Camargo I.J.

Corsini A.C.

Costa M.G.

Oliveira O.L.P.

Stelini Jr. A.

English

Repositorio da Producao Cientifica e Intelectual da Unicamp

Rev. Inst. Med. trcp. São Paulo
22(4) :192-196, julho-agosto, 1980
UDC 593 .16t.t3
616.937.3-092.9
SUSCEPTIBITITY OF INBRED MICE TO TRYPANOSOMA CRUZI STRA¡N Y
Antonio C. CORSINI, Mârcos G. COSTÀ, Odete L. P. OLIVEIRA, Irineu J. B. CAMARG0 and
Amaldo STELINI Jr. (1)
SUMMARY
Inbred mice infected i.p. with 100 parasites of T. cruzi Strain Y varied in their
resistance to infection from highfy susceptible CrIJe/J to resistant (CBA x Crt
B,L/L}) Fr. CBA v¿as found susceptible and Csz BV10 moderatly resistant. F, mice
are recommended as a good model for experimental Chagas'disease since an acute
and chronic phase are well characterized in these mice.
INTRODUCTION
Chagas' disease (CHAGAS, 1909) 3 is a se-
rious health problem in many Latin American
countdes (rtt¡HO, 10). Different clinical manifes'
tations of the disease have been described, for
instance, heart involvement is very common in
Brazil but unusual in Argentina and Chile
'where digestive üract manifestation are moÍe
common (REZENDEs). Brazilian Chagaq' di-
sease comprises heart, aesophagus and colon
"megas", isolated or combined in the sâme pa-
tient (I(OBERLE s).
Those geographical differences have been
thought to be due to different strains of Try-
panosoma wrazi and/or differences in the ge-
netic background of south-American populations
as well as influences due to the general state
of health of patients since sometimes multiple
parasitic diseases overlap in the same subject.
Since previous work of. PIZZI et al.7 inbred
mice have been shown to differ in their re-
sistance to T. cruzi. That resistance is not
linked to the major histocompatibility locus
since rnice sharing the same H, locus manifest-
ed different susceptibility (TRISCHMAN et
al. g).
A continuity of.resistance from highly sus-
ceptible CaH.e/J to resistant B.alb/C and
C,TB\/LO, possibly related to a single or multi-
ple genetic component, were demonstrated by
(1) Depa¡tment of Micrcbiology and Immunology, University
Corespondence to A, C. corsini at the above address
:1Ð2
TRISCHMAN et al. e using Brazil strain of T'
crrnzi.
On the other hand variations in susceptibi-
lity related to different strâins of T. cruzi were
described in CrrBVlO since this mouse strain
was found more susceptible to Tulahen than to
Brazil stfain. Moreover, higher inoculum led
to higher parasitaemiaS and increased mortali-
ty (TRISCHMAN et al.s).
The F.razilian strain Y isolated from a human
case of Chagas Disease- (PEREIRA DA SILVA
& NUSSENZTT,yEIG ó) has been largely used in
experimental Chagas'disease and has been
shown highly virulent to outbred Swiss mice
(BRENER2). Usually a large inoculum (104-10s)
has been currently used to infect mice i.p. On
the other hand as little as 100 parasites are
enough to infect A,/Sn mice that are highly
susceptible to strain Y (Mota-personal commu-
nication).
In order to verify the course of infection in
CaH, CBA, C'TFJ¿/L0 and (CBA x CrtBVlO) F1
mice were infected i.p. with 100 parasites and
parasitaemia scored daily.
The results showed that mice differed in
their susceptibility to strain Y and only (CBA x
CstBULl) F, were found resistant over a large
of Carnpinas 
- 
UNICAMP, 13.100 
- 
Ca,rnpinas '-- Brazil
CORSINI, A. C.; COSTA, M. G.; OLMIR{, O. L. P.;
inbred mice to Trypanosoma cruzi strain Y. Rev.
CAMARGO, L J. B. & STELINI JR., A. 
- 
Susceptibility of
Inst. Med. trop. São Paulo 22:192-196, 1980.
dose of parasites used to infect mice. Those F
mice are recommended as a good model to study
experimental Chagas'disease since an acute and
