-08. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Dias RR, Carvalho EC, Leite CC, Tedesco RC, Calabrese Kda S, Silva AC, DaMatta RA, de Fatima Sarro-Silva M. Toxoplasma gondii oral infection induces intestinal inflammation and retinochoroiditis in mice genetically selected for immune oral tolerance resistance. PLoS One. 2014. December 1;9(12):e113374. doi: 10.1371/journal.pone.0113374. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Djurković-Djaković O, Milenković V, Nikolić A, Bobić B, Grujić J. Efficacy of atovaquone combined with clindamycin against murine infection with a cystogenic (Me49) strain of Toxoplasma gondii. J Antimicrob Chemother. 2002. December;50(6):981–7. [DOI] [PubMed] [Google Scholar]
  21. Doggett JS, Nilsen A, Forquer I, Wegmann KW, Jones-Brando L, Yolken RH, Bordón C, Charman SA, Katneni K, Schultz T, Burrows JN, Hinrichs DJ, Meunier B, Carruthers VB, Riscoe MK. Endochin-like quinolones are highly efficacious against acute and latent experimental toxoplasmosis. Proc Natl Acad Sci U S A. 2012. September 25;109(39):15936–41. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Dubey JP, Lindsay DS, Speer CA. Structures of Toxoplasma gondii tachyzoites, bradyzoites, and sporozoites and biology and development of tissue cysts. Clin Microbiol Rev. 1998. April;11(2):267–99. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Dziadek B, Gatkowska J, Brzostek A, Dziadek J, Dzitko K, Grzybowski M, Dlugonska H. Evaluation of three recombinant multi-antigenic vaccines composed of surface and secretory antigens of Toxoplasma gondii in murine models of experimental toxoplasmosis. Vaccine. 2011. January 17;29(4):821–30. doi: 10.1016/j.vaccine.2010.11.002. [DOI] [PubMed] [Google Scholar]
  24. Dziadek B, Gatkowska J, Grzybowski M, Dziadek J, Dzitko K, Dlugonska H. Toxoplasma gondii: the vaccine potential of three trivalent antigen-cocktails composed of recombinant ROP2, ROP4, GRA4 and SAG1 proteins against chronic toxoplasmosis in BALB/c mice. Exp Parasitol. 2012. May;131(1):133–8. doi: 10.1016/j.exppara.2012.02.026. [DOI] [PubMed] [Google Scholar]
  25. Eissa MM, Barakat AM, Amer EI, Younis LK. Could miltefosine be used as a therapy for toxoplasmosis? Exp Parasitol. 2015. October;157:12–22. doi: 10.1016/j.exppara.2015.06.005. [DOI] [PubMed] [Google Scholar]
  26. EL-Malky MA, Al-Harthi SA, Mohamed RT, EL Bali MA, Saudy NS. Vaccination with Toxoplasma lysate antigen and CpG oligodeoxynucleotides: comparison of immune responses in intranasal versus intramuscular administrations. Parasitol Res. 2014. June;113(6):2277–84. doi: 10.1007/s00436-014-3882-0. [DOI] [PubMed] [Google Scholar]
  27. El-Malky M, Shaohong L, Kumagai T, Yabu Y, Noureldin MS, Saudy N, Maruyama H, Ohta N. Protective effect of vaccination with Toxoplasma lysate antigen and CpG as an adjuvant against Toxoplasma gondii in susceptible C57BL/6 mice. Microbiol Immunol. 2005;49(7):639–46. [DOI] [PubMed] [Google Scholar]
  28. El-Sayed NM, Ismail KA, Badawy AF, Elhasanein KF. In vivo effect of anti-TNF agent (etanercept) in reactivation of latent toxoplasmosis. J Parasit Dis. 2016. December;40(4):1459–1465. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. El-Zawawy LA, El-Said D, Mossallam SF, Ramadan HS, Younis SS. Preventive prospective of triclosan and triclosan-liposomal nanoparticles against experimental infection with a cystogenic ME49 strain of Toxoplasma gondii. Acta Trop. 2015. January;141(Pt A):103–11. doi: 10.1016/j.actatropica.2014.09.020. [DOI] [PubMed] [Google Scholar]
  30. Flegr J Effects of toxoplasma on human behavior. Schizophr Bull 2007. May;33(3):757–60. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Gatkowska J, Hiszczynska-Sawicka E, Kur J, Holec L, Dlugonska H. Toxoplasma gondii: an evaluation of diagnostic value of recombinant antigens in a murine model. Exp Parasitol. 2006. November;114(3):220–7. [DOI] [PubMed] [Google Scholar]
