Quantifying the level of under-detection of Trypanosoma brucei rhodesiense sleeping sickness cases.

To formally quantify the level of under-detection of Trypanosoma brucei rhodesiense sleeping sickness (SS) during an epidemic in Uganda, a decision tree (under-detection) model was developed; concurrently, to quantify the subset of undetected cases that sought health care but were not diagnosed, a deterministic (subset) model was developed. The values of the under-detection model parameters were estimated from previously published records of the duration of symptoms prior to presentation and the ratio of early to late stage cases in 760 SS patients presenting at LIRI hospital, Tororo, Uganda during the 1988--1990 epidemic of SS. For the observed early to late stage ratio of 0.47, we estimate that the proportion of under-detection in the catchment area of LIRI hospital was 0.39 (95% CI 0.37--0.41) i.e. 39% of cases are not reported. Based on this value, it is calculated that for every one reported death of SS, 12.0 (95% CI 11.0--13.0) deaths went undetected in the LIRI hospital catchment area - i.e. 92% of deaths are not reported. The deterministic (subset) model structured on the possible routes of a SS infection to either diagnosis or death through the health system or out of it, showed that of a total of 73 undetected deaths, 62 (CI 60-64) (85%) entered the healthcare system but were not diagnosed, and 11 (CI 11--12) died without seeking health care from a recognized health unit. The measure of early to late stage presentation provides a tractable measure to determine the level of rhodesiense SS under-detection and to gauge the effects of interventions aimed at increasing treatment coverage.

A burgeoning epidemic of sleeping sickness in Uganda.

The epidemic of Trypanosoma brucei rhodesiense sleeping sickness in eastern Uganda, which began in 1998 as a result of movements of the livestock reservoir of the parasite, has continued to spread. An additional 133 000 people have been put at risk of infection in Kaberamaido, another newly affected district. The few resources committed to control interventions in Soroti district have failed to contain the epidemic. The high prevalence of the parasite in cattle presents a significant risk for transmission to human beings and further spread of this neglected zoonotic disease. Targeted interventions are urgently needed to control epidemics and reduce the high mortality resulting from sleeping sickness.

Implications of the re-invasion of Southeast Uganda by Glossina pallidipes on the epidemiology of bovine trypanosomosis.

A study to assess the influence of re-invasion of Glossina pallidipes on the epidemiology of bovine trypanosomosis was conducted in Southeast Uganda. A total of 1,992 cattle were screened in villages, with (949) and without G. pallidipes (1043) for trypanosomosis using a combination of the BCT and HCT methods. The prevalence of trypanosomosis (15.5%), Trypanosoma brucei infection (1.4%), T. congolense infection (7.2%), T. vivax infection (5.3%) and mixed infection (1.6%) in cattle in villages with was significantly higher than in those without G. pallidipes: trypanosomosis (7.1%), T. brucei infection (0.6%), T. congolense infection (2.0%), T. vivax infection (3.3%) and mixed infection (1.2%) (overall trypanosome infection, chi2=35.5, d.f.=1, P<0.05; T. brucei infection, chi2=8.06, d.f.=1, P<0.05; T. congolense infection, chi2=22.8, d.f.=1, P<0.05 and T. vivax infection, chi2=6.4, d.f.=1, P<0.05). Infections of Trypanosoma congolense were predominant in cattle in villages with G. pallidipes, while T. vivax infections were predominant in cattle in villages without. In all villages, T. brucei infections were fewer than either T. congolense or T. vivax infections. The risk of transmission of T. brucei, T. congolense and T. vivax infections was 3, 2.7 and 1.6 times, respectively, higher in villages with G. pallidipes than in those without, despite the presence of G. f. fuscipes in either set of villages. The mean PCV (28.27+/-0.41, 95% CI) and mean herd size (3+/-0.46) of cattle in villages with G. pallidipes were significantly (P<0.05) lower than in those in villages without (mean PCV, 29.48+/-0.34; mean herd size, 4+/-0.72). It is evident that presence of G. pallidipes brings about an increase in the prevalence of T. congolense, which causes a more severe disease in cattle than other species of trypanosomes. This is a rare case of a re-invasion of a tsetse species whose disease transmission capability calls for refocusing of the traditional national tsetse and trypanosomosis control strategies to contain it.

Spatial and temporal risk factors for the early detection of Trypanosoma brucei rhodesiense sleeping sickness patients in Tororo and Busia districts, Uganda.

