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In Egypt, camel trypanosomiasis is widespread. From October 2021 to March 2022, we collected 181 blood samples from apparently healthy one-humped camels (Camelus dromedarius) in Cairo and Giza Governates. The objective of this study was to assess infection rates of trypanosomes using blood smear examination and PCR-sequencing assays. Trypanosomes were detected in 8.3% (15/181) of camels by blood smear and in 23.8% (43/181) by PCR targeting the internal transcribed spacer (ITS). Based on blood smear and ITS-PCR results, and the absence of tsetse flies in the study area, we hypothesized that the Trypanosoma species was likely T. evansi. Validation using PCR based on the variant surface glycoprotein (VSG) of T. evansi Rode Trypanozoon antigen type (RoTat) 1.2 (RoTat 1.2 VSG gene) on ITS-PCR-positive samples (n=43) confirmed that 88.4% (38/43) were RoTat 1.2 T. evansi, while 11.6% (5/43) were non-RoTat 1.2 T. evansi. This marks the second report of non-RoTat 1.2 T. evansi in dromedary camels in Egypt. Considering the underestimated zoonotic risk of T. evansi in Egypt, there is a potential threat to humans, underscoring the need for a "One Health" approach to safeguard animal and human health.

Background: Vector-borne pathogens (VBPs) are increasing in significance in veterinary medicine and public health settings, with wildlife playing a potentially crucial role in their transmission. Eurasian badgers (Meles meles) are widely distributed across Europe. However, information currently available on the prevalence of VBPs in badgers is limited. The objective of the current study was to investigate the occurrence of Anaplasmataceae, Bartonella spp., Mycoplasma spp., Rickettsia spp., Piroplasmida, Trypanosomatida and Filarioidea in badgers and subsequently, based on the results, assess the potential risk to domestic animals, other wildlife and humans., Methods: Between 2017 and 2021, blood or spleen samples from 220 badgers were collected in nine continental European countries: Austria (n = 7), Bosnia and Herzegovina (n = 2), Croatia (n = 22), France (n = 44), Germany (n = 16), Hungary (n = 7), Italy (n = 16), Romania (n = 80) and Serbia (n = 26). VBPs were identified by performing PCR analysis on the samples, followed by Sanger sequencing. Additionally, to distinguish between different Babesia lineages we performed restriction fragment length polymorphism (RFLP) analysis on piroplasm-positive samples, using HinfI as restriction enzyme. A phylogenetic analysis was performed on Mycoplasma spp., Results: The pathogens identified were Babesia sp. badger type A (54%), B (23%), and C (37%); Trypanosoma pestanai (56%); Mycoplasma sp. (34%); Candidatus Mycoplasma haematomelis (8%); Candidatus Mycoplasma haematominutum (0.5%); and Ehrlichia spp. (2%). Rickettsia spp., Bartonella spp. and filarioid nematodes were not detected among the tested samples., Conclusions: The large sample size and diverse study populations in this study provide valuable insights into the distribution and epidemiology of the analyzed pathogens. Some of the VBPs identified in our study show high similarity to those found in domestic animals, such as dogs. This finding suggests that badgers, as potential reservoirs for these pathogens, may pose a threat not only to other wildlife but also to domestic animals in close vicinity. Continuous surveillance is essential to monitor VBPs in wildlife as a means to enable the assessment of their impact on other wildlife species, domestic animals and human health., (© 2024. The Author(s).)

