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Home , Coccidia, Toxoplasma gondii

Rev. Biol. Trop., 45(1): 877-882,1997

Increasing of (, ) by Trypanosoma lewisi (Kinetoplastida, Trypanosomatidae) in white

OIga M. Guerrero, Misael Chinchilla and E. Abrahams Centro de Investigación en Enfermedades Tropicales (CIET). Departamento de Parasitología, Facultad de Microbiología, Universidad de Costa Rica, San José Costa Rica.

(Recibido 22-IV-1996. Corregido 9-VIII- 1996. Aceptado 23-IX-1996 .)

Abstra«;t: To demonstrate Iha! T. lewisi increases T. gondii multiplication in white rats, groups of five Wistar oc Sprague Dawley rats were inoculated with 106 T. lewisi trypomastigoles and four or seven days later infected wi!h Toxoplasma tachyzoites. survival time was monitored, and !he presence of T. gondii was confirmed in all dead rals by studying peritoneal exudate smears and lung tissue sections stained with haematoxylin-eosin. The presence of Toxoplasma cysts or antibodies was ehecked in the oí surviving rals. The ¡nerease is observed four days after trypanosome inoculatíon and is dependen! on fat strain, bul not on inoculum size oc age. Humoral and cellular factors may have a role in ¡he ¡nerease as has been reported for other experimental infections withAfrican trypanosomes and T. cruzi.

Key words: Toxoplasma gondii, Trypanosoma lewisi, irnmunosuppression, rats.

Since 1951-1955 it has been known that Immunosuppression of protozoan white rats are remarkably resistant to infections with other parasites has been Toxoplasma gondii infection (Ruchman and reported. For example Trypanosoma cruzi Fowler 1951, Lainson 1955). infection appears to be increased by some We have been tfying to explain the reasons immunosuppressive effects due to for such resistance. In our first studies we infections in host (Krettli 1977). demonstrated, with controlled inocula, that This effect has been also demonstrated in even 108 tachyzoites do not give rise to fatal African tripanosomíasis (Olsson et al. 1991. infection in adult white rats (Chinchilla et al. Darji et al. 1992). Particularly in 1981). We showed the importance of age , reactivation due to virus resistance (Chinchilla et al. 1981) and infection was reported in rruce (Pomeroy et al. macrophages activity (Chinchilla et al. 1982), 1989) and (Wiu et al. 1989). However, we for that natural resistance to T. gondii and have not found other studies of T. gondii studies in progress, including new information immunosuppression due to Trypanosoma about other factors that could play an infections. important role in this phenomenom, comfrrm We have found that Trypanosoma lewisi the low susceptibility of white rats to T. increases T. gondii infection of white , gondii. changing the susceptibility of this to 878 REVISTA DE BIOLOGIA TROPICAL the parasite. Studies of different factors RESULTS effecting this phenomena are presented in tbis papero Determination of T. lewisi previous infection time: In a controlled experiment, rats were infected with T. lewisi and after MATERIAL ANO METHODS different periods of time (O to 21 days), inoculated with T. gondii tachyzoites. Data : Wistar or Sprague Dawley rats indicate (Table 1) that T. lewisi inoculated to feed with a local chow and water ad libitum rats, four to six days previously to T. gondii were used in these experiments. Toxoplasma infection produces major increases of the tachyzoites for serology tests were obtained multiplication of this last parasite (see from NIH white mice peritoneal exudate. underlined numbers). In other experiment 30 or 60 days old rats Parasite strains: Tachyzoites of were inoculated with 104 trypomastigotes and Toxoplasma RH strain and partially four or seven days later infected with 106 T. characterized TCR-2 and TCR-3 strains (Holst gondii tachyzoites. As it is shown in Table 2, and Chinchilla 1990, Guerrero et al. 1991) both age animals infected with T. lewisi four were inoculated in different concentrations days before to Toxoplasma infection, died according to each experiment. These strains are earlier (see underlined data) than those infected mantained by regular passages (twice a week) seven days previously with trypanosomes. in the acute phase. Rats inoculated only with T. gondii (controls) T. lewisi strain was locally isolated from a survived 30 days, end of the experiments in Rattus norvegicus, gray rat, and mantained by anycase. weekly passage in white rats. Rat strain differences (Table 3): Either General experimental models: Groups 30 or 60 day old Wistar rats inoculated with of five Wistar or Sprague Dawley rats were Toxoplasma, previously infected with T. inoculated i.p. . with 106 T. lewisi lewisi, survived no more than 12 days. On the trypomastigotes obtained from one week other hand, Sprague Dawley rats survived more infected animals and four or seven days later, than 23 days. In both cases, animals inoculated according to the experiments, these rats were with T. gondii alone, survived 30 days, the end infected with different number of Toxoplasma of the experiment. tachyzoites. Simultaneously, other s were Toxoplasma strain differences: The inoculated only with T. gondii or T. lewisi, apparent immunosuppressive effect of T. representing the corresponding controls. AH lewisi was demonstrated for three strain studied infections were done intraperitoneally. (Table 4). However the survival time of the Survival time of rats was monitored, RH infected mice was lower. checking the presence of Toxoplasma in the peritoneal exudate or lungs in aH the dead rats. Effect of Toxoplasma inoculum: The Lung tissue sections were stained by average survival time was shorter in animals hematoxylin-eosin technique to demonstrate previously infected with T. lewisi, as compared lesions. Animal surviving for 30 days, the end with the controls, independent of the inocula póint for all the experiments, were studied for (lOs, 106, 107) of Toxoplaslna tachyzoites presence of brain. Toxoplasma cysts or for (Table 5). antibody presence. The Carbon Immuno-Assay All the Toxoplasma infected rats surviving (CIA) of known specificity (Chinchilla et al. 30 days were positive, either by brain cyst 1992) and used for us in other studies (Arias e t presence or the CIA test and many tachyzoites al. 1994) was performed for serologic analysis. were observed in peritoneal macrophages and A "t" student test for small samples was lung Iessions (Fig.1) of rats previously used for statistical analysis. inoculated with T. lewisi. Guerrero el aL: Increasing of Toxoplasma 879

