The classical diagnosis of human African trypanosomiasis (HAT) is a three-step approach:
Identification of suspects is based on determining whether they have lived in or visited an endemic area, in combination with clinical signs and symptoms or with serological evidence (presence of trypanosome-specific antibodies in blood).
Parasite detection is considered as the confirmation of infection. It is relatively easy for T b rhodesiense, but often difficult with T b gambiense, where failure to demonstrate the parasite is not uncommon, even if concentration techniques are applied. For that reason, in some epidemiological situations, treatment decision is taken following serological and clinical evidence.[27]Chappuis F, Stivanello E, Adams K, et al. Card agglutination test for trypanosomiasis (CATT) end-dilution titer and cerebrospinal fluid cell count as predictors of human African trypanosomiasis (Trypanosoma brucei gambiense) among serologically suspected individuals in Southern Sudan. Am J Trop Med Hyg. 2004 Sep;71(3):313-7.
http://www.ncbi.nlm.nih.gov/pubmed/15381812?tool=bestpractice.com
Once parasites have been detected, the disease stage, in combination with the subspecies of causative trypanosome, determines the therapeutic choices for treatment.
History and clinical examination
Initial suspicion should be high in patients who have visited a HAT-endemic area in sub-Saharan Africa and present with a history of a tsetse bite, although the bite may go unnoticed.
Symptoms and signs are non-specific and variable and are, therefore, insufficient for diagnosis and staging. The onset of symptoms is usually more acute in individuals originating from non-endemic areas.[7]Duggan AJ, Hutchinson MP. Sleeping sickness in Europeans: a review of 109 cases. J Trop Med Hyg. 1966 Jun;69(6):124-31.
http://www.ncbi.nlm.nih.gov/pubmed/5940665?tool=bestpractice.com
[28]Urech K, Neumayr A, Blum J. Sleeping sickness in travelers - do they really sleep? PLoS Negl Trop Dis. 2011 Nov;5(11):e1358.
https://journals.plos.org/plosntds/article?id=10.1371/journal.pntd.0001358
http://www.ncbi.nlm.nih.gov/pubmed/22069503?tool=bestpractice.com
Patients often present with a history of malaria treatment without improvement.[29]Odiit M, Shaw A, Welburn SC, et al. Assessing the patterns of health-seeking behaviours and awareness among sleeping-sickness patients in eastern Uganda. Ann Trop Med Parasitol. 2004 Jun;98(4):339-48.
http://www.ncbi.nlm.nih.gov/pubmed/15228715?tool=bestpractice.com
[30]Bukachi SA, Wandibba S, Nyamongo IK. The treatment pathways followed by cases of human African trypanosomiasis in western Kenya and eastern Uganda. Ann Trop Med Parasitol. 2009 Apr;103(3):211-20.
http://www.ncbi.nlm.nih.gov/pubmed/19341536?tool=bestpractice.com
Symptoms and signs that may be seen in the first stage of disease include:
Fatigue
Headache
Fever
General malaise
Enlarged cervical lymph nodes (Winterbottom's sign; mainly T b gambiense)
Inoculation chancre (mainly T b rhodesiense)
Pruritus
Rash
Oedema
Hepatosplenomegaly
Endocrine dysfunctions (in women: history of infertility, amenorrhoea, and miscarriage; in men: reduced libido, impotence)
Patients with gambiense HAT progress from first to second stage after a mean time of 500 days.[31]Checchi F, Filipe JA, Haydon DT, et al. Estimates of the duration of the early and late stage of gambiense sleeping sickness. BMC Infect Dis. 2008 Feb 8;8:16.
https://bmcinfectdis.biomedcentral.com/articles/10.1186/1471-2334-8-16
http://www.ncbi.nlm.nih.gov/pubmed/18261232?tool=bestpractice.com
In second-stage gambiense HAT, signs of first stage persist and signs of neurological involvement become apparent, including:[32]World Health Organization. Guidelines for the treatment of human African trypanosomiasis. Jun 2024 [internet publication].
https://www.who.int/publications/i/item/9789240096035
Disturbances of consciousness and sleep (although this can also be present in first-stage disease)
Impaired motor functions (e.g., ataxia, disturbance of gait, tremors, abnormal movements)
Sensory disorders
Mental changes (e.g., inattention, disorientation in time and/or space, slowed thought processes, memory deficit)
Abnormal behaviour (e.g., disinhibition, excitement, euphoria, aggressiveness, indifference)
In patients with rhodesiense HAT, the duration of symptoms suggests that the disease progresses into second stage between 3 weeks and 2 months of infection.[33]Odiit M, Kanshme F, Enyaru JCK. Duration of symptoms and case fatality of sleeping sickness caused by Trypanosoma brucei rhodesiense in Tororo, Uganda. East Afr Med J. 1997 Dec;74(12):792-5.