a, chronic phase are well characterized.
MATERIAL AND METÍIODS
1. Animals 
- 
CrHe/J, CB'A, C,TBL/10 and
(CBAxCrrBUl0)F, female and male mice bred
and kept in clean conditions in our animals fa'
cilities were used throughout. Mice were 3
months old when infected.
2. Trypanosome 
- 
Trypanosoma (Schizotri-
panum) cruzi strain Y (PEREIRA DA SILVA &
NUSSENZWEIG6) obtained from Dr. Zigman
Brener (Belo Horizonte, Brazil) in 1970, weekly
passaged in outbred Swiss 55 mice were used.
Swiss mice lvere infected i.p. with 10s parasites
and bled 7 days later at the peak of parasitae-
mia (BRENER2).
3. Infeotion of mice 
- 
Infected blood harv-
ested in heparin (10U,hl) from the axillary ple-
xus was diluted in Eagle MEM.1%' FCS. Mice
were infected i.p. with 100 parasites in 0.2 ml of
Eagle MEM serum free. Counts were made in
a Neubauer haemocitometer.
4. ParasÍúaernía 
- 
Parasites in peripheral
blood were scored daily in all mice using wet
btood films as described by BRENER 2. Mice
were cut at ther'tail tip and 5 pl of blood har-
r¡ested in a haemoglobin pipette after the first
drop had been discharged. The sample was dis-
persed under a coverslip Q2 x 22 mm) over sli-
des. Parasites were counted in 100 microscopic
fields (40 x magnification) and the results ex-
pressed per mm3 of blood.
Mice were marked in the footpad so para-
sitaemia could be followed-up daily in every sin-
gle mouse. Experiments were performed 4 times
per each strain using at least 10 animals,/group.
RESULTS 
:
1. Parasitaemia in CrHe/J and CBA IuIICE 
-CrEf mice were found highly susceptible to the
infection. Although this strain showed the long-
est pre-patent period (? days) they developed
an increasing parasitaemia from the 9th day up
to 5000 parasites,/mms at the llth day after infec-
tion.
Mice usually controlled. their parasitaemia
by day 12 but again parasitaemia increased from
the 13ttr day onwards killing 93.3% of. animals
around day 15 (Fig. 1, Table I).
=
=
ø
u
=q
À
PARASIIAEMIA
DAYS OF INFEClION
Fig. 1 
- 
Title: Pa¡asitaemia of CaII, CBA and C67B1/10 mice infected with T. cruzi.
Legend: Mice were infected. i.p. with 100 trypomastigotes. A-A-¡Crn; f,----f,CBA; *-*(-a6?81/10.
193
coRsINI, A. C.; COSTA, M. G.; OLMIRA, O. L. p.; c.aMARCiO, I. J. B. & STELTNI JR., A. 
- 
Susceptibility of
inbred mice to Trypanosom cruzi strain Y. Rev. Inst. Med. troB. São PâuIo 22:192-196, 1980.
TABLEI
Mortality of inbred mouse strains infected with T. crul¡
Mortality
20 days 60 days
crH
CBA
c57BL/70
(CBA x Cã?BLI10) Fl
Iì/25b (920/o) 25/25 (l00vo)
20/33 (66.60/o) 33/33 (1000/o)
70/67 (l6.4oto) 20/67 (32.8v0)
4/709 (3.?oto) 4/7Q9 (3.70/o)
a 
- 
Mice were infected i.p. with 100 trypomastigotes
b 
- 
Number dead per number infected
CrH showed a regular pattern of infection
with a small SD of the mean at the peak,
although larger SD lvere found on other days.
As low as 10 parasites were able to infected and
kill CaH mice. The pattetn of the infection was
similar to that of 100 parasites but a higher va-
riation rr¡as veÌified since some mice peaked
on day 12 or 13 after infection (data not sho¡wn).
CBA mice ïvere highly susceptible as rvell to
strain Y. Mice had a prepatent period of 5 days
and developed from then a high parasitae-
mia. Usually mice controlled their parasitaemia
around day l3 but a second wave killd as many
a,s 9O7o of mice around day 15. Seldom mice
survived infection longer than 20 days (Fig. 1,
Table I). Although CrH and CBA are highly sus-
ceptible to the infection, there are some differ-
ences between them.
Thus, CBA mice had a lo'wer parasitaemia
(2000/mm¡ parasites at the peak) and although
the pattern was similar in several experiments
