Comparative pathogenicity of single and mixed drug-resistant Trypanosoma brucei brucei and Trypanosoma congolense infections in rats.

Drug-resistant trypanosomes are widespread in sub-Saharan Africa and in conjunction with the drug-sensitive phenotypes cause a serious endemic wasting disease in animals. We evaluated the pathogenicity of single and mixed drug-resistant Trypanosoma brucei brucei and T. congolense isolates in 35 female rats, randomly divided into seven groups (1-7) of five rats. Group 1 was the uninfected control. Groups 2 and 3 were infected with drug-sensitive T. brucei brucei and T. congolense, respectively, whereas groups 4 and 5 were infected with multidrug-resistant T. brucei brucei and T. congolense respectively. Group 6 were infected with drug-sensitive T. brucei brucei and T. congolense while group 7 were infected with multidrug-resistant T. brucei brucei and T. congolense. Parasitaemia kinetics, haematological parameters, body weight, clinical signs, survival time, gross and histopathological changes in the spleen were evaluated. Parasitaemia occurred between day 3-9 post-infection in all the infected groups. Rats in groups 4 and 7 had markedly prolonged (p < 0.05) pre-patent period, days to first peak parasitaemia, survival time, and lower (p < 0.05) parasitaemia level than groups 2 and 6 rats while these parameters were comparable for groups 3 and 5 rats. Anaemia was noted in the infected groups but the severity did not vary amongst the infected groups. Severe clinical signs and splenic lesions were noted in rats infected with drug-sensitive trypanosome species compared to the multidrug-resistant species. Therefore, we conclude that the trypanosome isolates were pathogenic. However, the drug-sensitive T. brucei brucei and mixed drug-sensitive trypanosome infections were more pathogenic than their multidrug-resistant counterparts.

Comparative pathogenicity of drug-resistant and drug-sensitive Trypanosoma brucei and Trypanosoma congolense infections in Nigerian local dogs.

Animal trypanosomosis is an important endemic and wasting disease in sub-Saharan Africa. Its control relies on chemotherapy, and resistance to trypanocides has been widely reported. The pathogenicity of drug-resistant canine trypanosomes is not clear with scanty information available. Thus, this study assessed the comparative pathogenicity of drug-resistant and drug-sensitive Trypanosoma brucei and Trypanosoma congolense infections in dogs. Twenty Nigerian local dogs were used and were randomly assigned into five groups (A-E) of four dogs each. Group A served as the uninfected-control group, while groups B and C were infected with 106 drug-sensitive T. congolense and T. brucei. Groups D and E were infected with 106 multidrug-resistant T. congolense and T. brucei, respectively. The pre-patent period (PPP), clinical signs, level of parasitaemia (LOP), rectal temperature, body weight, packed cell volume (PCV), red blood cell count (RBC), haemoglobin concentration (HbC), mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC), total leucocyte count (TLC) and survivability were assessed. Groups D and E had longer (p < 0.05) mean PPP than groups B and C. Also, group E dogs had lower (p < 0.05) mean LOP, longer (p < 0.05) mean survivability, and higher (p < 0.05) mean body weight, PCV, HbC and RBC than group C dogs. The clinical signs were very severe in group C dogs, compared to group E dogs. However, these parameters did not differ statistically between groups B and D. Thus, multidrug-resistant T. brucei was of lower pathogenicity than drug-sensitive T. brucei, while multidrug-resistant and drug-sensitive T. congolense had comparable pathogenicity following infection in dogs.

Economic and feasibility comparison of the dRIT and DFA for decentralized rabies diagnosis in resource-limited settings: The use of Nigerian dog meat markets as a case study.