The increasing population of dogs and changes in the climatic conditions have resulted in the emergence and re-emergence of vector-borne diseases in canines. These vectors borne diseases in canines pose a diagnostic challenge to the field veterinarians because of co-infections with several pathogens. Comprehensive data on the prevalence of haemoparasites and haemorickettsiales in pet and community owned dogs from south India are scant. Hence, the present study aims to find and compare the prevalence of these infections in the pet and the community owned dogs of Kerala, a south Indian state. Two hundred and seventy-two pets and 150 community owned dogs were examined by microscopy and polymerase chain reaction (PCR) for infections with different heamoparasites and haemorickettsials from January 2018-November 2020 in the state of Kerala. A high prevalence of Babesia gibsoni infection (42.2-60.0 %) in pet and community owned dogs, followed by Babesia vogeli (5.8-39.3 %), Hepatozoon canis (0.7-28.0 %), Trypanosoma evansi (0.0-27.3 %), Ehrlichia canis (0.3-0.6 %) and Anaplasma platys (0.0-0.6 %) was observed in the present study. Eighty-eight per cent (132/150) of the community owned dogs and 49.2 % (134/272) of the pet dogs were positive for at least one pathogen. Phylogenetic analysis based on the partial nucleotide sequences of 18S rRNA and TRAP gene of B. gibsoni, 18S rRNA genes of B. vogeli and H. canis and RoTat, 1.2 virB9 and 16S rRNA genes of T. evansi, E. canis and A. platys, respectively was carried out. B. vogeli, H. canis, E. canis and A. platys revealed genetic relatedness between the Indian isolates and the isolates from other countries. However, B. gibsoni isolates from the Indian sub-continent were genetically unique compared to other Asian isolates. The clustering of T. evansi isolates from India in two clades viz., livestock origin (cattle, buffalo) and others indicated their genetic variability. The present study summarizes the prevalence of some of the haemoparasites and haemorickettsials in the dog populations of Kerala (south India) and also determined their genetic relationship with the isolates prevalent in dogs in other countries., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2024. Published by Elsevier B.V.)

Background: Trypanosomiasis continues to pose a global threat to human health, with human infection mainly caused by Trypanosoma brucei and Trypanosoma cruzi., Methods: We present a 30-year-old pregnant woman with persistent high fever from Shandong Province, China. High-throughput sequencing revealed the presence of Trypanosoma dionisii in blood. We conducted an analysis of the patient's clinical, epidemiological, and virological data., Results: The patients exhibited fever, shortness of breath, chest tightness, accompanied by change in liver function and inflammatory response. She made a full recovery without any long-term effects. T. dionisii was detected in blood collected 23 days after onset of illness. The 18S rRNA gene sequence showed close similarity to T. dionisii found in bats from Japan, while the gGAPDH gene was closely related to T. dionisii from bats in Mengyin County, Shandong Province. Phylogenetic analysis demonstrated the current T. dionisii belongs to clade B within its species group. Positive anti-Trypanosoma IgG antibody was detected from the patient on Day 23, 66 and 122 after disease onset, as well as the cord blood and serum from the newborn. Retrospective screening of wild small mammals captured from Shandong Province revealed a prevalence rate of 0.54% (7/1304) for T. dionisii; specifically among 0.81% (5/620) of Apodemus agrarius, and 0.46% (2/438) of Mus musculus., Conclusions: The confirmation of human infection with T. dionisii underscores its potential as a zoonotic pathogen, while the widespread presence of this parasite in rodent and bat species emphasizes the emerging threat it poses to human health., Competing Interests: Declaration of Competing Interest The authors declare no conflicts of interest., (Copyright © 2024. Published by Elsevier Ltd.)

A Trypanosoma screening was conducted on 130 pools comprising 1,241 ticks, collected from 674 selected farm ruminants in Peninsular Malaysia. Of these, nine pools were tested positive for Trypanosoma. Subsequent BLAST searches revealed that the 18S rRNA gene sequences were closely related to Trypanosoma rhipicephalis isolate Chaco CB, with percentage similarities ranging from 95.56 % to 99.84 %. Phylogenetic analysis showed that three of the nine sequences formed a clade with Trypanosoma rhipicephalis. The remaining six Trypanosoma sequences formed a distinct clade, separate from T. rhipicephalis and other Trypanosoma species, with genetic distances of 4.34 % and 4.33-4.58 %, respectively. This study marks the first report of tick-associated Trypanosoma in Malaysia and underscores significant research gaps regarding trypanosome interactions with tick hosts in the region., Competing Interests: Declaration of competing interest All authors declare no competing interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Background: Chagas disease is caused by Trypanosoma cruzi, whose genetic structure is divided into six discrete typing units (DTUs) known as TcI-TcVI. In the Yucatan Peninsula, Mexico, information regarding the DTUs circulating in wild mammals is scarce, while this is important knowledge for our understanding of T. cruzi transmission dynamics., Methods: In the current study, we sampled wild mammals in a sylvatic site of the Yucatan Peninsula and assessed their infection with T. cruzi by PCR. Then, for infected mammals, we amplified and sequenced nuclear and mitochondrial T. cruzi genetic markers for DTU identification., Results: In total, we captured 99 mammals belonging to the orders Chiroptera, Rodentia and Didelphimorphia. The prevalence of infection with T. cruzi was 9% (9/99; 95% CI [5, 16]), and we identified TcI in a Jamaican fruit bat, Artibeus jamaicensis. Moreover, we fortuitously identified Trypanosoma dionisii in another Jamaican fruit bat and detected an unidentified Trypanosoma species in a third specimen. While the latter discoveries were not expected because we used primers designed for T. cruzi, this study is the first to report the identification of T. dionisii in a bat from Yucatan, Mexico, adding to a recent first report of T. dionisii in bats from Veracruz, and first report of this Trypanosoma species in Mexico., Conclusion: Further research is needed to enhance our knowledge of T. cruzi DTUs and Trypanosoma diversity circulating in wildlife in Southeastern Mexico., (© The Author(s) 2024. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene.)