TABLE 1.

Survival of rats* with T. gondii infection and pre-infected with T. lewisi.

Number of T. lewisi Number of T. gondi Time interval between T. Survival time (d) % Surviving trypomast./inoculated animal tachyzJinoculated animal lewisi and Toxoplasma 30 d infections (d) o O 30 100 10" O 30 100 O O 30 100 10" 3 30 100 lO" 4 20.8 60 10" S 7 O 10" 6 2SA 80 10" 10 30 100 106 14 30 100 10" 17 30 100 10" 21 30 100

*5 rats foreach group.

TABLE 2

Effect of strain or age of rats on T.1ewisi "immunosuppression" against T. gondii, inoculated four d later

Rat strain Age T. lewisi infection (d) Infected Non-infected Survival time % surviving Survival time % surviving (d) (30 d) (d) (30 d) Wistar 30 11.4 20 30 100 60 12.7 2S 30 100 Sprague Daw1ey 30 23 60 30 100 60 30 100 30 100

All the differences between '{oxoplasma irnmune response. Since T. gondii is an multiplication of T. lewisi infected and non­ intraeellular parasite in which cellular infected animalsshowed a P

Likewise, the natural reslstance �oí . fa.ts� \Vas �·ssion· düe l(feilher� suppression �of�'I'b-2··by�-· inhibited by the same treatment (Chinchilla et generations of prostaglandin-producing al. 1992). However, this resistance was evident maerophages, or a prostaglándin-independent after several days, which is different to what suppressive mechanism that inhibit the we report in this study. The effect exerted by expression of TL-2 receptors. Furtbermore, T. lewisi on Toxoplasma infection ocurred in they establish that TNF a and IFN-gamma only four to five days (Table 1 and 2) and it play an important role in the pathway of T-cell was not dependent on parasite strain or age (probably CD4+) irnmunosuppression. Araujo (Table 3 and 4). This means that we are in (1992) has demonstrated, in very detailed presence of a strong effect against the acquired experiments, the importance of CD4� cells 880 REVISTA DE BIOLOGIA TROPICAL

TABLE 3

Confirmation that previous T.lewisi infection gives rise to increasing of T.gondü multiplication in rats*

Age Time interval between T. T. lewisi infection (d) lewisi and Toxoplasma infections (d) Infected Non-infected Survival time % surviving Survivaltime % surviving (d) (30 d) (d) (30 d) 30 o 30 100 30 100 4 15 35.7 30 100 7 23.3 70 30 100 60 O 30 100 30 100 4 15.6 37.5 30 100 7 27.7 90 30 100

* 5 rats foe each group.

TABLE 4

Relation ofToxoplasma strains with the effectof T.lewisi parasite sinoculated four days earlier in Wistar rats.