  32. Gatkowska J, Wieczorek M, Dziadek B, Dzitko K, Dlugonska H. Behavioral changes in mice caused by Toxoplasma gondii invasion of brain. Parasitol Res. 2012. July;111(1):53–8. doi: 10.1007/s00436-011-2800-y. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Gontijo da Silva M, Clare Vinaud M, de Castro AM. Prevalence of toxoplasmosis in pregnant women and vertical transmission of Toxoplasma gondii in patients from basic units of health from Gurupi, Tocantins, Brazil, from 2012 to 2014. PLoS One. 2015. November 11;10(11):e0141700. doi: 10.1371/journal.pone.0141700. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Han Y, Zhou A, Lu G, Zhao G, Wang L, Guo J, Song P, Zhou J, Zhou H, Cong H, He S. Protection via a ROM4 DNA vaccine and peptide against Toxoplasma gondii in BALB/c mice. BMC Infect Dis. 2017. January 11;17(1):59. doi: 10.1186/s12879-016-2104-z. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Hrdá S, Votýpka J, Kodym P, Flegr J. Transient nature of Toxoplasma gondii-induced behavioral changes in mice. J Parasitol. 2000. August;86(4):657–63. [DOI] [PubMed] [Google Scholar]
  36. Huang S, Holmes MJ, Radke JB, Hong DP, Liu TK, White MW, Sullivan WJ Jr. Toxoplasma gondii AP2IX-4 Regulates Gene Expression during Bradyzoite Development. mSphere. 2017. March 15;2(2). pii: e00054–17. doi: 10.1128/mSphere.00054-17. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Jebbari H, Roberts CW, Ferguson DJ, Bluethmann H, Alexander J. A protective role for IL-6 during early infection with Toxoplasma gondii. Parasite Immunol. 1998. May;20(5):231–9. [DOI] [PubMed] [Google Scholar]
  38. Johnson AM. Strain-dependent, route of challenge-dependent, murine susceptibility to toxoplasmosis. Z Parasitenkd. 1984;70(3):303–9. [DOI] [PubMed] [Google Scholar]
  39. Jones NG, Wang Q, Sibley LD. Secreted protein kinases regulate cyst burden during chronic toxoplasmosis. Cell Microbiol. 2017. February;19(2). doi: 10.1111/cmi.12651. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Kim JH, Lee J, Bae SJ, Kim Y, Park BJ, Choi JW, Kwon J, Cha GH, Yoo HJ, Jo EK, Bae YS, Lee YH, Yuk JM. NADPH oxidase 4 is required for the generation of macrophage migration inhibitory factor and host defense against Toxoplasma gondii infection. Sci Rep. 2017. July 25;7(1):6361. doi: 10.1038/s41598-017-06610-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Lee DH, Kim AR, Lee SH, Quan FS. Cross-protection induced by Toxoplasma gondii virus-like particle vaccine upon intraperitoneal route challenge. Acta Trop. 2016. December;164:77–83. doi: 10.1016/j.actatropica.2016.08.025. [DOI] [PubMed] [Google Scholar]
  42. Li XZ, Wang XH, Xia LJ, Weng YB, Hernandez JA, Tu LQ, Li LT, Li SJ, Yuan ZG. Protective efficacy of recombinant canine adenovirus type-2 expressing TgROP18 (CAV-2-ROP18) against acute and chronic Toxoplasma gondii infection in mice. BMC Infect Dis. 2015. March 4;15:114. doi: 10.1186/s12879-015-0815-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Lu G, Zhou J, Zhou A, Han Y, Guo J, Song P, Zhou H, Cong H, Hou M, Wang L, He S. SAG5B and SAG5C combined vaccine protects mice against Toxoplasma gondii infection. Parasitol Int. 2017. October;66(5):596–602. doi: 10.1016/j.parint.2017.06.002. [DOI] [PubMed] [Google Scholar]
  44. Luo F, Zheng L, Hu Y, Liu S, Wang Y, Xiong Z, Hu X, Tan F. Induction of Protective Immunity against Toxoplasma gondii in Mice by Nucleoside Triphosphate Hydrolase-II (NTPase-II) Self-amplifying RNA Vaccine Encapsulated in Lipid Nanoparticle (LNP). Front Microbiol. 2017. April 5;8:605. doi: 10.3389/fmicb.2017.00605. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Makino M, Uemura N, Moroda M, Kikumura A, Piao LX, Mohamed RM, Aosai F. Innate immunity in DNA vaccine with Toxoplasma gondii-heat shock protein 70 gene that induces DC activation and Th1 polarization. Vaccine. 2011. February 24;29(10):1899–905. doi: 10.1016/j.vaccine.2010.12.118. [DOI] [PubMed] [Google Scholar]