Phlebotomine sand flies are recognized as a primary vector of Leishmania and are also suspected vectors of Trypanosoma. The transmission cycle of these parasites relies on the distribution of sand fly vectors, parasites, and reservoir animals. This study aimed to detect Leishmania and Trypanosoma DNA and identify the sources of bloodmeals in post-feeding sand flies captured across Thailand. A total of 42,911 field female sand flies were collected from 11 provinces across Thailand using CDC light traps. Among these, 253 post-feeding sand flies were selected for analysis. The predominant species in this study was Sergentomyia khawi (33.60 %). The DNA was extracted from individual female sand flies. Polymerase chain reaction (PCR), specific to the internal transcribed spacer 1 (ITS1) and the small subunit ribosomal RNA (SSU rRNA) gene regions were used to detect the presence of Leishmania and Trypanosoma DNA, respectively. Additionally, cytochrome c oxidase subunit I (COI) gene region was utilized to identify the sources of host bloodmeals. Leishmania DNA was not detected in any specimens. The analysis of SSU rRNA sequences revealed the presence of Trypanosoma DNA (11.46 %, 29/253) in sand fly samples. Among these samples, T. noyesi (1.58 %, 4/253) was identified in Idiophlebotomus longiforceps and Phlebotomus asperulus, Trypanosoma Anura01+02/Frog2 (1.18 %, 3/253) in Se. khawi, and Trypanosoma Anura04/Frog1 (8.70 %, 22/253) in Se. khawi, Se. hivernus and Grossomyia indica. Bloodmeal analysis utilizing the COI gene revealed a diverse range of vertebrate hosts' blood, including bird, bat, frog and sun skink. Our findings confirm the presence of Trypanosoma DNA and identify the sources of bloodmeals from vertebrate hosts in various sand fly species, suggesting their potential as possible vectors for Trypanosoma in Thailand. Furthermore, our study is the first to provide molecular evidence using the COI gene to identify frogs as a host blood source for sand flies in Thailand. Further studies focusing on the isolation of live parasites in sand flies to confirm vector potential and examining the role of animal reservoirs will enhance our understanding of the host-parasite relationship and enable more efficient control for disease transmission., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Although several primers targeted to the internal transcribed-spacer 1 (ITS1) of the ribosomal DNA (rDNA) have been designed to improve the detection of African trypanosomes, no study tried to compare their agreement level and ability to amplify different trypanosome species in tsetse flies and mammals in various epidemiological settings. This study was designed to fill this gap, by targeting tsetse-infested areas of Cameroon. For this, archived DNA samples reporting at-least one trypanosome species with species-specific PCR primers were reviewed. Ten sets of primers targeting different ITS1 rDNA sequences of trypanosomes were selected for assessment using single-round and nested-PCR method. Amplification rates (sensitivity) and agreement level of different ITS1 assays were compared using Cohen's-Kappa and McNemar's x 2 statistic. Little agreement level (k = 0.05-0.52) were observed between different ITS1-primers PCRs detection of African trypanosome species despite significant (X 2 =54.3, p = 0.0001) high amplification rate 91.6 % (339/370). This sensitivity varied from quite low for T. simiae (11.9 %) and T. vivax (27.3 %) to fairly good for T. congolence (51.9 %), Trypanozoon (32.4 %) and T. theileri (40.3 %). Primers set targeting ITS1-A sequence of trypanosome species recorded the highest sensitivity (50.5 %) with fairly good agreement compared to 39.2 % for ITS1-C (k = 0.52), 32.4 % for ITS1-R (k = 0.47), 29.7 % for ITS1-N (k = 0.48) and 23.0 % for ITS1-KIN (k = 0.43) respectively. This study revealed a diversity in the sensitivity of different trypanosome species with different sets of ITS-primers enhancing the need to use the same sets of primers in different bio-ecological settings. The use of nested-PCR instead of single-round PCR enabled improvement of trypanosome infections detection in both tsetse and mammals. Among the sets of ITS1-primers tested, those designed by to amplify ITS1-A can be considered as the most appropriate for the detection of trypanosome infections in mammals and tsetse flies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing personal relationships or financial interests that could have appeared to influence the work reported in this paper. Authors also confirm that the manuscript has been read and approved by all named authors. Also that there are no other person who satisfied the criteria for authorship but are no listed. We further confirm that the order of authors listed in the manuscript has been approved by all of us., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Water frogs of the genus Pelophylax host a variety of parasites, from protozoa to helminths. Among the blood parasites, representatives of Apicomplexa, Trypanosoma and Nematoda show the highest prevalence. In this study, we focused on blood parasites of water frogs living in the Danube Delta, Romania. In total, 74 individuals of P. ridibundus and eight individuals of P. esculentus from six localities were examined. Blood parasites were detected microscopically and using a molecular marker (18S rDNA). 89.77% of frogs from all investigated localities were found to be infected with at least one parasitic group, specifically with haemogregarines (84.09%), nematodes (1.14%), and trypanosomes (63.64%). The parasitemia of haemogregarines and trypanosomes differed significantly among the studied locations. There was no statistically significant difference in parasitemia between male and female hosts. However, adults were found to have a significantly higher parasitemia in comparison with subadults infected with haemogregarines. Correlation between parasitemia and the body length of frogs infected with haemogregarines was also significant (r = 0.226). By comparing the 18S rDNA sequences with the corresponding GenBank sequences, Hepatozoon species identified in water frogs showed a close similarity (98.1-99.8%) to Hepatozoon magna. Trypanosomes showed the highest sequence similarity to Trypanosoma sp. isolate R10 clone L2-3, Trypanosoma ranarum, and Trypanosoma cobitis., Competing Interests: Declaration of competing interest All authors declare no conflict of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Background: Trypanosomiasis is an infectious disease caused by parasitic protozoa of the genus Trypanosome and primarily transmitted by tsetse flies. This study aimed to determine the density of tsetse flies and the rate of trypanosome infection in the Bedele and Dabo Hana districts of the Buno Bedele Zone in Ethiopia., Results: A cross-sectional study was conducted from January to February 2023 to catch tsetse flies, determine tsetse density, and estimate the trypanosome infection rate. We used 100 traps (40 NGU, 30 pyramidal, and 30 biconical) to catch the flies. The following standard procedures were followed to identify the specific trypanosome species in the collected tsetse flies: The flies were dissected, and the salivary glands were removed. We placed the salivary glands in a drop of saline solution on a microscope slide. A coverslip was placed over the salivary glands, the slide was examined under a microscope, and the trypanosomes were identified based on their morphology. A total of 3,740 tsetse flies were captured from 100 traps, resulting in an overall apparent density of 18.7 flies per trap per day. Within the study area, only one species of tsetse fly, Glossina tachinoides, was identified. Of the 1,320 dissected Glossina tachinoides, 1.82% were found to be infected with trypanosome parasites. Among these infections, 58.33% were attributed to Trypanosoma congolense, while the remaining 41.67% were caused by Trypanosoma brucei. The infection rate of trypanosomes was significantly higher in female tsetse flies (87.5%) as compared to male flies (12.5%). Furthermore, a significantly higher infection rate was observed in flies older than 20 days (83.33%) and in hunger stage 1 flies (58.33%) compared to hunger stages 2, 3, and 4., Conclusions: This study highlights the necessity of implementing control and suppression measures targeting the vector (tsetse flies) and the parasite (trypanosomes) to effectively manage and prevent pathogenic animal trypanosomiasis., (© 2024. The Author(s).)