http://www.ncbi.nlm.nih.gov/pubmed/9557424?tool=bestpractice.com
Because of this acuteness, neurological involvement may remain absent. Patients are usually admitted presenting with multi-organ or cardiac failure with abnormal electrocardiogram results.[28]Urech K, Neumayr A, Blum J. Sleeping sickness in travelers - do they really sleep? PLoS Negl Trop Dis. 2011 Nov;5(11):e1358.
https://journals.plos.org/plosntds/article?id=10.1371/journal.pntd.0001358
http://www.ncbi.nlm.nih.gov/pubmed/22069503?tool=bestpractice.com
[34]Cottle LE, Peters JR, Hall A, et al. Multiorgan dysfunction caused by travel-associated African trypanosomiasis. Emerg Infect Dis. 2012 Feb;18(2):287-9.
https://wwwnc.cdc.gov/eid/article/18/2/11-1479_article
http://www.ncbi.nlm.nih.gov/pubmed/22305185?tool=bestpractice.com
Laboratory investigations
Although not sufficient to establish the diagnosis of HAT, common blood abnormalities include:[35]Chappuis F, Loutan L, Simarro P, et al. Options for the field diagnosis of human African trypanosomiasis. Clin Microbiol Rev. 2005 Jan;18(1):133-46.
https://journals.asm.org/doi/10.1128/cmr.18.1.133-146.2005
http://www.ncbi.nlm.nih.gov/pubmed/15653823?tool=bestpractice.com
[36]Greenwood BM, Whittle HC. The pathogenesis of sleeping sickness. Trans R Soc Trop Med Hyg. 1980;74(6):716-25.
http://www.ncbi.nlm.nih.gov/pubmed/7010694?tool=bestpractice.com
Anaemia: mild for gambiense HAT, but severe in rhodesiense HAT
Thrombocytopenia: mild for gambiense HAT, may be severe in rhodesiense HAT
Moderate leukocytosis
Increased erythrocyte sedimentation rate (ESR)
Increased immunoglobulin (Ig) levels, especially IgM
Therefore, a full blood count, ESR, and serum immunoglobulins can be ordered to support the diagnosis.
The presence of a variety of non-specific antibodies has been reported during T b gambiense infection and consequently constitutes a risk for misdiagnosis, leaving HAT undetected.
Trypanosome-specific antibodies in blood are detectable by:
The card agglutination test for trypanosomiasis (CATT) for T b gambiense. This test is applied for screening of the population at risk in T b gambiense-endemic areas.
Enzyme-linked immunosorbent assay (ELISA) or immunofluorescence for T b gambiense and T b rhodesiense.
Rapid blood tests have been developed for detection of trypanosome-specific antibodies. They are an alternative to CATT and offer the advantage of being individual tests. They are, therefore, particularly suited for use in non-specialised centres that are occasionally confronted with T b gambiense HAT.[37]Sullivan L, Wall SJ, Carrington M, et al. Proteomic selection of immunodiagnostic antigens for human African trypanosomiasis and generation of a prototype lateral flow immunodiagnostic device. PLoS Negl Trop Dis. 2013;7(2):e2087.
https://journals.plos.org/plosntds/article?id=10.1371/journal.pntd.0002087
http://www.ncbi.nlm.nih.gov/pubmed/23469310?tool=bestpractice.com
[38]Büscher P, Gilleman Q, Lejon V. Rapid diagnostic test for sleeping sickness. N Engl J Med. 2013 Mar 14;368(11):1069-70.
https://www.nejm.org/doi/full/10.1056/NEJMc1210373
http://www.ncbi.nlm.nih.gov/pubmed/23484849?tool=bestpractice.com
[39]Büscher P, Mertens P, Leclipteux T, et al. Sensitivity and specificity of HAT Sero-K-SeT, a rapid diagnostic test for serodiagnosis of sleeping sickness caused by Trypanosoma brucei gambiense: a case-control study. Lancet Glob Health. 2014 Jun;2(6):e359-63.
https://www.thelancet.com/journals/langlo/article/PIIS2214-109X(14)70203-7/fulltext
http://www.ncbi.nlm.nih.gov/pubmed/25103304?tool=bestpractice.com
[40]Bisser S, Lumbala C, Nguertoum E, et al. Sensitivity and specificity of a prototype rapid diagnostic test for the detection of Trypanosoma brucei gambiense infection: a multi-centric prospective study. PLoS Negl Trop Dis. 2016 Apr 8;10(4):e0004608.
https://journals.plos.org/plosntds/article?id=10.1371/journal.pntd.0004608
http://www.ncbi.nlm.nih.gov/pubmed/27058033?tool=bestpractice.com
Identification of a suspected case of HAT, based either on clinical findings or on serological tests, should be followed by microscopic examination for the presence of parasites:
The stained, thick drop examination on blood is the most commonly applied microscopic examination and allows differential diagnosis of malaria. The technique might remain negative in T b gambiense infections, which are characterised by low parasite levels in blood.