  46. Martínez-Gómez F, García-González LF, Mondragón-Flores R, Bautista-Garfias CR. Protection against Toxoplasma gondii brain cyst formation in mice immunized with Toxoplasma gondii cytoskeleton proteins and Lactobacillus casei as adjuvant. Vet Parasitol. 2009. March 23;160(3–4):311–5. doi: 10.1016/j.vetpar.2008.11.017. [DOI] [PubMed] [Google Scholar]
  47. Martins-Duarte ES, Lemgruber L, de Souza W, Vommaro RC. Toxoplasma gondii: fluconazole and itraconazole activity against toxoplasmosis in a murine model. Exp Parasitol. 2010. April;124(4):466–9. doi: 10.1016/j.exppara.2009.12.011. [DOI] [PubMed] [Google Scholar]
  48. Miller CM, Smith NC, Ikin RJ, Boulter NR, Dalton JP, Donnelly S. Immunological interactions between 2 common pathogens, Th1-inducing protozoan Toxoplasma gondii and the Th2-inducing helminth Fasciola hepatica. PLoS One. 2009. May 25;4(5):e5692. doi: 10.1371/journal.pone.0005692. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Moiré N, Dion S, Lebrun M, Dubremetz JF, Dimier-Poisson I. Mic1–3KO tachyzoite a live attenuated vaccine candidate against toxoplasmosis derived from a type I strain shows features of type II strain. Exp Parasitol. 2009. October;123(2):111–7. doi: 10.1016/j.exppara.2009.06.003. [DOI] [PubMed] [Google Scholar]
  50. Mokua Mose J, Muchina Kamau D, Kagira JM, Maina N, Ngotho M, Njuguna A, Karanja SM. Development of Neurological Mouse Model for Toxoplasmosis Using Toxoplasma gondii Isolated from Chicken in Kenya. Patholog Res Int. 2017;2017:4302459. doi: 10.1155/2017/4302459. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Montoya JG, Liesenfeld O. (2004) Toxoplasmosis. The Lancet 262:1965–1976. [DOI] [PubMed] [Google Scholar]
  52. Mouveaux T, Oria G, Werkmeister E, Slomianny C, Fox BA, Bzik DJ, Tomavo S. Nuclear glycolytic enzyme enolase of Toxoplasma gondii functions as a transcriptional regulator. PLoS One. 2014. August 25;9(8):e105820. doi: 10.1371/journal.pone.0105820. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Pinzan CF, Sardinha-Silva A, Almeida F, Lai L, Lopes CD, Lourenço EV, Panunto-Castelo A, Matthews S, Roque-Barreira MC. Vaccination with Recombinant Microneme Proteins Confers Protection against Experimental Toxoplasmosis in Mice. PLoS One. 2015. November 17;10(11):e0143087. doi: 10.1371/journal.pone.0143087. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Randall LM, Hunter CA. Parasite dissemination and the pathogenesis of toxoplasmosis. Eur J Microbiol Immunol (Bp). 2011. March;1(1):3–9. doi: 10.1556/EuJMI.1.2011.1.3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Resende MG, Fux B, Caetano BC, Mendes EA, Silva NM, Ferreira AM, Melo MN, Vitor RW, Gazzinelli RT. The role of MHC haplotypes H2d/H2b in mouse resistance/susceptibility to cyst formation is influenced by the lineage of infective Toxoplasma gondii strain. An Acad Bras Cienc. 2008. March;80(1):85–99. [DOI] [PubMed] [Google Scholar]
  56. Roberts CW, Alexander J. Studies on a murine model of congenital toxoplasmosis: vertical disease transmission only occurs in BALB/c mice infected for the first time during pregnancy. Parasitology. 1992. February;104 Pt 1:19–23. [DOI] [PubMed] [Google Scholar]
  57. Saeij JP, Boyle JP, Boothroyd JC. Differences among the three major strains of Toxoplasma gondii and their specific interactions with the infected host. Trends Parasitol. 2005. October;21(10):476–81. [DOI] [PubMed] [Google Scholar]