We have carried out a study of risk factors for early detection of Trypanosoma brucei rhodesiense sleeping sickness. Records of sleeping sickness patients from 1987 to 2001 from Tororo and Busia districts in Uganda were reviewed for their village of origin and clinical stage (early or late). All villages that reported sleeping sickness and fixed post-diagnostic sleeping sickness health units in Tororo and Busia districts were geo-referenced. The spatial distribution of early and late stage patient detection by health units was analysed using Geographical Information Systems (GIS). Of 1316 sleeping sickness patients admitted at the Livestock Health Research Institute and Busolwe hospitals and Lumino health centre from Tororo and Busia districts, 471 (35.8%) were early stage, 825 (62.7%) were late stage, while 20 (1.5%) were not staged. Five hundred and eighty-five (44.5%) came from within a 10 km radius of the reporting health units. After multivariate analysis, the proportion of early stage patients detected was found to be significantly associated with patients originating from within a 10 km radius of the health unit (P < 0.01), with adults (>19 years) (P < 0.01), and with annual parish incidence (P < 0.01). Application of GIS and the early to late stages ratio are an informative and powerful means of determining efficiency of surveillance of sleeping sickness.

Assessing the patterns of health-seeking behaviour and awareness among sleeping-sickness patients in eastern Uganda.

For those with sleeping sickness, the consequences of delayed diagnosis include poor prognosis at treatment and an increased risk of tsetse infection. Data on their socio-demographic and clinical characteristics, health-seeking behaviour and delays in presentation and diagnosis were collected from 119 diagnosed cases of rhodesiense sleeping sickness in eastern Uganda. The median total delay, from onset of the illness to diagnosis, was 60 days. The median service-provider delay (30 days) was markedly longer than the median patient delay (17 days). Each of these delays was, however, considerable and independently associated with patients presenting with late-stage sleepiness, giving odds ratios and (95% confidence intervals) of 7.29 (3.10-17.14) and 2.98 (1.38-6.43), respectively. A blood examination at the first visit was also associated with the service-provider delay (odds ratio = 0.45; 95% confidence interval = 0.22-0.95). Most of the patients (77.4%) had either been referred to the local sleeping-sickness hospital by other members of their community or presented at the hospital on their own initiative; few had been referred by other components of the local health system. The results are disappointing, not only in showing long delays in diagnosis (and therefore in treatment) but also in indicating that much of the delay is attributable to the service provider failing to diagnose sleeping sickness among symptomatic individuals.

Nitric oxide and cytokine synthesis in human African trypanosomiasis.

Plasma and cerebrospinal fluid (CSF) concentrations of nitrate and the cytokines interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha, interleukin (IL)-10, and IL-4 were measured in 91 African trypanosomiasis patients before and after treatment. Nitrate levels overall were not significantly elevated over those for control persons, but a marginal increase in plasma nitrate was detected in patients reporting illness of <40 days' duration. Plasma IFN-gamma and total TNF-alpha concentrations increased during infection, but free TNF-alpha levels were low in all patients. The most dramatic cytokine response was for IL-10, which was significantly elevated in both plasma and CSF during infection but returned to control levels after treatment. The results indicate that human African trypanosomiasis leads to the development of a strong anti-inflammatory cytokine response.

The origins of a new Trypanosoma brucei rhodesiense sleeping sickness outbreak in eastern Uganda.

BACKGROUND: Sleeping sickness, caused by two trypanosome subspecies, Trypanosoma brucei gambiense and Trypanosoma brucei rhodesiense, is a parasitic disease transmitted by the tsetse fly in sub-Saharan Africa. We report on a recent outbreak of T b rhodesiense sleeping sickness outside the established south-east Ugandan focus, in Soroti District where the disease had previously been absent. Soroti District has been the subject of large-scale livestock restocking activities and, because domestic cattle are important reservoirs of T b rhodesiense, we investigated the role of cattle in the origins of the outbreak. METHODS: We identified the origins of cattle entering the outbreak area in the 4 years preceding the outbreak. A matched case-control study was conducted to assess whether the distance of villages from the main market involved with restocking was a risk factor for sleeping sickness. We investigated the spatial clustering of sleeping sickness cases at the start of the outbreak. FINDINGS: Over 50% (1510 of 2796) of cattle traded at the market were reported to have originated from endemic sleeping sickness areas. The case-control study revealed that distance to the cattle market was a highly significant risk factor for sleeping sickness (p<0.001) and that there was a significant clustering of cases (27 of 28) close to the market at the start of the outbreak (p<0.001). As the outbreak progressed, the average distance of cases moved away from the cattle market (0.014 km per day, 95% CI 0.008-0.020 km per day, p<0.001). INTERPRETATIONS: The results are consistent with the disease being introduced by cattle infected with T b rhodesiense imported to the market from the endemic sleeping sickness focus. The subsequent spread of the disease away from the market suggests that sleeping sickness is becoming established in this new focus. Public health measures directed at controlling the infection in the animal reservoir should be considered to prevent the spread of sleeping sickness.