Trypanosomatids have achieved significant evolutionary success in parasitizing various groups, yet reptiles remain relatively unexplored. The utilization of advanced molecular tools has revealed an increased richness of trypanosomatids in vertebrate hosts. The aim of this study was to identify the trypanosomatid species infecting Bothrops moojeni and Crotalus durissus kept in captivity from 2000 to 2022. Blood samples were obtained from 106 snakes: 73C. durissus and 33 B. moojeni. Whole blood was collected for hemoculture, blood smears and centrifugated to obtain the blood clot that had its DNA extracted and submitted to Nested PCR (18S rDNA gene) to detect Trypanosomatidae. Positive samples were quantified and submitted to both conventional (Sanger) and next generation sequencing (NGS). Cloning of the amplified PCR product was performed for only one individual of C. durissus. To exclude the possibility of local vector transmission, attempts to capture sandflies were conducted using six CDC-LT type light traps. Molecular diagnosis revealed that 34% of the snakes presented trypanosomatid DNA, 47.94% in C. durissus and 3.9% in B. moojeni. The cloning process generated four colonies identified as a new MOTU named Trypanosomatidae sp. CROT. The presence of DNA of five trypanosomatids (Trypanosoma cruzi TcII/VI, Trypanosoma sp. DID, Trypanosoma cascavelli, Trypanosomatidae sp. CROT, Leishmania infantum and Leishmania sp.) and one free-living kinetoplastid (Neobodo sp.) was revealed through NGS and confirmed by phylogenetic analysis. The haplotypic network divided the T. cascavelli sequences into two groups, 1) marsupials and snakes and 2) exclusive to marsupials. Therefore, the diversity of Kinetoplastea is still underestimated. Snakes have the ability to maintain infection with T. cruzi and L. infantum for up to 20 years and the DNA finding of Neobodo sp. in the blood of a C. durissus suggests that this genus can infect vertebrates., Competing Interests: Declaration of competing interest None., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Sand flies serve as crucial vectors in various medical and veterinary diseases. Sand fly-borne diseases pose a significant public health burden globally, as the causative agents can infect a diverse range of hosts, leading to severe consequences such as leishmaniasis and sand fly fever. Additionally, the widespread use of insecticides for agricultural purposes and mosquito control is not specifically targeted at sand flies, potentially leading to resistance development. We investigated sand fly species, their potential role as vectors of various parasitic agents, and insecticide resistance in the endemic regions of Natawi and Sadao districts in Songkhla, Thailand., Methods: Sand flies were collected using CDC light traps. The collected sand flies were then identified to species level using molecular techniques. Subsequent analyses included the detection of pathogens and the identification of pyrethroid resistance mutations within the voltage-sensitive sodium channel (Vgsc) domain IIS6 gene, followed by sequence analysis., Results: The study identified nine sand fly species belonging to the genera Phlebotomus and Sergentomyia. The DNA of Sergentomyia khawi was the only species found to test positive for one sample of Leishmania orientalis in Sadao district. This finding represents the first detection of L. orientalis in Thailand. Moreover, three samples of Leishmania martiniquensis and four samples of Trypanosoma sp. were found in the Natawi district. No I1011M, L1014F/S, V1016G, or F1020S mutations were detected in Vgsc gene., Conclusions: The results of this study provide valuable information on sand fly species and the continuous circulation of Leishmania spp. and Trypanosoma spp. in Songkhla, southern Thailand. Moreover, the development of geo-spatial information on vectors, parasites, and insecticide resistance in sand flies has the potential to provide well-informed risk assessments and evidence-based guidance for targeted vector control in Thailand. These results can serve as a foundation for integrating the One Health approach, which is crucial for disease control, considering the diverse ecological interactions among human and/or animal reservoir hosts, parasites, and sand fly vectors., (© 2024. The Author(s).)

