Conceived and designed the experiments: VJ BB PS RK KL MC VL PB. Performed the experiments: VJ JK HI OC FC DK RB. Analyzed the data: VJ BB JK HI OC FC DK RK KL MC RB VL PB. Contributed reagents/materials/analysis tools: VJ JK HI OC FC DK RK KL MC RB PB. Wrote the paper: VJ BB PS VL PB.
The authors have declared that no competing interests exist.
Because of its high sensitivity and its ease of use in the field, the card agglutination test for trypanosomiasis (CATT) is widely used for mass screening of sleeping sickness. However, the CATT exhibits false-positive results (i) raising the question of whether CATT-positive subjects who are negative in parasitology are truly exposed to infection and (ii) making it difficult to evaluate whether
TL was performed on plasma collected from CATT-positive persons identified within medical surveys in several West African HAT foci in Guinea, Côte d'Ivoire and Burkina Faso with diverse epidemiological statuses (active, latent, or historical). All HAT cases were TL+. All subjects living in a nonendemic area were TL−. CATT prevalence was not correlated with HAT prevalence in the study areas, whereas a significant correlation was found using TL.
TL appears to be a marker for contact with
Human African trypanosomiasis (HAT) due to
Human African trypanosomiasis (HAT) or sleeping sickness is caused by two subspecies of the protozoan flagellate
After the successful control campaigns dating from 1930 to 1960,
Mass screening of the population at risk of
In the current elimination context in West Africa, when prevalence becomes low or transmission has stopped, the limited specificity of CATT becomes a considerable drawback because it results in low positive predictive values
In a previous study, the immune trypanolysis test (TL) was shown to be a promising tool to help better understand the phenomenon of nonconfirmed CATT-positive subjects
The objective of the present study was to evaluate the use of TL with
All samples were collected within the framework of medical surveys conducted by the national HAT control programmes (NCP) according to the respective national HAT diagnostic procedures. No samples other than those collected for routine screening and diagnostic procedures were collected for the purposes of the present study. All participants were informed of the objective of the study in their own language and signed a written informed consent form. Children less than 12 years old were excluded. For participants between 12 and 18 years of age, informed consent was obtained from their parents. This study is part of a larger project aiming at improving HAT diagnosis for which approval was obtained from WHO (Research Ethics Review Committee) and Institut de Recherche pour le Développement (Comité Consultatif de Déontologie et d'Ethique) ethical committees.
Specimens were collected in three West African countries (Guinea, Côte d'Ivoire and Burkina Faso) in foci with a different epidemiological HAT status (
The Dubreka/Boffa focus is situated north of Conakry in the coastal mangroves and is currently the most active West African focus with a prevalence of about 1%
The Forécariah focus is situated south of Conakry in the coastal mangroves near Sierra Leone. Sporadic HAT cases reported at the Dubreka treatment centre come from this area.
The N'Zérékoré focus is situated in the woodlands between the savannah and the mesophilic forest near the border with Côte d'Ivoire. It is a historical HAT focus with a risk of re-emergence in the context of socio-political instability and populations moving between Liberia, Côte d'Ivoire and Guinea. Very few recent epidemiological data are available from this area.
The Bonon focus is situated in the Western central part of the country, between the savannah and the mesophilic forest. Between 1998 and 2003, HAT prevalence in Bonon was about 0.4%
Historical HAT foci of Folonzo, Loropéni and Batié (Fol/Lor/Bat) are located in the South-western part of the country. These areas were recently put under epidemiological surveillance because of a risk of re-emergence of the disease due to the return of agricultural workers from coffee plantations in Côte d'Ivoire where HAT is endemic. Tsetse flies and animal African trypanosomiasis are still present in the area
All persons participating in the study were identified during active screening campaigns organised by the NCPs in Guinea, Burkina Faso and Côte d'Ivoire during HAT surveillance activities. Only subjects positive to the CATT/
HAT (patients): CATT-P end titer ≥1/8 and parasitologically confirmed;
SERO (seropositives): CATT-P end titer ≥1/8 but no parasites detected;
SUSP (suspects): CATT-B-positive and CATT-P <1/8 but no parasites detected.
