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First isolation and genotyping of Toxoplasma gondii strains from domestic animals in Tunisia.

ABSTRACT: The isolation and molecular typing of Toxoplasma gondii strains provide an essential basis for a better understanding of the parasite's genetic diversity, determinants of its geographical distribution and associated risks to human health. In this study, we isolated and genetically characterized T. gondii strains from domestic animals in Southern and coastal area of Tunisia. Blood, hearts and/or brains were collected from 766 domestic animals (630 sheep and 136 free-range chickens). Strain isolation from these samples was performed using mouse bioassay and genotyping was carried out with a multiplex PCR technique using 15 microsatellite markers. Thirty viable strains of T. gondii were successfully isolated from tissues of sheep (19/142) and chickens (11/33). In addition, 3 strains could be successfully genotyped from animal tissues for which mouse bioassay was unsuccessful. A large predominance of type II strains (n = 29) was found in the sampled regions, followed by type III (n = 3) and, for the first time in Tunisia, a single isolate of Africa 4 lineage from a sheep. Analyses of population genetics showed the presence of a divergent population of type II lineage in Tunisia, supporting limited recent migrations of strains between Tunisia and other countries of the world.

SUBMITTER: Lachkhem A

PROVIDER: S-EPMC8085010 | biostudies-literature | 2021 Apr

REPOSITORIES: biostudies-literature

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First isolation and genotyping of Toxoplasma gondii strains from domestic animals in Tunisia.

Lachkhem Arwa A Galal Lokman L Lahmar Ibtissem I Passebosc Karine K Riahi Homayoun H Plault Nicolas N Dardé Marie-Laure ML Mercier Aurélien A Babba Hamouda H

Scientific reports 20210429 1

The isolation and molecular typing of Toxoplasma gondii strains provide an essential basis for a better understanding of the parasite's genetic diversity, determinants of its geographical distribution and associated risks to human health. In this study, we isolated and genetically characterized T. gondii strains from domestic animals in Southern and coastal area of Tunisia. Blood, hearts and/or brains were collected from 766 domestic animals (630 sheep and 136 free-range chickens). Strain isolat ...[more]

PMID: 33927299

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Project description:BACKGROUND: Recent studies of Toxoplasma gondii isolates from animals in different regions of China have shown a limited genetic diversity and type China 1 was the dominant genotype of T. gondii prevalent in Chinese animals. However, little has been known concerning the isolation and genotyping of T. gondii circulating in chickens, pigs and rodents in China. The aim of the study was to characterize samples of T. gondii isolates obtained from naturally infected cats, pigs and free-range chickens slaughtered for human consumption in China. METHODS: In the present study, brain tissues of 77 animals collected from different areas of China, including 24 free-range chickens (Gallus domesticus) , 13 voles (Rattus flavipectus), 23 pigs and 17 cats, were bioassayed in mice and viable T. gondii were isolated from the brains of eleven. These eleven T. gondii isolates were maintained in Kunming (KM) outbred mice and DNA isolated from tissues of infected mice was characterized using 11 PCR-restriction fragment length polymorphism (PCR-RFLP) markers: SAG1, SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, Apico, and CS3. Moreover, to determine mouse virulence of China 1 lineage of parasites, a TgCtgy5 genotype isolate was selected randomly and assessed in KM mice with different inoculation doses. RESULTS: Results of genotyping revealed that ten isolates were type China 1 (ToxoDB PCR-RFLP genotype #9), and TgCksz1 was a new genotype that was reported for the first time designated here as ToxoDB PCR-RFLP #225. No clonal types I, II and III lineages were found. DNA sequencing of four introns (EF1, HP2, UPRT1 and UPRT7) and two genes (GRA6 and GRA7) from representative isolates confirmed the results of PCR-RFLP genotyping. The TgCtgy5 isolate was highly virulent in KM mice; all infected mice died of acute toxoplasmosis, irrespective of the inoculation dose. The results indicate that mouse virulent isolates of T. gondii are predominantly circulating in cats in China. CONCLUSIONS: T. gondii isolated from chickens, pigs, cats and rodents in different locations in China were genotyped and the results reconfirmed the limited diversity of T. gondii in China and showed that type China 1 lineage was dominant in this country.

