Molecular detection of Sodalis glossinidius, Spiroplasma species and Wolbachia endosymbionts in wild population of tsetse flies collected in Cameroon, Chad and Nigeria.

BACKGROUND: Tsetse flies are cyclical vectors of African trypanosomiasis (AT). The flies have established symbiotic associations with different bacteria that influence certain aspects of their physiology. Vector competence of tsetse flies for different trypanosome species is highly variable and is suggested to be affected by bacterial endosymbionts amongst other factors. Symbiotic interactions may provide an avenue for AT control. The current study provided prevalence of three tsetse symbionts in Glossina species from Cameroon, Chad and Nigeria. RESULTS: Tsetse flies were collected and dissected from five different locations. DNA was extracted and polymerase chain reaction used to detect presence of Sodalis glossinidius, Spiroplasma species and Wolbachia endosymbionts, using species specific primers. A total of 848 tsetse samples were analysed: Glossina morsitans submorsitans (47.52%), Glossina palpalis palpalis (37.26%), Glossina fuscipes fuscipes (9.08%) and Glossina tachinoides (6.13%). Only 95 (11.20%) were infected with at least one of the three symbionts. Among infected flies, six (6.31%) had Wolbachia and Spiroplasma mixed infection. The overall symbiont prevalence was 0.88, 3.66 and 11.00% respectively, for Sodalis glossinidius, Spiroplasma species and Wolbachia endosymbionts. Prevalence varied between countries and tsetse fly species. Neither Spiroplasma species nor S. glossinidius were detected in samples from Cameroon and Nigeria respectively. CONCLUSION: The present study revealed, for the first time, presence of Spiroplasma species infections in tsetse fly populations in Chad and Nigeria. These findings provide useful information on repertoire of bacterial flora of tsetse flies and incite more investigations to understand their implication in the vector competence of tsetse flies.

Mitigation of Trypanosoma congolense-Associated Anemia and Expression of Trans-sialidase (TconTS) Gene Variants by Eugenol.

PURPOSE: African Animal Trypanosomosis (AAT) caused by Trypanosoma congolense is a parasitic disease affecting the livestock industry in sub-Saharan Africa and usually results in severe anemia, organ damage, and ultimately the death of the infected host. The present study was designed to investigate the possible chemotherapeutic effect of eugenol on T. congolense infections and its inhibitory effect on the trans-sialidase (TconTS) gene expression. METHODS: Animals were infected with T. congolense and treated with 15 and 30 mg/kg body weight (BW) of eugenol for ten (10) days. RESULTS: The eugenol (15 mg/kg BW) significantly (P < 0.05) reduced the T. congolense proliferation, increased animal survival, and reduced serum urea level. However, both dosages of eugenol significantly (P < 0.05) ameliorated T. congolense-induced anemia, renal hypertrophy, splenomegaly, and reduced total damage score in the liver and kidney of infected animals. In addition, the compound significantly (P < 0.05) downregulated the expression levels of TconTS1, TconTS2, TconTS3, and TconTS4 but the effect was more pronounced (sevenfold reduction) on TconTS1. CONCLUSIONS: The oral administration of eugenol suppressed T. congolense proliferation and prevented some major pathologies associated with trypanosomiasis infection. The reversal of renal hypertrophy and splenomegaly by the compound in addition to the reduction in the expression level of the TconTS gene variants could explain the observed anemia ameliorative potential of the compound.

Therapeutic efficacy of ß-sitosterol treatment on Trypanosoma congolense infection, anemia development, and trans-sialidase (TconTS1) gene expression.

Background: African animal trypanosomiasis hinders sustainable livestock productivity in sub-Saharan Africa. About 17 million infected cattle are treated with trypanocides annually but most of the drugs are associated with drawbacks, necessitating the search for a promising chemotherapeutic agent. Objectives: In this study, the effects of ß-sitosterol on Trypanosoma congolense infection were investigated along with its effect on the trans-sialidase gene expressions. Results: Oral treatment with ß-sitosterol at 15 and 30 mg/kg body weight (BW) for 14 days significantly (p < 0.05) reduced parasitemia and ameliorated the parasite-induced anemia. Also, the parasite-induced increase in serum urea level and renal histopathological damage scores in addition to renal hypertrophy was significantly (p < 0.05) reverted following treatment with 30 mg/kg BW ß-sitosterol. The compound also significantly (p < 0.05) down-regulated the expression of TconTS1 but not TconTS2, TconTS3, and TconTS4. Correlation analysis between free serum sialic acid with the TconTS1 and TconTS2 gene variants revealed negative correlations in the ß-sitosterol-treated groups although they were non-significant (p > 0.05) in the group treated with 15 mg/kg BW ß-sitosterol. Similarly, a non-significant negative (p > 0.05) correlation between the biomolecule and the TconTS3 and TconTS4 gene variants was observed in the ß-sitosterol-treated groups while positive correlations were observed in the infected untreated control group. Conclusion: The observed effect of ß-sitosterol on T. congolense infection could make the compound a possible template for the design of novel trypanocides.