The origin and numbers of participants in each group are detailed in
Country | Focus | SUSP | SERO | HAT | Total |
Guinea | Dubreka/Boffa | 0 | 17 | 37 | 54 |
Guinea | Forécariah | 0 | 30 | 28 | 58 |
Côte d'Ivoire | Bonon | 86 | 24 | 6 | 116 |
Guinea | N'Zérékoré | 0 | 16 | 0 | 16 |
Burkina Faso | Fol/Lor/Bat | 18 | 25 | 0 | 43 |
Fol/Lor/Bat = Folonzo, Loropéni and Batié.
SUSP, SERO and HAT are defined in the
Cloned populations of
A total of 43,373 persons were screened with CATT/
Study site | Examined population | CATT-B pos | CATT-P pos | HAT | |||
number | % | number | % | number | % | ||
Dubreka/Boffa | 6795 | 124 | 1.82 | 56 | 0.82 | 39 | 0.57 |
Forécariah | 17571 | 167 | 0.95 | 63 | 0.36 | 28 | 0.16 |
Bonon | 3305 | 128 | 3.87 | 40 | 1.21 | 6 | 0.18 |
N'Zérékoré | 4853 | 102 | 2.10 | 16 | 0.33 | 0 | 0.00 |
Fol/Lor/Bat | 10849 | 132 | 1.22 | 34 | 0.31 | 0 | 0.00 |
Total | 43373 | 653 | 1.51 | 209 | 0.48 | 73 | 0.17 |
Fol/Lor/Bat = Folonzo, Loropéni and Batié; pos = positive.
CATT-P pos = CATT-P end titer ≥1/8.
The results of TL on the 287 subjects included in the study are summarised in
Category | SUSP | SERO | HAT | ||||||||
Study sites | Prev | TL+ | TL- | %TL+ | TL+ | TL- | %TL+ | TL+ | TL- | %TL+ | |
Dub/Bof | 0.57% | 0 | 0 | na | 15 | 2 | 88.2 | 37 | 0 | 100 | |
Forécariah | 0.16% | 0 | 0 | na | 18 | 12 | 60 | 28 | 0 | 100 | |
Bonon | 0.18% | 10 | 76 | 11.6 | 7 | 17 | 29.2 | 6 | 0 | 100 | |
N'Zérékoré | 0.00% | 0 | 0 | na | 0 | 16 | 0 | 0 | 0 | na | |
Fol/Lor/Bat | 0.00% | 0 | 18 | 0 | 1 | 24 | 4 | 0 | 0 | na | |
Total | 10 | 94 | 9.6 | 41 | 71 | 36.6 | 71 | 0 | 100 |
Dub/Bof = Dubreka/Boffa; Fol/Lor/Bat = Folonzo, Loropéni and Batié, prev = HAT prevalence; na = not available; SUSP, SERO and HAT are defined in the
All 71 HAT patients were TL+. All 18 SUSP from For/Lor/Bat in Burkina Faso were TL− while in Bonon 10 of 86 (11.6%) were TL+. Among the SERO subjects, 41 of 112 (36.6%) were TL+. Interestingly, the percentage of TL+ subjects in the SERO group was correlated with HAT prevalence (r2 = 0.84,
South Western Burkina = Folonzo, Loropéni and Batié. The left Y-axis represents the prevalence of HAT (number of HAT cases/examined population) and SERO (number of subjects with CATT-P end titer ≥1/8 but no parasites detected/examined population). The right Y-axis represents the proportion of SERO individuals that were positive to the trypanolysis test.