i.e. two successive waves of parasites in periphe-
ral blood, the peak of the first wave varied from
day 9 to 11. Conversely CrH had the same behe-
viour in several experiments. No differences
rrere noticed between female ot male C"H and
CBA mice.
2. Parasitaemia in C'TBL/I0 
- 
C'.B.L,/LO
mice were found moderatly resistant to the in-
fection sinoe 30% of animals died. Parasitaemia
developed after a pre-patent period of 6 days
reaching a peak of 300 parasites,hm3 on day
11. From this point onwards animals cleared
their peripheral blood and kept very low num-
bers of circulating parasites. Mortality (25-30o/o)
was verified from the day 2O and beared no re-
lation with the parasitaemia (Fig. 1, Table I).
Survivors lyere kept as long as 6 months
194
apparently with no signs of disease. No differ-
ences between sexes were noticed.
3. Parasitaemia in (CBA x CrrBLll0) F, 
-Female and male (CBA x CszB.WL0) F, mice
IMere found resistant to the infection with 100
parasites of the strain Y. Mice had a pre-patent
period of 6 days then developed an increasing
parasitaemia up to the 1lth day. From the peak
onwards animals usualþ cleared parasites from
the peripheral blood. This pattern was found
quite regular with no changes in several expe-
riments (Fig. 2, Table II).
Higher inocula (103, 104 and 10s parasites)
showed for 103 parasites a shorter prepatent
period (4 days) and a peak reached sooner (9trt
day) but still with the same pattern and 1000/o
survival (Fig.2, Table II).
Conversely, 10+ parasites had also a shorter
pre-patent period and a higher peak but killed
as many as 500/o of animals on day 20. Survi-
vors'were apparently healthy (Fig. 2, Table II).
On the other hand 1S parasites killed all
rnice around day 7 after infection. Ðeath was
not related to parasitaemia, since parasites in-
crea,sed only up to 600,/mm¡ 04 day 7.
DISCUSSION
Our results showed that mice differed in
their susceptibility to T. cruzi strain Y. CrH and
CBA mice \¡sere found highly susceptible
although with different patterns in their para^si-
taemia. Coincidently these strains peaked all
on day 11 although with different pre-patent pe-
riods and parasites numbers at the peak. CrH
developed the highest parasitaemia (5000 para-
siteVmmg). CBA with a lo'ïver number of para.
sites at the peak developed usually a second
wave \¡vhich killed the animals.
There was not any direct relationship
between parasites numbers in peripheral blood
and death as rvas shown by TRISCHMAN et
al. s using Brazil strain.
Although CBA mice did not show always
the same peaking day as far as parasitaemia
w¿ìs concerned, the pattern was the same in
different experiments. Those differences may
be related to Y intra-strain variations (AIIAUJO
& NEVES 1) unrevealed in another inbred mou-
se strain. Ttrese strain Y variations are possible
to occur since we did not use cloned parasites.
coRsrNr, A. c.; cosTA, M. G.; OLMIRA, O. L. P.; CAMARCiO, L J. B. & STET,INI JR., A. 
- 
Susceptibility of
inbred mice to lly¡ranosoma crui strain Y. Rev, Insú. Med. trop. São Pâulo 22:192-196' 1980'
TABLEII
Moftality of (CBA x C5?BLI10) Fr mice, infected with
T' crr¡a
Numþer of infecting
Trypomastigotes
Mortality
20 days 60 days
On the other hand C'TBI./LO has been des'
cribed as resistant to T. cruzi strain Y (COS'
TA4) C'7E.IJ10 \¡r¡as described resistant as well
to Brazil strain with a large inoculum (104)
(TRISCHMAN et al.e). Our results shoïYed a li'
mited resistance since 25-30%'of animals died
around day 20 of infection with very low pa'
rasitaemia.
Converseþ, (CBA x cszBl,/l0) F, mice were
found highly resistant since they developed a
low parasitaemia, cleared the parasites fro(n