Molecular detection of Sodalis glossinidius, Spiroplasma and Wolbachia endosymbionts in wild population of tsetse flies collected in Cameroon, Chad and Nigeria.

Background Tsetse flies are cyclical vectors of African trypanosomiasis. They have established symbiotic associations with different bacteria, which influence certain aspects of their physiology. The vector competence of tsetse flies for different trypanosome species is highly variable and is suggested to be affected by various factors, amongst which are bacterial endosymbionts. Symbiotic interactions may provide an avenue for the disease control. The current study provided the prevalence of 3 tsetse symbionts in Glossina species from Cameroon, Chad and Nigeria. Results Tsetse flies were collected from five different locations and dissected. DNA was extracted and polymerase chain reaction PCR was used to detect the presence of Sodalis glossinidius , Spiroplasma sp and Wolbachia using specific primers. A total of 848 tsetse samples were analysed: Glossina morsitans submorsitans (47.52%), Glossina palpalis palpalis (37.26%), Glossina fuscipes fuscipes (9.08%) and Glossina tachinoides (6.13%). Only 95 (11.20%) were infected with at least one of the 3 symbionts. Among the infected, 6 (6.31%) were carrying mixed infection ( Wolbachia and Spiroplasma ). The overall symbiont prevalence was 0.88%, 3.66% and 11.00% respectively, for Sodalis , Spiroplasma and Wolbachia . Prevalence varied between countries and tsetse species. No Spiroplasma was detected in samples from Cameroon and no Sodalis was found in samples from Nigeria. Conclusion The present study revealed for the first time, the presence of infection by Spiroplasma in tsetse in Chad and Nigeria. These findings provide useful information to the repertoire of bacterial flora of tsetse flies and incite to more investigations to understand their implication in the vector competence of tsetse flies.

Chemotherapeutic potentials of ß-ionone against Trypanosoma congolense infection: Inhibition of parasite proliferation, anemia development, trans-sialidase (TconTS3 and TconTS4) gene expressions, and phospholipase A2.

Trypanosoma congolense is a pathogenic African animal trypanosome species causing devastating conditions leading to death of an infected host. The drawbacks of the existing trypanocidal drugs have led to the search for new drug candidates. In this study, ß-ionone at 15 and 30 mg/kg body weight (BW) was orally administered to T. congolense infected rats for 14 days followed by an assessment of anemia, organ damages, and the expression of T. congolense trans-sialidase gene variants. A significant decrease in parasitemia (p < .05) was observed in the animals treated with 15 mg/kg BW ß-ionone besides increased animal survival rate. A trypanosome-induced decrease in packed cell volume (PCV) and histopathological changes across tissues was significantly (p < .05) ameliorated following treatment with both doses of ß-ionone. This is in addition to reversing the parasite-induced upsurge in free serum sialic acid (FSA) and expression of T. congolense trans-sialidase gene variants (TconTS1, TconTS3, and TconTS4). Correlation analysis revealed a positive correlation (p > .05) between FSA with the TconTS gene expressions. In addition, the compound inhibited partially purified T. congolense sialidase and phospholipase A2 via mixed inhibition pattern with inhibition binding constants of 25.325 and 4.550 µM, respectively, while molecular docking predicted binding energies of -5.6 kcal/mol for both enzymes. In conclusion, treatment with ß-ionone suppressed T. congolense proliferation and protected the animals against some of the parasite-induced pathologies whilst the effect on anemia development might be due to inhibition of sialidase and PLA2 activities as well as the expression levels of TconTS3 and TconTS4.

SARS-CoV-2 seroprevalence at urban and rural sites in Kaduna State, Nigeria, during October/November 2021, immediately prior to detection of the Omicron variant.

