Case Report: Trypanosoma lewisi or T. lewisi -like Infection in a 37-Day-Old Indian Infant
Am. J. Trop. Med. Hyg., 85(2), 2011, pp. 221–224 doi:10.4269/ajtmh.2011.11-0002
221
Am. J. Trop. Med. Hyg., 85(2), 2011, pp. 221–224
doi:10.4269/ajtmh.2011.11-0002
Copyright © 2011 by The American Society of Tropical Medicine and Hygiene
INTRODUCTION
Trypanosomes are flagellated protozoan parasites infecting
a wide range of animals and man. Human infection with
Trypanosoma brucei ( T.b.) gambiense or T.b. rhodesiense
causes African sleeping sickness (African trypanosomiasis)
and Trypanosoma cruzi causes Chagas disease (American trypanosomiasis).
These infections have not been reported from
the Indian subcontinent.
Other species of trypanosomes are known to affect animals
in different parts of the world but human infection with
them is rare. Trypanosoma lewisi is an infection of rats and
there have been seven previous reports of human infection
with the organism. Here, we report a case of a T. lewisi infection
in a 37-day-old child from Bagpat, Uttar Pradesh in India.
This infant is arguably the youngest case to be reported. The
symptoms and treatment of the infant are discussed and the
literature is reviewed. We also discuss the morphological identification
of the parasite and its confirmation by molecular
analysis.
CASE REPORT
A 37-day-old infant, resident of Bagpat, Uttar Pradesh,
India, with no history of travel outside Uttar Pradesh was
admitted at St. Stephens Hospital, Delhi, India, in August
2010, with pyrexia (going up to 39°C), poor feeding (anorexia),
and lethargy for 1 day. On the day before admission, the child
got up from his sleep in the afternoon, screaming. His mother
went into the room and noticed three red spots on the leg.
The area became indurated and inflamed, about 2 cm around
the marks. The induration subsided in a couple of hours. The
sting marks remained to form a scab ( Figure 1 ). At that time
his mother attributed the crying and sting marks to wasps. The
child was cuddled and was consolable.
The next day he developed fever, seemed listless, and went
off feeds. He then had several episodes of generalized tonic
seizures for which he was brought to the hospital. Initially
lorazepam at 0.1 mg/kg/dose, and later phenobarbitone
(20 mg/kg/dose over 20 minutes, and repeated at 10 mg/kg
over 10 minutes) followed by phenytoin (20 mg/kg/dose over
20 minutes) were administered in succession before the seizures
were controlled.
On examination, the infant had no hepatosplenomegaly or
lymphadenopathy. Investigations on admission were essentially
normal ( Table 1 ). The infant had hemoglobin 10.6 gm/dL, white
blood cell counts 7,400/mm 3 , and platelet counts 102,000/mm 3 .
Blood smear showed trypomastigote forms of Trypanosoma
with the distinct subterminal kinetoplast, nucleus located
toward the anterior half, and a flagellum arising from near the
kinetoplast forming an undulating membrane before emerging
free from the anterior end ( Figure 2 ). Wet smear preparation
showed many motile trypanosomes. Biochemical tests
were unremarkable. Cerebrospinal fluid (CSF) was partially
traumatic and showed normal cell counts and biochemistry but
trypanosomes were seen in the specimen . The CSF examination
was repeated after 24 hours. This sample was not contaminated
by blood and did not show the parasite.
Specimens of blood on filter paper and blood thin smears
were sent to two laboratories for confirmation of trypanosomiasis
infection (Division of Parasitology, Indian Veterinary
Research Institute (IVRI), Izatnagar, U.P. and Institute
of Tropical Medicine, Antwerp, Belgium). On the basis of
detailed morphologic characters as described previously and
micrometry (average length 30.8 μm and width 1.9 μm) the
eukaryote was tentatively identified as a member of the subgenus
Herpetosoma, resembling Trypanosoma lewisi .