  58. Schultz TL, Hencken CP, Woodard LE, Posner GH, Yolken RH, Jones-Brando L, Carruthers VB. A thiazole derivative of artemisinin moderately reduces Toxoplasma gondii cyst burden in infected mice. J Parasitol. 2014. August;100(4):516–21. doi: 10.1645/13-451.1. [DOI] [PubMed] [Google Scholar]
  59. Scorza T, D’Souza S, Laloup M, Dewit J, De Braekeleer J, Verschueren H, Vercammen M, Huygen K, Jongert E. A GRA1 DNA vaccine primes cytolytic CD8(+) T cells to control acute Toxoplasma gondii infection. Infect Immun. 2003. January;71(1):309–16. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Singh U, Brewer JL, Boothroyd JC. Genetic analysis of tachyzoite to bradyzoite differentiation mutants in Toxoplasma gondii reveals a hierarchy of gene induction. Mol Microbiol. 2002. May;44(3):721–33. [DOI] [PubMed] [Google Scholar]
  61. Su C, Evans D, Cole RH, Kissinger JC, Ajioka JW, Sibley LD. Recent expansion of Toxoplasma through enhanced oral transmission. Science. 2003. January 17;299(5605):414–6. [DOI] [PubMed] [Google Scholar]
  62. Su C, Khan A, Zhou P, Majumdar D, Ajzenberg D, Dardé ML, Zhu XQ, Ajioka JW, Rosenthal BM, Dubey JP, Sibley LD. Globally diverse Toxoplasma gondii isolates comprise six major clades originating from a small number of distinct ancestral lineages. Proc Natl Acad Sci U S A. 2012. April 10;109(15):5844–9. doi: 10.1073/pnas.1203190109. [DOI] [PMC free article] [PubMed] [Google Scholar]
  63. Subauste C Animal models for Toxoplasma gondii infection. Curr Protoc Immunol. 2012. February;Chapter 19:Unit 19.3.1–23. doi: 10.1002/0471142735.im1903s96. [DOI] [PubMed] [Google Scholar]
  64. Sugi T, Ma YF, Tomita T, Murakoshi F, Eaton MS, Yakubu R, Han B, Tu V, Kato K, Kawazu S, Gupta N, Suvorova ES, White MW, Kim K, Weiss LM. Toxoplasma gondii Cyclic AMP-Dependent Protein Kinase Subunit 3 Is Involved in the Switch from Tachyzoite to Bradyzoite Development. MBio. 2016. May 31;7(3). pii: e00755–16. doi: 10.1128/mBio.00755-16. [DOI] [PMC free article] [PubMed] [Google Scholar]
  65. Suzuki Y, Orellana MA, Wong SY, Conley FK, Remington JS. Susceptibility to chronic infection with Toxoplasma gondii does not correlate with susceptibility to acute infection in mice. Infect Immun. 1993. June;61(6):2284–8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  66. Velge-Roussel F, Moretto M, Buzoni-Gatel D, Dimier-Poisson I, Ferrer M, Hoebeke J, Bout D. Differences in immunological response to a T. gondii protein (SAG1) derived peptide between two strains of mice: effect on protection in T. gondii infection. Mol Immunol. 1997. October;34(15):1045–53. [DOI] [PubMed] [Google Scholar]
  67. Vyas A, Kim SK, Giacomini N, Boothroyd JC, Sapolsky RM. Behavioral changes induced by Toxoplasma infection of rodents are highly specific to aversion of cat odors. Proc Natl Acad Sci U S A. 2007. April 10;104(15):6442–7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  68. Walker R, Gissot M, Croken MM, Huot L, Hot D, Kim K, Tomavo S. The Toxoplasma nuclear factor TgAP2XI-4 controls bradyzoite gene expression and cyst formation. Mol Microbiol. 2013. February;87(3):641–55. doi: 10.1111/mmi.12121. [DOI] [PMC free article] [PubMed] [Google Scholar]
  69. Wang JL, Elsheikha HM, Zhu WN, Chen K, Li TT, Yue DM, Zhang XX, Huang SY, Zhu XQ . Immunization with Toxoplasma gondii GRA17 Deletion Mutant Induces Partial Protection and Survival in Challenged Mice. Front Immunol 2017. June 29;8:730. doi: 10.3389/fimmu.2017.00730. [DOI] [PMC free article] [PubMed] [Google Scholar]
  70. Wang M, Yang XY, Zhang NZ, Zhang DL, Zhu XQ. Evaluation of Protective Immune Responses Induced by Recombinant TrxLp and ENO2 Proteins against Toxoplasma gondii Infection in BALB/c Mice. Biomed Res Int. 2016;2016:3571962. [DOI] [PMC free article] [PubMed] [Google Scholar]