Sleeping sickness: a tale of two diseases.

Sleeping sickness presents clinically as two distinct diseases, reflecting the fact that two very different trypanosomes are responsible. The African Rift separating East and West Africa defines the distribution of the two diseases. In this review, Susan Welburn, Eric Fèvre, Paul Coleman, Martin Odiit and Ian Maudlin discuss the biology and distribution of these two diseases in relation to the evolution of hominids in Africa.

Identification of human-infective trypanosomes in animal reservoir of sleeping sickness in Uganda by means of serum-resistance-associated (SRA) gene.

BACKGROUND: The expansion of sleeping sickness caused by Trypanosoma brucei rhodesiense beyond its traditional focus in southeast Uganda has been linked with large-scale livestock restocking. To assess the risk presented to the human population by domestic livestock, human-infective T b rhodesiense must be distinguished from non-human-infective T brucei brucei, since both parasites can be present in cattle. We investigated the use of a simple genetic marker to characterise parasites collected from cattle in villages within the new sleeping sickness focus in Soroti District, Uganda. METHODS: 70 T brucei sl samples of known human infectivity status collected from human beings and cattle in Tororo District, Uganda, from 1989 to 1991 were screened for the presence of the human-serum-resistance-associated (SRA) gene by conventional PCR. In 2000-01, blood samples from 200 randomly selected cattle in six villages and two markets in Soroti District were screened for T brucei sl parasites by PCR; positive samples were screened for the presence of the SRA gene. FINDINGS: The SRA gene was present in all 29 samples from patients with sleeping sickness in Tororo District. Of the 41 samples collected from cattle at the same time, the SRA gene was present in the eight samples that tested resistant to human serum in vitro, whereas it was absent from all 33 isolates that were sensitive to human serum in vitro. Of the 200 cattle sampled in Soroti District, we estimated that up to 18% (95% CI 12-23) were infected with T b rhodesiense. INTERPRETATION: Detection of the SRA gene could provide the basis for a simple diagnostic test to enable targeted control of T b rhodesiense in the domestic livestock reservoir, thereby reducing the public-health burden of sleeping sickness in east Africa.

Detection of trypanosomes in suspected sleeping sickness patients in Uganda using the polymerase chain reaction.

Diagnosis of sleeping sickness (trypanosomiasis) is difficult because of the fluctuating levels of parasitaemia encountered in patients. In the present study we found that the polymerase chain reaction (PCR) demonstrated trypanosome infection in 20 out of 35 (57.1%) blood samples and in 21 out of 34 (61.7%) cerebrospinal fluid (CSF) samples collected from an area endemic for sleeping sickness in north-west Uganda. A total of 14 blood samples and 13 CSF samples that were positive for trypanosomes by double centrifugation were also positive by PCR, demonstrating good concordance between the two methods. However, 6 (28.6%) of the 21 blood samples that were parasitologically negative were positive by PCR, while 8 (38.0%) out of 21 CSF samples that were negative by double centrifugation were positive by PCR. These 14 negative samples could therefore be from sleeping sickness cases even though a positive PCR test is not evidence for the presence of trypanosomes. Furthermore, of these 8 CSF samples, 4 had been designated as early cases, based on the absence of trypanosomes and on a count of < or = 5 white blood cells (WBC) per microliter. This suggests that some late-stage cases could potentially be missed according to the present criteria, and it is therefore important to perform clinical trials to determine whether these cases could be treated successfully with the first-stage drug alone. The remaining four CSF samples had been classified as late-stage cases, based on a count of > 6 WBC per microliter, even though trypanosomes could not be detected in these samples by either double centrifugation or PCR. A cut-off point of 5 WBC per microliter, which is used as a rule of thumb to stage sleeping sickness patients, seems to leave some late-stage cases undetected since trypanosomes were detected in four CSF samples from suspected cases with < 5 WBC per microliter.