T. gondii strain T. lewisi infection (4 d before) Infected Non-infected Survival time % surviving Survival time % surviving (d) (30 d) (d) (30 d) RH 16 40 30 100 TCR-2 20.4 60 30 100 TCR-3 24 75 30 100

TABLE 5

Effect ofToxoplasma inoculum on" immunosuppression" of Wistar rats, induced by T.lewisi, infected four days earlier

T gondii inocula T. lewisi infection (4 d before) (Tachyz.lanimal) Non-infected Infected Survival time % surviving Survival time %surviving (d) (30 d) (d) (30 d) 25 80 6.4 O 30 100 11.4 20 7 80 25.3 10

and IFN-a in the irnmune response against T. Albright and Albright (1991) have gondii. Sorne of these studies and others with suggested that in infeetions with T. brucei (Bakhiet et al. 1990) as well as with Trypanosoma musculi, a mouse parasite T. congolense (FIyn and Sileghen 1991) biologieally similar to T. lewisi, there are explain sOme irnmunosuppressive meehanisms several faetors bloeking of antibody aetivity,

that ean be suspected for our Toxoplasma - T. interferenee that affeet humoral immunity lewisi model. Sztein and Kierszenbaum (1992) suppression. This eould be another explanation suggest that alterations of the expression of for the apparent immunosuppressive effeet that TeR, CD4, CDg and IL-2R due to T. cruzi we have observed in our model. infeetions, eould be responsable of lymphoeyte This phenomenon appears to be related interferenee, giving rise to with the animal genetie origen as is shown in immunosuppression. 'This is another Table 3. Aetually the effeet was more evident interesting aspectto take into aceount for· our in Wistar than in Sprague Dawley rats. experiments. Differenees in animal immune response to intracellular parasites depending on animal, Guerrero et al.: íncreasing of Toxoplasma 881 specially rodent strains, have been reporte Chinchilla, M. M. Alfaro & O.M. Guerrero 198!. Adaptación natural de la rata blanca a Toxoplasma (Handman 1979). Specifically, for rat strains, gondii. Rev. Biol. Trop. 29: 273-282. in a study to detennine Pneumocystis carinii karyotypes obtained by corticosteroid Chinchilla, M. O.M. Guerrero & E. Solano 1982. Lack of multiplication of Toxoplasma in macrophages of rats treatment, Hong et al. (1992) needed only five in vitro.J. Parasitol. 68: 952-955. to eight weeks for immunosuppression of Chinchilla, M. O.M. Guerrero, L. Reyes & A. Castro. Wistar rats. On the contrary a nine to ten 1992. Efecto de los corticosteroides en la trasmisión weeks treatment was neccesary to obtain the congénita de toxoplasmosis experimental. Rev. Biol. same effeet for Sprague Dawley rats. Trop. 40: 135-137.

In conclusion, these preliminary studies Chinchilla, M. L. Reyes. O.M. Guerrero & F. Hernández. demonstrate that T. lewisi infections can 1992. Specificity of !he carbon immunoassay (CIA) Toxoplasma ¡nerease Toxoplasma multiplication in rats due test for !he diagnosis of ¡nfeclion. Veto Parasitol.44: 315-320. to an apparent immunosuppressive effect. The presence of many tachyzoites in lung lesions Darji, A. R. Lucas, S. Magez, E. Torreele, 1. Palacios, M. Sileghem, E. Bajyana Songa, R. Hamers & P. De (Fig. 1 a, b, e) as well as in peritoneal Baetselier. 1992. Mechanisms underlying macrophages (Fig. Id) oí these animals, trypanosorne-elicited irnmunosuppression. Ann. Soco compared with the controIs, support the Belg. Med. Trop. 72 (Supp!. 1): 27-38. interpretation that T. gondii was the cause of Flyn J.N. & M. Sileghern. 1991. l1te role of the death. macrophage in induction of immunosuppression in Furthermore survival of mts infected with Trypanosoma congolense-infected caldeo Immullo­ logy 74: 310-316. T. lewisi or T. gondií alone let us think that the effect is not produced by other disease, as Frenkel, J.K. 1985. lmmunity in toxoplasmosis. PAHO has been mentioned for bartonelosis infections Bulletin 19: 354-36. (Perla and Marmorston94 1 1) which can Guerrero, O.M. M. Chinchilla, R. Marin, G. Catarinella modified the natural resistance. & A. Castro. 1991. Estudio comparativo de dos cepas de Toxoplasma gondii. Parasitol.Día. 15: 97-10L

Handman, E. R. Ceredig& G.F. Milchells. 1979. Murine ACKNOWLEDGMENTS cutaneous leishmaniasis: Disease parterns in intact and nude rnice of various genotypes and examination of sorne differences between nonna! and infected This work was supported by grants # 430- macrophages. AJEBAK 57: 9-29.

92-904 and # 430-92-905 from !he University Holst, I. & M. Chinchilla. i 990. Development and of Costa Rica. We thank J.K. Frenkel for distribution of cysts of an avirulent strain of suggestions about this manuscript. Toxoplasmagondií and ¡he humoral immune response in rnice.Rev. Biol. Trop. 38: 189-193.

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