BACKGROUND: Rabies lyssavirus (RABV) is the aetiologic agent of rabies, a disease that is severely underreported in Nigeria as well as elsewhere in Africa and Asia. Despite the role that rabies diagnosis plays towards elucidating the true burden of the disease, Nigeria-a country of 180 million inhabitants-has a limited number of diagnostic facilities. In this study, we sought to investigate two of the World Organization for Animal Health (OIE)-recommended diagnostic assays for rabies-viz; the direct fluorescent antibody test (DFA) and the direct rapid immunohistochemical test (dRIT) in terms of their relative suitability in resource-limited settings. Our primary considerations were (1) the financial feasibility for implementation and (2) the diagnostic efficacy. As a case study, we used suspect rabies samples from dog meat markets in Nigeria. METHODS/PRINCIPAL FINDINGS: By developing a simple simulation framework, we suggested that the assay with the lowest cost to implement and routinely use was the dRIT assay. The costs associated with the dRIT were lower in all simulated scenarios, irrespective of the number of samples tested per year. In addition to the cost analysis, the diagnostic efficacies of the two assays were evaluated. To do this, a cohort of DFA-positive and -negative samples collected from dog meat markets in Nigeria were initially diagnosed using the DFA in Nigeria and subsequently sent to South Africa for diagnostic confirmation. In South Africa, all the specimens were re-tested with the DFA, the dRIT and a quantitative real-time polymerase chain reaction (qRT-PCR). In our investigation, discrepancies were observed between the three diagnostic assays; with the incongruent results being resolved by means of confirmatory testing using the heminested reverse transcription polymerase reaction and sequencing to confirm that they were not contamination. CONCLUSIONS/SIGNIFICANCE: The data obtained from this study suggested that the dRIT was not only an effective diagnostic assay that could be used to routinely diagnose rabies, but that the assay was also the most cost-effective option among all of the OIE recommended methods. In addition, the results of our investigation confirmed that some of the dogs slaughtered in dog markets were rabies-positive and that the markets posed a potential public health threat. Lastly, our data showed that the DFA, although regarded as the gold standard test for rabies, has some limitations-particularly at low antigen levels. Based on the results reported here and the current challenges faced in Nigeria, we believe that the dRIT assay would be the most suitable laboratory test for decentralized or confirmatory rabies diagnosis in Nigeria, given its relative speed, accuracy, cost and ease of use.

Molecular Detection of Rabies Lyssaviruses from Dogs in Southeastern Nigeria: Evidence of TransboundaryTransmission of Rabies in West Africa.

Despite being the first country to register confirmed cases of Mokola and Lagos bat lyssaviruses (two very distant lyssaviruses), knowledge gaps, particularly on the molecular epidemiology of lyssaviruses, still exist in Nigeria. A total of 278 specimens were collected from dogs in southeastern Nigeria between October 2015 and July 2016, and 23 (8.3%) of these tested positive for lyssaviruses with the direct fluorescent antibody test (DFA). The lyssaviruses were genetically characterized by amplifying the highly conserved nucleoprotein (N) gene of the rabies lyssaviruses (RABVs) of the viral genome. Phylogenetic analyses of the nucleotide sequences showed that all the RABV sequences in this study were of the Africa-2 lineage. Our results demonstrated that transboundary transmission of rabies lyssavirus is a key event, given that one of the RABV sequences (MN196576) clustered with rabies variants from neighboring Niger Republic. Furthermore, three RABVs from dogs from Anambra State clustered separately forming a novel and distinct group. Our results demonstrated that transboundary transmission of RABLVs is a key driver in the spread of rabies in West Africa. In order for the successful control of this zoonotic disease, a multinational stepwise surveillance and elimination of rabies in Africa by 2030 is probably the solution for regional elimination.

Evaluation of antitrypanosomal activity of Pterocarpus santalinoides L'H'erit ex DC hydroethanol leaf extract in rats experimentally infected with Trypanosoma brucei.