PIP: This study evaluates the effectiveness of polymerase chain reaction (PCR) in detecting trypanosomes in the blood and cerebrospinal fluid (CSF) of suspected sleeping sickness patients in Uganda. A total of 35 blood samples and 34 CSF samples were analyzed. Trypanosomes were detected in 20 of 35 (57.1%) blood samples, and in 21 of 34 (61.7%) CSF samples by PCR. However, 6 (28.6%) of the 20 blood samples that were parasitologically negative were positive by PCR, while 8 (38.0%) out of 21 CSF samples that were negative by double centrifugation were positive by PCR. These 14 negative samples could therefore be from sleeping sickness cases even though a positive PCR test is not an evidence for the presence of trypanosomes. Furthermore, of these 8 CSF samples, 4 had been designated as early cases, based on the absence of trypanosomes and on a count of 5 or fewer white blood cells (WBCs) per microliter. The remaining 4 CSF samples had been classified as late-stage cases, based on a count of 6 WBCs per microliter, even though trypanosomes could not be detected in these samples by either double centrifugation or PCR. Usually, 5 WBCs per microliter is considered to be the cut-off point in the staging and treatment of sleeping sickness patients. In conclusion, it is imperative to carry out a detailed clinical study on the use of PCR for trypanosomiasis diagnosis and staging of patients in order to demonstrate the relation between PCR and the outcome of treatment.

Trypanosomosis agglutination card test for Trypanosoma brucei rhodesiense sleeping sickness.

OBJECTIVE: To develop a simple field test for diagnosis of Trypanosoma brucei rhodesiense in man. DESIGN: Trypanosomosis Agglutination Card Test (TACT) was developed for the diagnosis of sleeping sickness due to Trypanosoma brucei rhodesiense infection, based on stabilised procyclic forms derived from Utat 4.1. Procyclics were fixed in buffered formalin at 4 degrees for 24 hours and further stabilised in acid/alcohol mixture for 30 minutes. The fixed antigen was stained with Coomassie blue and suspended in 0.1 M PBS/sodium azide buffer pH 7.2 at a concentration of 1 x 10(8) trypanosomes/ml and kept at room temperature. This antigen was used to screen 100 sera from rabbits infected with T. b. rhodesiense, eight from normal rabbits, and 220 only sera 60 of which were from sleeping sickness patients, 50 from normal persons and 110 from other parasitic infections. SETTING: Laboratory testing of the antigen types against the rabbit and human sera infected with cloned variable antigen types of T. b. rhodesiense, was routinely carried on test cards under room temperature. SUBJECTS/PARTICIPANTS: Serum samples from normal and infected rabbits and human subjects. RESULTS: All sera from infected rabbits and 59 from sleeping sickness patients reacted strongly with the antigen showing agglutination reaction which ranged from 1:4 to 1:1024 serum dilution. There was minimal cross reaction with other parasitic infections as follows: one out of 20 malaria patients none of the 20 hookworm patients, one out of 30 for schistosomiasis patients, none of the 10 amoebiasis patients and one out of 20 for filariasis patients. Agglutination titres from all these non-sleeping sickness patients were below 1:16. Based on rabbit positive and negative sera, TACT gave a sensitivity and specificity of 100% and 80% while for human sera a sensitivity of 98.3% and specificity of 96% were observed. CONCLUSION: These preliminary results show that TACT could be a promising screening field test for T. b. rhodesiense sleeping sickness.

Evidence for the occurrence of Trypanosoma brucei rhodesiense sleeping sickness outside the traditional focus in south-eastern Uganda.

The occurrence of Trypanosoma brucei rhodesiense west of the River Nile, in Masindi district in the mid-western part of Uganda, is confirmed. Masindi borders the traditional belt of T. b. gambiense infection in the north-west, Gulu in the north and the Democratic Republic of Congo in the west. Of the 702 persons tested for sleeping sickness in Masindi, 113 (16%) were positive by the card agglutination test for trypanosomiasis (CATT). Trypanosomes were observed in samples of cerebrospinal fluid (CSF) from two (0.3%) of the subjects: a 7-year-old girl, who had been ill for 2 weeks and yet was in good general condition, with three white blood cells (WBC)/microliter CSF; and a 47-year-old woman who had been ill for 8 months, looked sickly, had seven WBC/microliter CSF, but was still able to dig in her gardens. Rats and mice inoculated with blood from the two parasitologically confirmed cases became parasitaemic on day 3 post-inoculation, indicating that the parasites were T. b. rhodesiense. Isoenzyme analysis revealed that the parasites isolated from one of these confirmed cases belonged to a zymodeme (449) which has not been previously observed among isolates from south-eastern or north-western Uganda. Although the isolate shared PGM2 and ICD3 patterns with T. b. gambiense and T. b. rhodesiense, respectively, it did not have the SOD3:5 pattern characteristic of T. b. gambiense. The spread of T. b. rhodesiense beyond its traditional focus and the development of areas where this subspecies and T. b. gambiense are co-endemic will complicate the control of sleeping sickness in Uganda; although the CATT is very useful for the mass screening of populations for T. b. gambiense area, it is not applicable in the detection of T. b. rhodesiense.

