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Plasmodium falciparum full life cycle and Plasmodium ovale liver stages in humanized mice.

Soulard V, Bosson-Vanga H, Lorthiois A, Roucher C, Franetich JF, Zanghi G, Bordessoulles M, Tefit M, Thellier M, Morosan S, Le Naour G, Capron F, Suemizu H, Snounou G, Moreno-Sabater A, Mazier D - Nat Commun (2015)

Bottom Line: Experimental studies of Plasmodium parasites that infect humans are restricted by their host specificity.Thus, we obtain the complete hepatic development of P. falciparum, the transition to the erythrocytic stages, their subsequent multiplication, and the appearance of mature gametocytes over an extended period of observation.Thus, TK-NOG mice are highly suited for in vivo observations on the Plasmodium species of humans.

View Article: PubMed Central - PubMed

Affiliation: 1] Sorbonne Universités, UPMC Univ Paris 06, CR7, Centre d'Immunologie et des Maladies Infectieuses (CIMI-Paris), 91 Bd de l'hôpital, F-75013 Paris, France [2] INSERM, U1135, CIMI-PARIS, 91 Bd de l'hôpital, F-75013 Paris, France [3] CNRS, ERL 8255, CIMI-PARIS, 91 Bd de l'hôpital, F-75013 Paris, France.

ABSTRACT
Experimental studies of Plasmodium parasites that infect humans are restricted by their host specificity. Humanized mice offer a means to overcome this and further provide the opportunity to observe the parasites in vivo. Here we improve on previous protocols to achieve efficient double engraftment of TK-NOG mice by human primary hepatocytes and red blood cells. Thus, we obtain the complete hepatic development of P. falciparum, the transition to the erythrocytic stages, their subsequent multiplication, and the appearance of mature gametocytes over an extended period of observation. Furthermore, using sporozoites derived from two P. ovale-infected patients, we show that human hepatocytes engrafted in TK-NOG mice sustain maturation of the liver stages, and the presence of late-developing schizonts indicate the eventual activation of quiescent parasites. Thus, TK-NOG mice are highly suited for in vivo observations on the Plasmodium species of humans.

No MeSH data available.


Related in: MedlinePlus

Asexual and sexual blood stages of P. falciparum in sporozoite-infected TK-NOG mice engrafted with human hepatocytes and RBC.(a). Experimental procedure for the double engraftment of TK-NOG mice with human hepatocytes and RBC before infection with P. falciparum sporozoites. (b). In vitro asexual blood stage culture: at day 8 of infection, 200 ul of venous blood was put in culture to follow emergence of parasitaemia. Results of one experiment with three mice are expressed as the number of parasites within 2 μl of blood for each mouse. Results are representative of three independent experiments performed with a total of eight mice. Representative pictures of a trophozoite and a schizont observed on Giemsa-stained thick blood smears at day 0 of in vitro culture (= day 8 of infection) and of trophozoites observed on Giemsa-stained thin blood smears at day 4 of culture. (c). In vivo asexual blood stage development: individual parasitaemias were determined using Giemsa-stained thin blood smears from a total of 4 mice from two independent experiments. Representative pictures of schizonts observed on Giemsa-stained thick blood smears during the course of infection, scale bar, 5 μm. (d). In vivo sexual blood stage development: representative pictures of gametocytes at different stages of their development as observed in Giemsa-stained thick blood smears from two mice.
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f2: Asexual and sexual blood stages of P. falciparum in sporozoite-infected TK-NOG mice engrafted with human hepatocytes and RBC.(a). Experimental procedure for the double engraftment of TK-NOG mice with human hepatocytes and RBC before infection with P. falciparum sporozoites. (b). In vitro asexual blood stage culture: at day 8 of infection, 200 ul of venous blood was put in culture to follow emergence of parasitaemia. Results of one experiment with three mice are expressed as the number of parasites within 2 μl of blood for each mouse. Results are representative of three independent experiments performed with a total of eight mice. Representative pictures of a trophozoite and a schizont observed on Giemsa-stained thick blood smears at day 0 of in vitro culture (= day 8 of infection) and of trophozoites observed on Giemsa-stained thin blood smears at day 4 of culture. (c). In vivo asexual blood stage development: individual parasitaemias were determined using Giemsa-stained thin blood smears from a total of 4 mice from two independent experiments. Representative pictures of schizonts observed on Giemsa-stained thick blood smears during the course of infection, scale bar, 5 μm. (d). In vivo sexual blood stage development: representative pictures of gametocytes at different stages of their development as observed in Giemsa-stained thick blood smears from two mice.

Mentions: Groups of TK-NOG mice were then first engrafted with hHEP and when plasma hAlb levels reached 5.0–9.8 mg ml−1 (liver humanization≥60%), they were injected with hRBC daily for 6 days, before inoculation with 1.75 × 106 to 3.5 × 106P. falciparum sporozoites. Daily injections of hRBC were continued until the end of the observations. In this manner, optimal levels of hRBC (≥80%) were present in the mice at the time (day 6) when the hepatic schizonts are expected to reach full maturity and release infective merozoites (Supplementary Table 1). At day-8 post-sporozoite inoculation a sample of 200 μl of peripheral mouse blood was collected and used to initiate in vitro cultures that were then followed over the next 8 days to assess for the presence of P. falciparum blood stage parasites (Fig. 2a). In three independent experiments, cultured blood from all eight mice showed parasites within the first 4 days of in vitro culture (Fig. 2b). The robustness of the protocol is further underlined by the fact that three distinct batches of hHEP and of sporozoites (though all from the same line, NF54) were used for these experiments.