Blood concentration techniques, such as the microhaematocrit centrifugation technique, buffy coat technique, or mini-anion exchange centrifugation technique (mAECT), are indicated for detection of T b gambiense. Repetitive examinations might be needed for findings of T b gambiense. mAECT is considered the best test, with sensitivity around 75% to 85%.[41]Miezan TW, Meda AH, Doua F, et al. Evaluation of the parasitologic techniques used in the diagnosis of human Trypanosoma gambiense trypanosomiasis in the Ivory Coast [in French]. Bull Soc Pathol Exot. 1994;87(2):101-4.
http://www.ncbi.nlm.nih.gov/pubmed/8061525?tool=bestpractice.com
[42]Lutumba P, Robays J, Miaka C, et al. Validity, cost and feasibility of the mAECT and CTC confirmation tests after diagnosis of African of sleeping sickness [in French]. Trop Med Int Health. 2006 Apr;11(4):470-8.
http://www.ncbi.nlm.nih.gov/pubmed/16553930?tool=bestpractice.com
Sensitivity of mAECT can be increased to between 91% and 96% when the buffy coat of larger volumes of blood is applied onto the column.[43]Camara M, Camara O, Ilboudo H, et al. Sleeping sickness diagnosis: use of buffy coats improves the sensitivity of the mini anion exchange centrifugation test. Trop Med Int Health. 2010 Jul;15(7):796-9.
https://onlinelibrary.wiley.com/doi/10.1111/j.1365-3156.2010.02546.x
http://www.ncbi.nlm.nih.gov/pubmed/20497407?tool=bestpractice.com
[44]Mumba Ngoyi D, Ali Ekangu R, Mumvemba Kodi MF, et al. Performance of parasitological and molecular techniques for the diagnosis and surveillance of gambiense sleeping sickness. PLoS Negl Trop Dis. 2014 Jun 12;8(6):e2954.
https://journals.plos.org/plosntds/article?id=10.1371/journal.pntd.0002954
http://www.ncbi.nlm.nih.gov/pubmed/24921941?tool=bestpractice.com
It is generally accepted that cases of trypanosomiasis are defined by the microscopic demonstration of trypanosomes in any body fluid. Occasionally, cases of gambiense HAT have been defined serologically, such as the case of a patient living an endemic area who has a positive serology in CATT, using different titration thresholds.[27]Chappuis F, Stivanello E, Adams K, et al. Card agglutination test for trypanosomiasis (CATT) end-dilution titer and cerebrospinal fluid cell count as predictors of human African trypanosomiasis (Trypanosoma brucei gambiense) among serologically suspected individuals in Southern Sudan. Am J Trop Med Hyg. 2004 Sep;71(3):313-7.
http://www.ncbi.nlm.nih.gov/pubmed/15381812?tool=bestpractice.com
[45]Simarro PP, Ruiz JA, Franco JR, et al. Attitude towards CATT-positive individuals without parasitological conformation in the African trypanosomiasis (T b gambiense) focus of Quiçama (Angola). Trop Med Int Health. 1999 Dec;4(12):858-61.
https://onlinelibrary.wiley.com/doi/10.1046/j.1365-3156.1999.00494.x
http://www.ncbi.nlm.nih.gov/pubmed/10632994?tool=bestpractice.com
[46]Inojosa WO, Augusto I, Bisoffi Z, et al. Diagnosing human African trypanosomiasis in Angola using card agglutination test: observational study of active and passive case finding strategies. BMJ. 2006 Jun 24;332(7556):1479.
https://www.bmj.com/content/332/7556/1479
http://www.ncbi.nlm.nih.gov/pubmed/16777858?tool=bestpractice.com
Other body fluids that can be examined microscopically for the presence of trypanosomes include:
Lymph node aspirate for T b gambiense
Chancre aspirate for T b rhodesiense
Cerebrospinal fluid (CSF)
Disease staging
Once the infection has been confirmed, or in the case of a high index of suspicion for infection (such as an elevated CATT and titre and/or neurological symptoms), a lumbar puncture should be performed and the CSF examined for disease staging to help guide treatment.[32]World Health Organization. Guidelines for the treatment of human African trypanosomiasis. Jun 2024 [internet publication].
https://www.who.int/publications/i/item/9789240096035
Patients who can be managed without lumbar puncture
Patients who require lumbar puncture
If lumbar puncture and CSF examination is required, disease staging for gambiense and rhodesiense HAT is as follows:[32]World Health Organization. Guidelines for the treatment of human African trypanosomiasis. Jun 2024 [internet publication].
https://www.who.int/publications/i/item/9789240096035
The second stage of gambiense HAT can be further classified as severe.