At IVRI, Izatnagar, DNA was extracted from 200 μL of the
blood sample using the GENEAID Genomic DNA Mini Kit
(Taipei, Taiwan) . Polymerase chain reaction (PCR) was performed
for amplification of the internal transcribed spacer
1 (ITS1) region, which is flanked by the 18S and 5.8S ribosomal
genes, and this yielded an amplicon of 623 bp. This corresponds
to the size of this region for T. lewisi . 1 Furthermore,
the organism was experimentally inoculated intraperitoneally
in laboratory-bred Swiss mice. The mice were negative for trypanosome
up to the 28th day post-inoculation. Trypanosoma
lewisi is host specific and fails to develop in mice. 1 A follow-up
investigation was conducted at Bagpat by the National Center
for Disease Control (formerly NICD), Delhi. Ten rats ( Rattus
rattus ) were trapped from the surroundings of the patient’s
house and blood was examined both microscopically and by
PCR at IVRI. Two of the 10 rats were also found to be infected
with the same hemoflagellate.
At the Institute of Tropical Medicine Antwerp (Belgium),
the World Health Organization (WHO) reference laboratory,
DNA was extracted from the blood on filter paper using the
QIAamp DNA micro kit (Qiagen, Valencia, CA). To check
if DNA was successfully extracted from the blood specimens,
Case Report: Trypanosoma lewisi or T. lewisi -like Infection in a 37-Day-Old Indian Infant
Archana Verma , Samiksha Manchanda , Nirmal Kumar , Archna Sharma , Masha Goel , Partha Sarathi Banerjee , Rajat Garg ,
Brahma Pal Singh , Fatima Balharbi , Veerle Lejon , Stijn Deborggraeve , Udai Veer Singh Rana , and Jacob Puliyel *
Department of Paediatrics, St. Stephens Hospital, Delhi, India; Department of Pathology, St. Stephens Hospital, Delhi, India;
Division of Parasitology, Indian Veterinary Research Institute, U.P., India; Institute of Tropical Medicine Antwerp, Belgium;
National Centre for Disease Control, Delhi, India
Abstract. Trypanosomes were observed in the peripheral blood smear of a 37-day-old Indian infant admitted off feeds,
with fever and convulsions. Trypanosoma ( Herpetosoma ) lewisi was identified in the blood. The species identification was
confirmed by morphometry, polymerase chain reaction, and sequencing. Human infection with this organism is rare. Only
seven cases of this infection have been reported previously in humans. The cases reported are reviewed to develop a composite
picture of this disease.
* Address correspondence to Jacob Puliyel, St. Stephens Hospital, Tis
Hazari, Delhi, India 110054. E-mail: puliyel@gmail.com
222 VERMA AND OTHERS
a control PCR targeting the human beta-globin gene was performed.
To identify the trypanosome, PCRs specific for the
Trypanosomatidae, 2 Trypanozoon , 3 T. evansi , 4 and T.lewisi 1
were performed. The PCR for T. evansi and Trypanozoon were
negative but it was positive for in the Trypanosomatidae . The
length of the amplified ITS1 DNA sequence corresponded
with that of T. lewisi described by Desquesnes and colleagues. 1
The ITS1 DNA sequence differentiates T. lewisi (amplicon
of 623 bp) from T. brucei and T. evansi (amplicon of 520 bp).
The ITS1 PCR product was sequenced at the University of
Antwerp, Belgium. The DNA sequence was analyzed with the
Bioedit software (Ibis Therapeutics, Carlsbad, CA) and aligned
with the T. lewisi DNA sequence reported by Desquesnes and
others using Multalin 5 (see Box 1 ).
Sequencing results. The upper ends of the ITS1 PCR
product could not be sequenced because sequencing was done
directly on the PCR product. Sequencing similarity of 90%
was observed.
Human trypanosomiasis is rare in India and the anti-
Trypanosoma medication for this child (Pentamidine and
Suramine) had to be sent from WHO, Geneva; this took
5 days. Pending availability of definitive treatment, the infant
was started empirically on Liposomal Amphotericin B along
with Ceftriaxone as given in infants with clinical signs and
symptoms of sepsis. Amphotericin B is an antifungal medication
used also in Leishmania infections, which is endemic in
India. The infant became asymptomatic after 3 days. The serial
blood reports are shown in Table 1 . Injection of Pentamidine
was started on Day 5 of admission with cardiac monitoring.
Blood sugar and blood pressure were also monitored. Ceftriaxone
and Phenytoin were stopped on Day 10 of admission.
Pen tamidine was continued for a total of 10 days. The numbers
of trypanosomes progressively decreased. On the seventh day
of admission, peripheral smear did not show the parasite.
The child was discharged on 17/08/2010. At the followup
examination on 4/10/10, the chile child was asymptomatic and
showed no parasites on peripheral blood smear and had normal
blood counts. His high-density lipoprotein (HDL) levels
were estimated at this time and found to be normal.