BACKGROUND: Nigeria is Africa's most populated country. By November 2021 it had experienced three waves of SARS-CoV-2 infection. Peer-reviewed seroprevalence data assessing the proportion of the Nigerian population that have been infected were extremely limited. METHODS: We conducted a serosurvey in one urban site (n = 400) and one rural site (n = 402) in Kaduna State, Nigeria between 11 October 2021 and 8 November 2021. Z-tests were used to compare seroprevalence across age groups, locations and sexes. T tests were used to determine whether age or household size are associated with seropositivity. Associations between seropositivity and recent history of common Covid-19 symptoms were tested using logistic regression. RESULTS: SARS-CoV-2 antibodies were detected in 42.5% an 53.5% of participants at the urban and rural sites, respectively The overall age- and sex- stratified seroprevalence was 43.7% (42.2% for unvaccinated individuals). The data indicate an infection rate in Kaduna State ≥359-fold the rate derived from polymerase chain reaction-confirmed cases. In the urban site, seroprevalence among females and participants aged <20 was lower than other groups. Reporting loss of sense of taste and/or smell was strongly associated with seropositive status. Associations with seropositivity were also found for the reporting of dry cough, fever, headache, nausea and sore throat. CONCLUSIONS: This study provides baseline SARS-CoV-2 seroprevalence in Kaduna State, Nigeria, immediately prior to the spread of the Omicron variant. It indicates that in October/November 2021, approximately 56% of the population did not have detectable antibodies, and population subgroups with particularly low seroprevalence remain. It highlights limitations in using PCR-confirmed cases to estimate infection rates. The data will inform public health strategies in Nigeria and other sub-Saharan African countries with limited SARS-CoV-2 seroprevalence data.

Single-Nucleotide Polymorphism Associates' ß-Tubulin Isotype-1 Gene in Onchocerca volvulus Populations in Ivermectin-Treated Communities in Taraba State, Nigeria.

PURPOSE: The occurrence of Single-Nucleotide Polymorphisms (SNPs) associated with repeated ivermectin treatment and sub-optimal responses reported by previous findings is of great concern in Onchocerciasis endemic areas. This study investigated SNPs' occurrence after 15 years of ivermectin intervention in Onchocerciasis endemic communities in two Local Government Areas of Taraba State, Nigeria. METHODS: Microfilariae samples were collected by skin snip from individuals treated with ivermectin for 10-15 years of annual distribution and preserved in RNAlater® in a 1.5 ml micro-centrifuge tube. Genomic DNA was extracted from microfilariae and residual skin, amplification in two regions within the ß-tubulin gene, sequenced and analyzed for SNPs using Bioinformatics tools. RESULTS: Three distinct SNP positions: 1183 (T/G), 1188 (T/C) and 1308 (C/T) on the ß-tubulin gene on the targeted 1083-1568 bp fragment, associate's with the ivermectin-treated population. Furthermore, SNPs positions detected in this study are 1730 (A/G) and 1794 (T/G) in the ß-tub gene in the 1557-1857 (bp) region. The 1794 (T/G) SNP position (Phe243Val) in the exon within the ß-tubulin gene region were observed in this study. CONCLUSION: The present study indicates that SNPs are observed in Onchocerca volvulus, thus strengthening the warning that genetic changes could occur in some parasite populations in some ivermectin-treated areas.

Epizootiology and Molecular Identification of Trypanosome Species in Livestock Ruminants in the Gambia.