Project description:BackgroundBartonellosis is an emerging vector-borne disease caused by different intracellular bacteria of the genus Bartonella (Rhizobiales: Bartonellaceae) that is transmitted primarily by blood-sucking arthropods such as sandflies, ticks and fleas. In Tunisia, there are no data available identifying the vectors of Bartonella spp. In our research, we used molecular methods to detect and characterize Bartonella species circulating in fleas collected from domestic animals in several of the country's bioclimatic areas.ResultsA total of 2178 fleas were collected from 5 cats, 27 dogs, 34 sheep, and 41 goats at 22 sites located in Tunisia's five bioclimatic zones. The fleas were identified as: 1803 Ctenocephalides felis (83%) (Siphonaptera: Pulicidae), 266 C. canis (12%) and 109 Pulex irritans (5%) (Siphonaptera: Pulicidae). Using conventional PCR, we screened the fleas for the presence of Bartonella spp., targeting the citrate synthase gene (gltA). Bartonella DNA was detected in 14% (121/866) of the tested flea pools [estimated infection rate (EIR) per 2 specimens: 0.072, 95% confidence interval (CI): 0.060-0.086]. The Bartonella infection rate per pool was broken down as follows: 55% (65/118; EIR per 2 specimens: 0.329, 95% CI: 0.262-0.402) in C. canis; 23.5% (8/34; EIR per 2 specimens: 0.125, 95% CI: 0.055-0.233) in P. irritans and 6.7% (48/714; EIR per 2 specimens: 0.032, 95% CI: 0.025-0.045) in C. felis. Infection rates, which varied significantly by bioclimatic zone (P < 0.0001), were highest in the humid areas. By sequencing, targeting the gltA gene and the 16S-23S rRNA Intergenic Spacer Regions (ITS), we identified three Bartonella zoonotic species: B. elizabethae, B. henselae, B. clarridgeiae, as well as uncharacterized Bartonella genotypes.ConclusionsTo the best of our knowledge, this is the first time that fleas in Tunisia have been shown to carry zoonotic species of Bartonella. The dog flea, Ctenocephalides canis, should be considered the main potential vector of Bartonella. Our study not only provides new information about this vector, but also offers a public health update: medical practitioners and farmers in Tunisia should be apprised of the presence of Bartonella in fleas and implement preventive measures.

Project description:The present study aimed to isolate and perform molecular and phenotypic characterization of Toxoplasma gondii strains infecting Iberian pigs bred under semi-free conditions and destined for human consumption. Blood and heart tissue samples from 361 fattening pigs from 10 various herds selected in the main areas of Iberian pig production were collected at a slaughterhouse; the sera were tested for anti-T. gondii antibodies using a commercial indirect ELISA kit, and a mouse bioassay was carried out using heart muscle of seropositive individual representatives from each geographical location. Seventy-nine (21.9%) of the 361 animals tested positive for anti-T. gondii antibodies according to the serology test. Fifteen samples of myocardial tissue were subjected to bioassay and 5 isolates (TgPigSp1 to TgPigSp5) were obtained. The isolates were characterized by using 11 PCR-RFLP genetic markers; three isolates had a ToxoDB #3 genotype (3/5) and two isolates had a ToxoDB #2 genotype (2/5). The TgPigSp1 and TgPigSp4 isolates were selected for virulence in mice characterization as instances of each different RFLP-genotype found. The TgPigSp1 isolate (#2 genotype) was virulent in mice with notable cumulative mortality (87.5%) and morbidity rates (100%); the TgPigSp4 (#3) was nonvirulent and triggered mild clinical signs in 42.1% of seropositive mice. Infection dynamics and organ distribution of both isolates were analyzed; the data revealed significant differences, including substantially higher parasite load in the lung during the acute phase of infection, in mice infected with TgPigSp1 than in the case of TgPigSp4 (median parasite load 7.6 vs. 0 zoites/mg, respectively; p < 0.05). Furthermore, degrees of severity of detected histopathological lesions appeared to be related to higher parasite burdens. Taking into account the unexpectedly high mortality rate and parasite load associated with the clonal genotype III, which is traditionally considered nonvirulent in mice, the need for further investigation and characterization of the T. gondii strains circulating in any host in Europe is emphasized.

Dataset Information

First isolation and genotyping of Toxoplasma gondii strains from domestic animals in Tunisia.

Publications

First isolation and genotyping of Toxoplasma gondii strains from domestic animals in Tunisia.

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