Emerging investigations
Although brain MRI has no role in diagnosis of HAT, meningeal thickening and bilateral confluent hyperintensive T2 signal of supratentorial white matter, brainstem, and cerebellum may be seen. Abnormalities remain present after successful cure.[47]Kager PA, Schipper HG, Stam J, et al. Magnetic resonance imaging findings in human African trypanosomiasis: a four-year follow-up study in a patient and review of the literature. Am J Trop Med Hyg. 2009 Jun;80(6):947-52.
http://www.ncbi.nlm.nih.gov/pubmed/19478256?tool=bestpractice.com
[48]Boukobza M, Lariven S, Houzé S, et al. Unusual MRI findings in African Trypanosoma brucei gambiense trypanosomiasis: dentate nuclei and hypothalamic lesions. Am J Trop Med Hyg. 2020 Jan;102(1):5-6.
https://www.ajtmh.org/view/journals/tpmd/102/1/article-p5.xml
http://www.ncbi.nlm.nih.gov/pubmed/31971136?tool=bestpractice.com
Immune trypanolysis is an emerging test in which live T b gambiense trypanosomes are incubated with human serum or plasma.[49]Van Meirvenne N, Magnus E, Büscher P. Evaluation of variant specific trypanolysis tests for serodiagnosis of human infections with Trypanosoma brucei gambiense. Acta Trop. 1995 Dec;60(3):189-99.
http://www.ncbi.nlm.nih.gov/pubmed/8907397?tool=bestpractice.com
[50]Jamonneau V, Bucheton B, Kaboré J, et al. Revisiting the immune trypanolysis test to optimise epidemiological surveillance and control of sleeping sickness in West Africa. PLoS Negl Trop Dis. 2010 Dec 21;4(12):e917.
https://journals.plos.org/plosntds/article?id=10.1371/journal.pntd.0000917
http://www.ncbi.nlm.nih.gov/pubmed/21200417?tool=bestpractice.com
The specificity of immune trypanolysis is considered to be 100%, and a positive immune trypanolysis appears to be an indicator for contact with T b gambiense. The test can be performed only in highly specialised laboratories.
Various CSF proteins and markers of cellular immune system activation, as well as detection of RNA, are being investigated as markers for staging and treatment outcome assessment.[51]Tiberti N, Hainard A, Lejon V, et al. Cerebrospinal fluid neopterin as marker of the meningo-encephalitic stage of Trypanosoma brucei gambiense sleeping sickness. PLoS One. 2012;7:e40909.
https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0040909
http://www.ncbi.nlm.nih.gov/pubmed/22815865?tool=bestpractice.com
[52]Amin DN, Mumba Ngoyi D, Nhkwachi GM, et al. Identification of stage biomarkers for human African trypanosomiasis. Am J Trop Med Hyg. 2010 Jun;82(6):983-90.
https://pmc.ncbi.nlm.nih.gov/articles/PMC2877438
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[53]Tiberti N, Lejon V, Hainard A, et al. Neopterin is a cerebrospinal fluid marker for treatment outcome evaluation in patients affected by Trypanosoma brucei gambiense sleeping sickness. PLoS Negl Trop Dis. 2013;7(2):e2088.
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[54]Ngay Lukusa I, Van Reet N, Mumba Ngoyi D, et al. Trypanosome SL-RNA detection in blood and cerebrospinal fluid to demonstrate active gambiense human African trypanosomiasis infection. PLoS Negl Trop Dis. 2021 Sep;15(9):e0009739.
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[55]Ilboudo H, Camara O, Ravel S, et al. Trypanosoma brucei gambiense spliced leader RNA is a more specific marker for cure of human African trypanosomiasis than T. b. gambiense DNA. J Infect Dis. 2015 Dec 15;212(12):1996-8.
https://academic.oup.com/jid/article/212/12/1996/2911942
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[56]González-Andrade P, Camara M, Ilboudo H, et al. Diagnosis of trypanosomatid infections: targeting the spliced leader RNA. J Mol Diagn. 2014 Jul;16(4):400-4.
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