DISCUSSION
We describe here a case of T. lewisi infection in a 37-dayold
infant. There are only seven previously reported cases of
T. lewisi -like infection in humans ( Table 2 ). Infants seem to be
Figure 1. Sting marks seen on child’s leg.
Table 1
Investigation reports
Day 1 Day 2 Day 3 Day 5 Day 7 Day 8 Day 9 Day 10 Day 11 Day 12 Day 13 Day 14 Day 15
Hemoglobin (g/dL) 9.7 7.7 9.8 7.9 8.0 8.6 5.8 7.4 9.2 8.2 9.1 8.0 8.6
Total leukocyte
counts (×10 3 /μL) 8.7 8.0 9.4 5.9 6.2 8.3 6.7 6.8 11.7 7.5 7.8 8.5 8.9
Platelet counts
(×10 3 /μL) 27 41 51 68 317 151 542 467 416 591 477 601 570
Peripheral smear
remark
Tryp
seen
Tryp
seen
Tryp
seen
Tryp
seen
Tryp not
seen
Tryp not
seen
Tryp not
seen
Tryp not
seen
Tryp not
seen
Tryp not
seen
Tryp not
seen
Tryp not
seen
Tryp not
seen
Blood urea nitrogen
(mg/dL) 17.4 3.9 12.7 6.7 4.5 5.2 5.2 6.1 6.3 8.9
Creatinine (mg/dL) 0.3 0.3 0.1 0.3 0.2 0.3 0.3 0.2 0.3 0.3
Sodium (mmol/L) 132 145 133 133 134 145 146 143 146 134
Potassium (mmol/L) 4.6 4.91 5.6 4.6 5.0 6.31 5.7 5.68 5.53 5.1
Calcium (mg/dL) 9.9 9.1 10.1 9.8
Cerebrospinal fluid (CSF) analysis: Day 1 – Trypanosomes seen in blood stained CSF.
Day 2 – No trypanosomes seen in CSF. No biochemical abnormality.
Tryp – Trypanosoma .
Figure 2. Magnification 100×.
TRYPANOSOMA LEWISI INFECTION 223
Table 2
Blood smear showing Trypomastigotes *
Author year (reference) Place Age of patient Presenting complaints Method of identification Treatment given Outcome
Johnson, 1933 12 Malaysia 4 months Anorexia, lassitude, fever Morphology in blood None Recovered
Shrivastava and
others 1974 13
Madhya Pradesh,
India
35 years Fever, lassitude Morphology in blood Immune
fluorescence antibody test
Formol-gel test
None described Recovered
Shrivastava and
others 1974 13
Madhya Pradesh,
India
40 year Fever, lassitude Morphology in blood Immune
fluorescence antibody test
Formol-gel test
None described Recovered
Howie and others,
2003 15
Gambia 2 months Fever, generalized edema Morphology in blood and CSF
PCR analysis
Melarso- Prol Recovered
Kaur and others, 2006 14 Bombay India 2 months Fever Morphology in blood None described Recovered
Sarataphan and
others, 2007 11
Thailand 45 days Fever, cough, anorexia,
depression
Morphology on blood smear
dissimilar to T. lewisi . ITS1
sequence analysis and
amplicon size similar to
T. lewisi -like Herpetosoma
Injection
gentamycin
Recovered
Banerjee and
others, 2008 6
Pune, Maharashtra 55 years Intermittent fever,
anorexia, pedal oedema,
lethargy, splenomegaly,
and hepatomegaly
Morphology similar to T. lewisi .
ITS1 amplicon size similar to
T. lewisi
Injection
suramin
Died
* CSF = cerebrospinal fluid; PCR = polymerase chain reaction; ITS1 = internal transcribed spacer.
more vulnerable to this infection. Five of the eight infections
so far have been in infants. This might be attributed to immunologically
naive status of this group of patients. 6 Our infant is
arguably the youngest case to be reported until now.
The first case of human trypanosomiasis reported from
India was a case of T. evansi in a herdsman of village Seoni in
Chandrapur district of Maharashtra in 2004. 7– 9 Trypanosoma
evansi is also the most commonly occurring hemoflagellate
of domestic animals in India. Trypanosoma evansi can be
cultured easily in mice within 3 to 5 days. Failure of development
in mice is a good indicator that the organism, in all probability,
is not T. evansi . Trypanosoma evansi can also be cultured
in rats, but it takes more time to develop. Trypanosoma lewisi
is more host specific and fails to develop in mice but often
can be cultured in rats. Parasitemia is intermittent and rate of
development depends upon the strain of rats used.