LiTat | SUSP | SERO | HAT | |||
Study sites | 1.3 | 1.5 | 1.6 | nb | nb | nb |
Dubreka/Boffa | + | + | + | na | 10 | 30 |
+ | + | − | na | 5 | 7 | |
Forécariah | + | + | + | na | 2 | 15 |
+ | + | − | na | 15 | 13 | |
+ | − | − | na | 1 | 0 | |
Bonon | + | + | + | 6 | 7 | 6 |
+ | + | − | 1 | 0 | 0 | |
+ | − | + | 1 | 0 | 0 | |
+ | − | − | 1 | 0 | 0 | |
− | − | + | 1 | 0 | 0 | |
Fol/Lor/Bat | + | + | + | 0 | 1 | na |
Fol/Lor/Bat = Folonzo, Loropéni and Batié; nb = number; na = not available; SUSP, SERO and HAT are defined in the
This study shows that high prevalence of CATT-positive individuals can be found even in areas were transmission has stopped, presumably owing to false positivity. On the contrary, positivity of TL in SERO subjects was significantly correlated with HAT prevalence and not in nonendemic areas. Thus TL is a useful tool, both to define the epidemiological status of an area when no HAT cases are diagnosed and to improve the monitoring of CATT-positive subjects with no parasitological confirmation, who are currently left out of HAT control strategies in most endemic countries.
The HAT prevalence rates observed in this study are in agreement with recent data on HAT epidemiology in West Africa. Guinea was the most affected country, with 0.57% HAT prevalence in Dubreka-Boffa and 0.16% in the Forécariah focus. No HAT cases were diagnosed in the N'Zérékoré focus. With 0.18% prevalence, HAT is still endemic in the Bonon focus in Côte d'Ivoire. The disease did not re-emerge in the historical foci of Burkina Faso despite the return of agricultural workers from active HAT foci in Côte d'Ivoire since 2002
TL was found to be highly sensitive (100% of HAT cases were TL+). Among SUSP and SERO persons, TL+ individuals were only found in areas with proven transmission, except one SERO person in the Fol/Lor/Bat focus in Burkina Faso, which is no longer active. It is noteworthy that this person had worked for 4 years in coffee and cacao plantations in a known HAT focus in Côte d'Ivoire where he may have been exposed to
Assuming TL is a marker of exposure to
These individuals could be patients in an early step of infection with as yet undetectable trypanosomes in blood, lymph or cerebrospinal fluid.
They could be asymptomatic carriers with very low parasitaemia. In this study, 53% of the SERO TL+ persons were positive on PCR using the TBR1/2 primers
They could have experienced a transient episode of
Concordant with TL positivity being a marker of contact with
At the individual level, TL can represent a tool for NCPs to identify among CATT-positives those who should be followed up by CATT and parasitological investigations until CATT becomes negative or the person is confirmed as a HAT patient. Whether these seropositives should undergo treatment remains an open question as long as their role in HAT transmission is unknown. We are currently carrying out follow-ups of SERO subjects. Preliminary results indicate that SERO TL+ individuals maintain a strong serological response (CATT and TL) over time, whereas SERO TL− subjects become CATT-negative within several months. Furthermore, HAT patients confirmed during these follow-ups were all from the SERO TL+ cohort (Bucheton, personal communication). In some countries, treatment of unconfirmed persons with CATT-P titers ≥1/16 is already recommended
At the population level, TL performed on CATT-positive individuals could be a valuable decision tool for NCPs to plan control measures (
HAT = presence of HAT cases; No HAT = absence; SERO = presence of subjects with CATT-P end titer ≥1/8 but no parasites detected, No SERO = absence; TL+ = positive in trypanolysis test; TL− = negative; * except for a special event, such as population movements, occurs.
From a practical point of view, the implementation of TL in NCP is hampered by its technological requirements (cryobiology and laboratory animal facilities, availability of VAT-specific control sera, etc.). An alternative test that is applicable in the field and that allows combining several VATs in a single test is the indirect agglutination test LATEX/
In conclusion, application of the TL test within the framework of medical surveys has provided a better picture of HAT epidemiology in West Africa, thanks to a better characterisation of parasitologically unconfirmed CATT positive subjects. The proportion of TL+ subjects among CATT+ individuals was associated with active HAT foci and can thus be used as a marker for exposure to
We particularly acknowledge all the technicians from (i) the HAT team of the Institut Pierre Richet (Abidjan, Côte d'Ivoire), (ii) the HAT team of the Sinfra health district (Côte d'Ivoire), (iii) the Centre de Santé Urbain of Bonon (Côte d'Ivoire) (iv) the HAT team of the CIRDES (Bobo-Dioulasso, Burkina-Faso) and (v) the HAT NCP of Burkina-Faso, Côte d'Ivoire and Guinea for their help in sampling.