peripheral þlood and kept a low or undetected
On the other hand higher inoculum of 10r
parasites killed 500/o of animals by 20 day of
infection. As many as 10s parasites killed all
mice by da,y 7 of infection.
Our results confÍrmed the existence of a
continoum of resistance in inbred strains of
mice towards infection with strain Y as was
described for Brazil strain (TRISCHMAN et
al. s) in ? weeks old mice infected with 10¿ pa-
rasites.
l0¡
104
106
1/15b (6.60/o)
10/20 60vo)
10/10 (100ouo)
l/15 G'.60/o)
L6/20 (80v0)
a 
- 
Mice 'were infected i.p.
b 
- 
Nurnber dead per number infected
/
o
.
E
Ø
u
ts
Ø
c
PARASITAEMIA
o 5 lo 15 20
DAYS OF INFECTION
Fig,. 2 
- 
Titte: Pârasita€mia of (cBA x CurBl/lo) F1 mice inJected with 1. crozi.
Legend: Mice were infected i.p. {'ith A-À-¡tO'; *-*108 o" *- - - - *104 trvpoma'stigotes.
,f.
parasitaemia for a long time been apparently
healthy. \üe have mice infected for 10 months.
, .4, large inoculum of 103 parasites showed a
briefer prepatent period and an earlier peak but
mice still controlled the infection (Fig. 2). This
behaviour of (CBA x CrrBUL0) F, towards in-
fection with strain Y makes them a suitable
model for experimental Chagas'disease since an
acute and chronic phase are \ryell characterized.
Experirnents are been carried out to study the
behaviour of Fr mioe towards reinfection.
195
coRsrNr, A. c.; cosTA, M. G.; OLTVEÌRA, O. L. p.; c.aMARco,
inbred mice to ltyl,anosoma cruzl strain y. Bev. Insú, Med.
I. J. B. & STELINI JR., A. 
- 
Susceptibility of
úrop. São Paulo 22:192-196, 1980.
Differences were found in relation to CrrBL/
10, since this strain was found only moderately
resistant.
Some differences related to sex were notic-
ed in (CBA x CrrBUtÛ) F, mice since female
mice usually showed a lower parasitaemia than
male mice.
On the other hand we could not confirm
the results of COSTA ¿ since C.,BV10 was nor
found as resistant as described. -óOSfAn describ-
ed survival of these mice for months with ino-
çulum of 500 parasites. Mice were susceptible
'only to 5.000 and 50.000 parasites.
', fntra-strain variations may explain these
differences and points to the necessity to clonef. cruzÍ strains.
;' fn conclusion as low as 100 parasites of T.
ôruzi strain Y are enough to infect mice and ro
separate inbred strains of mice in susceptible
or resistant. We feel that a small inoculum is
closer to the natural infection than the larger
ones.
RESUMO
Suscetibilidade de camundongos isogênicos à
infecção pela cepa Y do Trypanosoma cruzi
Linhagens de camundongos isogênicos in-
fectados intraperitonealmente com 100 parasitas
da cepa Y do T. cruzi diferem quanto à resis-
tência à infecção.
A linhagem CrIJe/J é altamente suscetível à
infecção contrariamente à (CBA x C."BV10) F,
que.é resistente.
A linhagem CBA mostrou-se suscetível e a
C,TBL/|0 moderadamente resistente.
Os camundongos (CBA x CrrBVt0) F, são
recomendados como um bom modelo para estu-
dos experimentais da doença de Chagas uma vez
que nestes camundongos a fase aguda e a fase
crônica estão bem caracterizadas.
ACKNOWLEDGEMENTS
This work was supported in part by grants
from CNPq and FAPESP. lVe thank Mrs. Isma-
lia Menegon Doné for ther assistance in breed-
ing the mice.
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- 
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- 
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- 
SWG (1) 7?.3, 197?.
t0
-.'q
Recebido pala publicagão em 18/10/1979.
196


Susceptibility Of Inbred Mice To Trypanosoma Cruzi Strain Y

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Susceptibility Of Inbred Mice To Trypanosoma Cruzi Strain Y

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