Motivation for the study. The role of bats as hosts of Trypanosoma spp. in the Atlantic department in Colombia, as well as its taxonomic diversity has been poorly studied. Main findings. This is the first report of frequency of infection by Trypanosoma spp. in bats in the Atlántico Department in Colombia. Implications. The great adaptive capacity of bats to different ecological niches and its role as hosts of Trypanosoma spp. for wild and urban ecotopes represents a risk factor in transmission cycles of epidemiological importance. We conducted a study to evaluate the frequency of infection by Trypanosoma spp. in bats captured in wild and urban ecotopes in the Department of Atlántico in the Caribbean region of Colombia from March 2021 to May 2022. Bats were taxonomically identified, and sex, relative age, and reproductive conditions were determined. A blood sample was used for parasitological analysis and DNA extraction to amplify a region of the 18S rRNA. 125 bats were collected, with the most abundant families being Molossidae (62/125; 49.6%) and Phyllostomidae (43/125; 34.4%). Molossus molossus collected in wild habitats showed an infection frequency of 8.1% (5/61) and 4.1% (3/61) through parasitological and molecular analysis, respectively. In comparison, Noctilio albiventris collected in urban habitats showed an infection frequency of 16.6% (2/12) for both analyses. These findings represent the first records of M. molossus harboring trypanosomes for the Department of Atlántico and of N. albiventris harboring trypanosomes in Colombia.