INTRODUCTION: African Animal Trypanosomiasis (AAT) or nagana in animals, is caused by the blood-borne parasitic protozoa called trypanosomes, and is potentially fatal. It is estimated that Africa loses $4‒5 billion annually due to the death of livestock to nagana in the tsetse belt. PURPOSE: Although The Gambia lies within this belt, there is scanty data regarding the epizootiology of nagana in The Gambia. Here, records of reported cases of nagana for the period 2010-2019 at the International Trypanotolerance Centre (ITC) in The Gambia were analyzed retrospectively. METHODS: For insights into the current prevalence of AAT, blood samples of 384 cattle, 42 goats, and 59 sheep from the Central River Region (CRR) and Lower River Region (LRR) were analyzed microscopically for parasite identification. Furthermore, trypanosomes were characterized by polymerase chain reaction (PCR) using a panel of primers that identify trypanosomes to the level of the species and subspecies by targeting a portion of the internally transcribed spacer-one (ITS-1) of the ribosomal RNA. RESULTS: The retrospective study indicates that Trypanosoma vivax (66%) and T. congolense (33.4%) were the predominant species. Based on the archive records of ITC, the villages Touba, Misera, and Sambel Kunda all in the CRR of the Gambia are the most burdened with AAT. Microscopic examination of blood samples from cattle showed a prevalence of 1.56%, whereas the PCR-based analysis gave a higher prevalence of 12.5%. The molecular analysis revealed the presence of T. vivax (3.65%), T. congolense kilifi (2.6%), T. b. brucei (1.3%), T. congolense savannah/forest (0.52%), T. b. gambiense (0.52%). Interestingly, 4.43% of mixed infections i.e. multiple trypanosome species in individual animals were recorded. In 18% of the mixed infection cases, T. godfreyi, T. simiae were coinfecting cattle alongside T. congolense. The molecular identification including the phylogenetic analysis implicated T. congolense as the most predominant trypanosome species infecting animals in The Gambia. CONCLUSION: The incidence of nagana in The Gambia is documented and the prevalent trypanosomes identified to be T. vivax, different types of T. congolense, and T. brucei including the gambiense subspecie. Finally, nagana is less profound in sheep and goats compared to cattle, with seasonal and regional variations playing a significant role in the disease dynamics.

Molecular identification and prevalence of trypanosomes in cattle distributed within the Jebba axis of the River Niger, Kwara state, Nigeria.

BACKGROUND: Trypanosomiasis is a fatal disease that threatens the economy of at least 37 countries in sub-Saharan Africa, particularly with regard to livestock farming. In this study, we investigated the prevalence of trypanosome infection in cattle, and molecularly identified the species of trypanosomes in infected cattle and the spatial distribution of trypanosome-infected herds along the Jebba axis of the River Niger. METHODS: A randomized cross-sectional study was conducted along the Jebba axis of the River Niger by screening cattle from 36 herd clusters by nested PCR using ITS-1 generic primers. Data generated were analysed using the Chi-square test at a 95% confidence interval. RESULTS: Microscopic examination revealed three infected cattle out of 398 examined, representing 0.8% prevalence. Twelve animals (3.0%) were positive by PCR. Our results showed a decline in the packed cell volume of infected animals (24.7%). The infection rates were categorized as single infection in 11/12 (91.7%) and mixed infection in 1/12 (8.3%). Animals were most frequently infected by Trypanosoma congolense (50.0%), with T. congolense Savannah being the most prevalent subspecies (71.4%). Aside from the infection rate by age (10.0%) and relative distance of animals from the River Niger (56.2%), statistical differences in every other parameter tested were based on mere probabilistic chance. Spatial data showed that the disease was prevalent among herds located less than 3 km from the River Niger. CONCLUSIONS: Six species of trypanosomes were identified in cattle herds along the Jebba axis of the River Niger, with T. congolense being the most prevalent. Age and relative distance of herds from the River Niger may be risk factors for trypanosome infection in cattle herds in this area.

Genetic diversity of Dengue virus serotypes circulating among Aedes mosquitoes in selected regions of northeastern Nigeria.

The flaviviruses are mosquito borne pathogens that continue to pose a considerable public health risk to animals and humans. The members of this group includes, Dengue virus (DENV), Yellow fever virus (YVF), Japanese encephalitis virus (JEV), West Nile virus (WEV) and Zika virus (ZKV). The DENV mosquito vector is endemic to tropical and subtropical climates, placing ∼40% of the world's population at direct risk of dengue infection. Currently, in Nigeria the status of DENV serotypes circulating among mosquito vectors is unknown. Our study was designed to identify and characterize the DENV serotypes circulating in Aedes mosquito populations collected in selected sites in Nigeria. The mosquitoes were collected and identified morphologically to species level using colored identification keys of Rueda. Generally, each species identified was tested in pools of 20 individuals of each Aedes species. RT-PCR and semi nested PCR were used to detect DENV serotypes in mosquitoes and characterized using Sanger sequencing methods. The results showed that DENV serotypes were detected in 58.54% (24/41) of the pools of Aedes mosquitoes from Mubi, Numan and Yola screened. All DENV1-4 serotypes were detected in Ae. aegypti. While DENV 1, 2 and 4 were detected in Ae. albopictus. And only DENV 2 was detected in Ae. galloisi with DENV4 serotype being reported for the first time in Nigeria. DENV2 (37.8%) was the most detected serotypes, while double and triple co-infections of serotypes were detected in 24.4% of the pools. Phylogenetic analysis revealed a strong evolutionary relatedness of DENV serotypes in our study with that of South and Southeast Asia, North America, and other African countries. This is the first reports on the natural DENV serotypes co-infection among Aedes species pools in Nigeria, which can create possible interaction with other flaviviruses causing animal and human diseases. In addition, our study postulates the possible linkage between DENV serotypes infection and human febrile flu-like disease burden being experienced by host communities in northeastern Nigeria.