  71. Wang T, Yin H, Li Y, Zhao L, Sun X, Cong H. Vaccination with recombinant adenovirus expressing multi-stage antigens of Toxoplasma gondii by the mucosal route induces higher systemic cellular and local mucosal immune responses than with other vaccination routes. Parasite. 2017;24:12. doi: 10.1051/parasite/2017013. [DOI] [PMC free article] [PubMed] [Google Scholar]
  72. Watts E, Zhao Y, Dhara A, Eller B, Patwardhan A, Sinai AP. Novel Approaches Reveal that Toxoplasma gondii Bradyzoites within Tissue Cysts Are Dynamic and Replicating Entities In Vivo. MBio. 2015. September 8;6(5):e01155–15. doi: 10.1128/mBio.01155-15. [DOI] [PMC free article] [PubMed] [Google Scholar]
  73. Witting PA. Learning capacity and memory of normal and Toxoplasma-infected laboratory rats and mice. Z Parasitenkd. 1979;61(1):29–51. [DOI] [PubMed] [Google Scholar]
  74. Xiao J, Li Y, Prandovszky E, Kannan G, Viscidi RP, Pletnikov MV, Yolken RH. Behavioral Abnormalities in a Mouse Model of Chronic Toxoplasmosis Are Associated with MAG1 Antibody Levels and Cyst Burden. PLoS Negl Trop Dis. 2016. April 28;10(4):e0004674. doi: 10.1371/journal.pntd.0004674. [DOI] [PMC free article] [PubMed] [Google Scholar]
  75. Xiao J, Yolken RH. Strain hypothesis of Toxoplasma gondii infection on the outcome of human diseases. Acta Physiol (Oxf). 2015. April;213(4):828–45. doi: 10.1111/apha.12458. [DOI] [PMC free article] [PubMed] [Google Scholar]
  76. Xu Y, Zhang NZ, Tan QD, Chen J, Lu J, Xu QM, Zhu XQ. Evaluation of immuno-efficacy of a novel DNA vaccine encoding Toxoplasma gondii rhoptry protein 38 (TgROP38) against chronic toxoplasmosis in a murine model. BMC Infect Dis. 2014. September 30;14:525. doi: 10.1186/1471-2334-14-525. [DOI] [PMC free article] [PubMed] [Google Scholar]
  77. Yang WB, Zhou DH, Zou Y, Chen K, Liu Q, Wang JL, Zhu XQ, Zhao GH. Vaccination with a DNA vaccine encoding Toxoplasma gondii ROP54 induces protective immunity against toxoplasmosis in mice. Acta Trop. 2017. December;176:427–432. doi: 10.1016/j.actatropica.2017.09.007. [DOI] [PubMed] [Google Scholar]
  78. Yin H, Zhao L, Wang T, Zhou H, He S, Cong H. A Toxoplasma gondii vaccine encoding multistage antigens in conjunction with ubiquitin confers protective immunity to BALB/c mice against parasite infection. Parasit Vectors. 2015. September 30;8:498. doi: 10.1186/s13071-015-1108-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  79. Zenner L, Darcy F, Capron A, Cesbron-Delauw MF. Toxoplasma gondii: kinetics of the dissemination in the host tissues during the acute phase of infection of mice and rats. Exp Parasitol. 1998. September;90(1):86–94. [DOI] [PubMed] [Google Scholar]
  80. Zhang G, Huong VT, Battur B, Zhou J, Zhang H, Liao M, Kawase O, Lee EG, Dautu G, Igarashi M, Nishikawa Y, Xuan X. A heterologous prime-boost vaccination regime using DNA and a vaccinia virus, both expressing GRA4, induced protective immunity against Toxoplasma gondii infection in mice. Parasitology. 2007. September;134(Pt 10):1339–46. [DOI] [PubMed] [Google Scholar]
  81. Zhang NZ, Huang SY, Zhou DH, Chen J, Xu Y, Tian WP, Lu J, Zhu XQ. Protective immunity against Toxoplasma gondii induced by DNA immunization with the gene encoding a novel vaccine candidate: calcium-dependent protein kinase 3. BMC Infect Dis. 2013. October 31;13:512. doi: 10.1186/1471-2334-13-512. [DOI] [PMC free article] [PubMed] [Google Scholar]
  82. Zhang NZ, Xu Y, Wang M, Petersen E, Chen J, Huang SY, Zhu XQ. Protective efficacy of two novel DNA vaccines expressing Toxoplasma gondii rhomboid 4 and rhomboid 5 proteins against acute and chronic toxoplasmosis in mice. Expert Rev Vaccines. 2015;14(9):1289–97. doi: 10.1586/14760584.2015.1061938. [DOI] [PubMed] [Google Scholar]

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