ETHNOPHARMACOLOGICAL RELEVANCE: The leaves of Pterocarpus santalinoides are used alone or with other plants by traditional healers in some Southern Nigerian and Ivorian rural villages to treat blood parasitic infections including trypanosomiasis and malaria. However, their efficacy and safety remains doubtful. AIM: To evaluate the antitrypanosomal activity of Pterocarpus santalinoides hydroethanol leaf extract (PSELE) in vitro and in vivo. MATERIALS AND METHODS: The phytochemical and acute toxicity studies of PSELE were performed using standard methods. Eleven different concentrations of the extract were screened for in vitro antitrypanosomal activity. For the in vivo study, twenty-five female albino rats were randomly assigned into five groups of five rats each. Group A was the uninfected and untreated group while groups B - E were infected intraperitoneally with 106 trypanosomes. Group C rats were treated once on day 11 post infection (PI) with 7 mg/kg diminazene aceturate while groups D and E rats were also treated on same day with 200 mg/kg and 400 mg/kg PSELE respectively for seven days. The level of parasitaemia (LOP), survivability, packed cell volume (PCV), total leucocyte count (TLC), total erythrocyte count (TEC), body weight and rectal temperature were used to assess the antitrypanosomal effect of PSELE. RESULTS: Phytochemical analysis revealed the presence of flavonoids, tannins, carbohydrates and reducing sugar. No sign of toxicity or mortality was observed following acute toxicity study at a single dose of 2000 mg/kg. The LC50 of PSELE was 0.0625 mg/ml. Parasitaemia was first detected on day 8 and all infected rats became parasitaemic on day 10 (PI). PSELE significantly reduced (p < 0.05) LOP, improved PCV, TEC and prolonged survival time of the rats compared with the infected untreated group B. CONCLUSION: It was therefore concluded that PSELE is safe, possesses some antitrypanosomal activity and may serve as a lead for the development of an effective alternative antitrypanosomal drug.

Detection of lyssavirus antigen and antibody levels among apparently healthy and suspected rabid dogs in South-Eastern Nigeria.

OBJECTIVES: Domestic dogs are the main reservoir of rabies virus (RABV) infection in Nigeria, thus surveillance of rabies in dog populations is crucial in order to understand the patterns of spread of infection and ultimately devise an appropriate rabies control strategy. This study determined the presence of lyssavirus antigen in brain tissues and anti-rabies antibodies in sera of apparently healthy and suspected-rabid dogs slaughtered for human consumption at local markets in South-Eastern Nigeria. RESULTS: Our findings demonstrated that 8.3% (n = 23) of brain tissues were lyssavirus positive and 2.5% (n = 25) of sera had rabies antibody levels as percentage blocking of 70% and above correlating with a cut-off value ≥ 0.5 IU/mL in the fluorescent antibody neutralization test. There was an inverse correlation between lyssavirus positivity and rabies antibody levels confirming that infected individuals most often do not develop virus neutralizing antibodies to the disease. The low percentage of rabies antibodies in this dog population suggests a susceptible population at high risk to RABV infection. These findings highlight a huge challenge to national rabies programs and subsequent elimination of the disease from Nigeria, considering that majority of dogs are confined to rural communal areas, where parenteral dog vaccination is not routinely undertaken.

Comparative haematological study of single and mixed infections of mongrel dogs with Trypanosoma congolense and Trypanosoma brucei brucei.

The haematological effects of single and mixed infections of Trypanosoma congolense and Trypanosoma brucei brucei were compared in experimentally infected mongrel dogs. Twenty mongrel dogs of both sexes aged between 3 and 6 months, and weighing between 2.5 and 5.9 kg were used for the study. The dogs were kept in clean metal cages in a fly-proof house and were adequately fed and given water ad libitum. The twenty dogs were divided into four groups of five dogs each. Group I dogs were uninfected control, group II dogs were infected with T. congolense, group III dogs were infected with T. brucei brucei and group IV dogs were infected with both T. congolense and T. brucei brucei. Parasitaemia occurred in the infected dogs in groups II, III, and IV; 10-13 days post-infection (PI) with the mean pre-patent period (PPP) of 12, 10, and 11 days respectively. Mixed infection persisted throughout the duration of the experiment. T. brucei predominated T. congolense in the mixed infection constituting about 70% of the trypanosomes. The significant (P<0.05) decrease in the mean haemoglobin concentration (Hb) and packed cell volume (PCV) caused by the infection did not differ significantly (P>0.05) between the infected groups. Also the significant (P<0.05) reduction in the total white blood cell count (TWBC) caused by the infection did not differ significantly (P>0.05) between the infected groups. The decline in the total WBC count was due primarily to significant (P<0.05) reduction in the lymphocyte counts of the infected dogs. It was thus concluded that single or mixed infection of mongrel dogs with T. congolense and T. brucei brucei resulted in anaemia and leucopenia which did not differ significantly (P>0.05) among the infected groups.