Duration of symptoms and case fatality of sleeping sickness caused by Trypanosoma brucei rhodesiense in Tororo, Uganda.

Although there have been recent molecular biological studies for evidence of possible changes in trypanosome biochemistry, such studies are not yet complemented by parallel clinical studies to determine the possible implications to the sleeping sickness patient. The study of the duration of symptoms and the case fatality of T. b. rhodesiense showed that the disease progressed to the stage of central nervous system involvement between three weeks to two months of infection. Most (> 80%) deaths occurred within six months of illness. The case fatality rate of treated sleeping sickness patients was 6% of which the rate in the late-stage of sleeping sickness was more than two and a half times that in the early stage. The incidence of melarsoprol encephalopathy was 2.5% and case fatality due to this condition was 1.0% and similar to previous findings. Thus it appears the virulence of T. b. rhodesiense circulating in south east Uganda has not changed during the past decades.

Isoenzyme comparison of Trypanozoon isolates from two sleeping sickness areas of south-eastern Uganda.

The study characterized 151 Trypanozoon isolates from south-east Uganda by isoenzyme electrophoresis. Stocks were from a range of hosts, including man, cattle, pigs, dogs and Glossina fuscipes fuscipes: 104 isolates were from the Busoga area, 47 were from the Tororo district. Stocks were characterized on thin layer starch gel using eight enzyme systems: ALAT, ASAT, ICD, MDH, ME, NHD, NHI, PGM. Enzyme profiles were generally typical of East Africa; new patterns for ICD and ME were detected. Trypanosomes were classified on the basis of their profile by similarity coefficient analysis and the unweighted pair-group method using arithmetic averages (UPGMA). The majority of trypanosomes were classified in one or other of two genetically distinct groups which corresponded to the strain groups busoga and zambezi, both of which are associated with Rhodesian sleeping sickness in East Africa. Contingency table analyses indicated associations between certain isoenzymes of ICD and PGM, according to host and geographical origin. Significant relationships between trypanosome strain group and geographic origin were also demonstrated for some host groups.

Multicentre evaluation of an antigen-detection ELISA for the diagnosis of Trypanosoma brucei rhodesiense sleeping sickness.

The performance of an enzyme-linked immunosorbent assay (antigen ELISA) for the detection, in serum or cerebrospinal fluid, of an invariant trypanosome antigen to diagnose Trypanosoma brucei rhodesiense sleeping sickness was evaluated in four clinical treatment centres. The test, which was carried out in polystyrene test-tubes, was positive in 88 (88.9%) of 99 parasitologically confirmed cases that were tested at the National Institute for Medical Research, Tabora, United Republic of Tanzania; 99 (94.3%) of 105 cases tested at the National Sleeping Sickness Control Programme, Jinja, Uganda; 86 (87.8%) of 98 cases tested at the Uganda Trypanosomiasis Research Organisation, Tororo, Uganda; and 59 (96.7%) of 61 cases tested at the Tropical Diseases Research Centre, Ndola, Zambia. The overall detection rate was 91.5%. There was no cross-reactivity with the agents of the common bacterial, viral, or parasitic diseases prevalent in the areas where the studies were conducted. The only false-positive result involved a blood donor from a trypanosomiasis endemic focus. The test was simple to perform, was read visually, and is therefore a potential tool for diagnosing human African trypanosomiasis.

Characterization by isoenzyme electrophoresis of Trypanozoon stocks from sleeping sickness endemic areas of south-east Uganda.

An epidemic of sleeping sickness, which started in 1976 in a focus within the county of Luuka in Central Busoga, has spread to cover the three districts of Busoga and large parts of the neighbouring districts of Tororo and Mukono. Forty-three isolates of the subgenus Trypanozoon from Busoga and Tororo (27 from man, 9 from cows, 2 from pigs and 5 from tsetse flies) were compared by thin-layer starch-gel electrophoresis for seven enzymes. Thirty zymodemes were identified; 17 of them were found circulating in the human population. The zymodemes seen previously in Busoga were still circulating together with several new ones. Of the 16 isolates from cattle, pigs and tsetse flies, only two had the same profile, indicating a high degree of diversity. Two zymodemes from cows and a pig were identical to those found in man, implicating domestic stock in the transmission of human disease in south-east Uganda. A computer analysis of the results produced six main zymodeme groups. One comprised only isolates from man; two were composed of isolates from man, domestic animals and tsetse; and three consisted of stocks from domestic animals only. These groups quite probably indicate the different cycles of transmission involving man, tsetse fly and domestic stock.