It was postulated earlier that a trypanolytic factor in human
plasma prevents infection with zoonotic trypanosomes. Lack
Box 1
Alignment sequence of the internal transcribed spacer 1 (ITS1) polymerase chain reaction (PCR) product from the child’s blood and T. lewisi ITS1
product, Desquesnes and others. 1
224 VERMA AND OTHERS
of this trypanolytic factor is seen in persons with low HDL
levels and this can make a person vulnerable to trypanosomiasis.
10 Our infant had normal HDL levels.
Fever and lassitude were the chief presenting complaints
in all the cases reported previously. Four of the seven cases
reported previously were identified only by morphology. The
ITS1 PCR has been used for identification of human infection
with T. lewisi in two cases so far. In one infant from Thailand,
ITS1 PCR identified T. lewisi infection but the morphology of
the organism was different, 11 and this suggests that the identification
based on ITS1 PCR is not specific. The ITS1 PCR
tallied with the morphology in our case. We further confirmed
species by DNA sequencing.
In four cases no specific treatment was administered 12– 14 ;
these cases recovered clinically and showed disappearance of
trypanosomes from blood films. Follow-up did not show any
relapses. However, a 55 year old, with the infection, died in spite
of receiving Suramine. 6 One case reported use of Gentamycin
injections for treatment. 11 The Gambian infant was treated with
Melarsoprol. 15 Our infant responded symptomatically within
3 days of admission, while being administered Liposomal
Amphotericin B and Ceftriaxone. The parasitemia however
persisted albeit in reduced numbers up to Day 7. Pentamidine
was started on Day 5, but in retrospect, it is difficult to say
that it was needed for clearing the parasites. Specific treatment
with anti-trypanosomal drugs (Melarsoprol and Suramine)
was given in only two of the seven previously reported cases.
It appears to be a self-limiting infection in humans. However,
given the present evidence it will be prudent to prescribe antibiotics
in an infant with clinical evidence of sepsis, even if
T. lewisi is detected in the blood film. Aggressive anti-trypanosomal
treatment is perhaps not indicated.
In rats, T. lewisi is an infection transmitted by the excreta of
fleas through contamination of rat food or ingestion of fleas
by the rats. The route of transmission to humans is unclear.
Our child had bite marks over the left leg. We cannot say with
certainty if these were flea bites transmitting infection to our
child .
Received January 1, 2011. Accepted for publication April 30, 2011.
Acknowledgment: We acknowledge WHO (Dr. P. Simarro, World
Health Organization, Control of Neglected Tropical Diseases, Innovative
and Intensified Disease Management, Geneva, Switzerland) for
technical assistance. The American Society of Tropical Medicine and
Hygiene (ASTMH) assisted with publication expenses.
Authors’ addresses: Archana Verma, Samiksha Manchanda, and
Nirmal Kumar, Department of Pediatrics, St. Stephens Hospital,
Delhi, India, E-mails: dr_archanaverma@yahoo.com , manchanda
sami@yahoo.com , and nsk9_2000@yahoo.com . Archna Sharma and
Masha Goel, Department of Pathology, St. Stephens Hospital, Delhi,
India, E-mails: drarchna_sharma@yahoo.com and Mashagoel86@
gmail.com . Partha Sarathi Banerjee, Rajat Garg, and Brahma Pal
Singh, Division of Parasitology, Indian Veterinary Research Institute,
U.P., India, E-mails: banerjeeparth62@gmail.com , rajatgarg_2000@
yahoo.com , and bpsingh_para@yahoo.co.in . Fatima Balharbi, Veerle
Lejon, and Stijn Deborggraeve, Department of Parasitology, Institute
of Tropical Medicine, Antwerp, Belgium, E-mails: fbalharbi@itg.be ,
vlejon@itg.be , and sdeborggraeve@itg.be . Udai Veer Singh Rana,
National Centre for Disease Control, Delhi, India, E-mail: druvsrana@
gmail.com . Jacob Puliyel, St. Stephens Hospital, Delhi, India, E-mail:
puliyel@gmail.com .
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