Plasmodium falciparum full life cycle and Plasmodium ovale liver stages in humanized mice.

Soulard V, Bosson-Vanga H, Lorthiois A, Roucher C, Franetich JF, Zanghi G, Bordessoulles M, Tefit M, Thellier M, Morosan S, Le Naour G, Capron F, Suemizu H, Snounou G, Moreno-Sabater A, Mazier D - Nat Commun (2015)

Asexual and sexual blood stages of P. falciparum in sporozoite-infected TK-NOG mice engrafted with human hepatocytes and RBC.(a). Experimental procedure for the double engraftment of TK-NOG mice with human hepatocytes and RBC before infection with P. falciparum sporozoites. (b). In vitro asexual blood stage culture: at day 8 of infection, 200 ul of venous blood was put in culture to follow emergence of parasitaemia. Results of one experiment with three mice are expressed as the number of parasites within 2 μl of blood for each mouse. Results are representative of three independent experiments performed with a total of eight mice. Representative pictures of a trophozoite and a schizont observed on Giemsa-stained thick blood smears at day 0 of in vitro culture (= day 8 of infection) and of trophozoites observed on Giemsa-stained thin blood smears at day 4 of culture. (c). In vivo asexual blood stage development: individual parasitaemias were determined using Giemsa-stained thin blood smears from a total of 4 mice from two independent experiments. Representative pictures of schizonts observed on Giemsa-stained thick blood smears during the course of infection, scale bar, 5 μm. (d). In vivo sexual blood stage development: representative pictures of gametocytes at different stages of their development as observed in Giemsa-stained thick blood smears from two mice.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4525212&req=5

f2: Asexual and sexual blood stages of P. falciparum in sporozoite-infected TK-NOG mice engrafted with human hepatocytes and RBC.(a). Experimental procedure for the double engraftment of TK-NOG mice with human hepatocytes and RBC before infection with P. falciparum sporozoites. (b). In vitro asexual blood stage culture: at day 8 of infection, 200 ul of venous blood was put in culture to follow emergence of parasitaemia. Results of one experiment with three mice are expressed as the number of parasites within 2 μl of blood for each mouse. Results are representative of three independent experiments performed with a total of eight mice. Representative pictures of a trophozoite and a schizont observed on Giemsa-stained thick blood smears at day 0 of in vitro culture (= day 8 of infection) and of trophozoites observed on Giemsa-stained thin blood smears at day 4 of culture. (c). In vivo asexual blood stage development: individual parasitaemias were determined using Giemsa-stained thin blood smears from a total of 4 mice from two independent experiments. Representative pictures of schizonts observed on Giemsa-stained thick blood smears during the course of infection, scale bar, 5 μm. (d). In vivo sexual blood stage development: representative pictures of gametocytes at different stages of their development as observed in Giemsa-stained thick blood smears from two mice.
Mentions: Groups of TK-NOG mice were then first engrafted with hHEP and when plasma hAlb levels reached 5.0–9.8 mg ml−1 (liver humanization≥60%), they were injected with hRBC daily for 6 days, before inoculation with 1.75 × 106 to 3.5 × 106P. falciparum sporozoites. Daily injections of hRBC were continued until the end of the observations. In this manner, optimal levels of hRBC (≥80%) were present in the mice at the time (day 6) when the hepatic schizonts are expected to reach full maturity and release infective merozoites (Supplementary Table 1). At day-8 post-sporozoite inoculation a sample of 200 μl of peripheral mouse blood was collected and used to initiate in vitro cultures that were then followed over the next 8 days to assess for the presence of P. falciparum blood stage parasites (Fig. 2a). In three independent experiments, cultured blood from all eight mice showed parasites within the first 4 days of in vitro culture (Fig. 2b). The robustness of the protocol is further underlined by the fact that three distinct batches of hHEP and of sporozoites (though all from the same line, NF54) were used for these experiments.

Bottom Line: Experimental studies of Plasmodium parasites that infect humans are restricted by their host specificity.Thus, we obtain the complete hepatic development of P. falciparum, the transition to the erythrocytic stages, their subsequent multiplication, and the appearance of mature gametocytes over an extended period of observation.Thus, TK-NOG mice are highly suited for in vivo observations on the Plasmodium species of humans.

View Article: PubMed Central - PubMed

Affiliation: 1] Sorbonne Universités, UPMC Univ Paris 06, CR7, Centre d'Immunologie et des Maladies Infectieuses (CIMI-Paris), 91 Bd de l'hôpital, F-75013 Paris, France [2] INSERM, U1135, CIMI-PARIS, 91 Bd de l'hôpital, F-75013 Paris, France [3] CNRS, ERL 8255, CIMI-PARIS, 91 Bd de l'hôpital, F-75013 Paris, France.

ABSTRACT
Experimental studies of Plasmodium parasites that infect humans are restricted by their host specificity. Humanized mice offer a means to overcome this and further provide the opportunity to observe the parasites in vivo. Here we improve on previous protocols to achieve efficient double engraftment of TK-NOG mice by human primary hepatocytes and red blood cells. Thus, we obtain the complete hepatic development of P. falciparum, the transition to the erythrocytic stages, their subsequent multiplication, and the appearance of mature gametocytes over an extended period of observation. Furthermore, using sporozoites derived from two P. ovale-infected patients, we show that human hepatocytes engrafted in TK-NOG mice sustain maturation of the liver stages, and the presence of late-developing schizonts indicate the eventual activation of quiescent parasites. Thus, TK-NOG mice are highly suited for in vivo observations on the Plasmodium species of humans.

No MeSH data available.


Related in: MedlinePlus