Genetic diversity of trypanosome species in tsetse flies (Glossina spp.) in Nigeria.

BACKGROUND: Trypanosomes cause disease in humans and livestock in sub-Saharan Africa and rely on tsetse flies as their main insect vector. Nigeria is the most populous country in Africa; however, only limited information about the occurrence and diversity of trypanosomes circulating in the country is available. METHODS: Tsetse flies were collected from five different locations in or adjacent to protected areas, i.e. national parks and game reserves, in Nigeria. Proboscis and gut samples were analysed for trypanosome DNA by molecular amplification of the internal transcribed spacer 1 (ITS1) region and part of the trypanosome specific glycosomal glyceraldehyde-3-phosphate dehydrogenase (gGAPDH) gene. RESULTS: The most abundant Trypanosoma species found in the tsetse gut was T. grayi, a trypanosome infecting crocodiles. It was ubiquitously distributed throughout the country, accounting for over 90% of all cases involving trypanosomes. Trypanosoma congolense was detected in gut samples from all locations except Cross River National Park, but not in the proboscis, while T. brucei (sensu lato) was not detected at all. In proboscis samples, T. vivax was the most prominent. The sequence diversity of gGAPDH suggests that T. vivax and T. grayi represent genetically diverse species clusters. This implies that they are highly dynamic populations. CONCLUSIONS: The prevalence of animal pathogenic trypanosomes throughout Nigeria emphasises the role of protected areas as reservoirs for livestock trypanosomes. The genetic diversity observed within T. vivax and T. grayi populations might be an indication for changing pathogenicity or host range and the origin and consequences of this diversity has to be further investigated.

Clastogenic effects of Trypanosoma brucei brucei and Trypanosoma evansi mixed infection in bone marrow of Wistar rats.

The clastogenic effect of mixed infection of Trypanosoma evansi and Trypanosoma brucei brucei in the bone marrow (BM) cells of Wistar albino rats was investigated. Clastogenic effects were observed in the BM cells using the micronucleus assay. The findings indicate that T. evansi, T. b. brucei and mixed infection with both parasites induced the formation of micronucleated polychromatic erythrocyte (MN-PCEs) in the BM cells significantly (P < 0.05) by 60, 63 and 81 micronuclei/1000 PCE respectively. Mixed infection induced formation of MN-PCEs increase by about 1.33 fold when compared with single infections of T. b. brucei and T. evansi. These data give a preliminary evidence of possible genotoxic effects in trypanosomiasis.

Biochemical diversity in the Trypanosoma congolense trans-sialidase family.

Trans-sialidases are key enzymes in the life cycle of African trypanosomes in both, mammalian host and insect vector and have been associated with the disease trypanosomiasis, namely sleeping sickness and nagana. Besides the previously reported TconTS1, we have identified three additional active trans-sialidases, TconTS2, TconTS3 and TconTS4, and three trans-sialidase like genes in Trypanosoma congolense. At least TconTS1, TconTS2 and TconTS4 are found in the bloodstream of infected animals. We have characterised the enzymatic properties of recombinant proteins expressed in eukaryotic fibroblasts using fetuin as model blood glycoprotein donor substrate. One of the recombinant trans-sialidases, TconTS2, had the highest specific activity reported thus far with very low sialidase activity. The active trans-sialidases share all the amino acids critical for the catalytic reaction with few variations in the predicted binding site for the leaving or acceptor glycan. However, these differences cannot explain the orders of magnitudes between their transfer activities, which must be due to other unidentified structural features of the proteins or substrates selectivity. Interestingly, the phylogenetic relationships between the lectin domains correlate with their specific trans-sialylation activities. This raises the question whether and how the lectin domains regulate the trans-sialidase reaction. The identification and enzymatic characterisation of the trans-sialidase family in T. congolense will contribute significantly towards the understanding of the roles of these enzymes in the pathogenesis of Animal African Trypanosomiasis.