Bats are hosts for diverse Trypanosoma species, including trypanosomes of the Trypanosoma cruzi clade. This clade is believed to have originated in Africa and diversified in many lineages worldwide. In several geographical areas, including Cameroon, no data about trypanosomes of bats has been collected yet. In this study, we investigated the diversity and phylogenetic relationships of trypanosomes of different bat species in the central region of Cameroon. Trypanosome infections were detected in six bat species of four bat families, namely Hipposideridae, Pteropodidae, Rhinolophidae, and Vespertilionidae, with an overall prevalence of 29% and the highest infection rate in hipposiderid bat species. All trypanosomes were identified as belonging to the Trypanosoma livingstonei species group with one clade that might represent an additional subspecies of T. livingstonei. Understanding the prevalence, distribution, and host range of parasites of this group contributes to our overall knowledge of the diversity and host specificity of trypanosome species that phylogenetically group at the base of the T. cruzi clade., (© 2024. The Author(s).)

Trypanosoma evansi is reportedly divided into two genotypes: types A and B. The type B is uncommon and reportedly limited to Africa: Kenya Sudan, and Ethiopia. In contrast, type A has been widely reported in Africa, South America, and Asia. However, Trypanosoma evansi type non-A/B has never been reported. Therefore, this study aims to determine the species and genotype of the Trypanozoon subgenus using a robust identification algorithm. Forty-three trypanosoma isolates from Indonesia were identified as Trypanosoma evansi using a molecular identification algorithm. Further identification showed that 39 isolates were type A and 4 isolates were possibly non-A/B types. The PML, AMN-SB1, and STENT3 isolates were likely non-A/B type Trypanosoma evansi isolated from buffalo, while the PDE isolates were isolated from cattle. Cladistic analysis revealed that Indonesian Trypanosoma evansi was divided into seven clusters based on the gRNA-kDNA minicircle gene. Clusters 6 and 7 are each divided into two sub-clusters. The areas with the highest genetic diversity are the provinces of Banten, Central Java (included Yogyakarta), and East Nusa Tenggara. The Central Java (including Yogyakarta) and East Nusa Tenggara provinces, each have four sub-clusters, while Banten has three.

The current study proposes to investigate the diversity and phylogeny of trypanosomes parasitizing wild birds from the Brazilian Atlantic Forest. Cytological examination was carried out by light microscopy of blood smears and positive birds were selected for amplification of the 18S rDNA sequence through PCR. The resulting amplicons were subjected to purification, cloning, and sequencing analysis. Phylogenetic reconstruction was conducted, including all avian trypanosomes representative's lineages. A total of ten bird samples from species of Turdus flavipes (N=1/12), T. albicollis (N=1/8), Tachyphonus coronatus (N=6/121), Thamnophilus caerulescens (N=1/22) and Synallaxis spixi (N=1/8) were positive for Trypanosoma spp. In the six specimens of T. coronatus, five distinct lineages of Trypanosoma spp. 18S-rRNA were observed in ninety sequences obtained, and using the strategy of cloning independent PCR, it was possible to observe that two of them were related to T. avium (JB01/JB02), and three were closed related to T. bennetti (JB03/ JB04/JB05). Addionaly, all fifteen sequences obtained from T. caerulescens/ S. spixi/T. flavipes/T. albicollis were identical. The present research is the first study to access molecular diversity and polyparasitism by avian trypanosomes in Brazil. The current research exhibits the wide genetic variability in avian trypanosomes and its non-specific relationship with its avian hosts.

Background: Animal African trypanosomiasis, which is caused by different species of African trypanosomes, is a deadly disease in livestock. Although African trypanosomes are often described as blood-borne parasites, there have been recent reappraisals of the ability of these parasites to reside in a wide range of tissues. However, the majority of those studies were conducted on non-natural hosts infected with only one species of trypanosome, and it is unclear whether a similar phenomenon occurs during natural animal infections, where multiple species of these parasites may be present., Methods: The infective trypanosome species in the blood and other tissues (adipose and skin) of a natural host (cows, goats and sheep) were determined using a polymerase chain reaction-based diagnostic., Results: The animals were found to harbour multiple species of trypanosomes. Different patterns of distribution were observed within the host tissues; for instance, in some animals, the blood was positive for the DNA of one species of trypanosome and the skin and adipose were positive for the DNA of another species. Moreover, the rate of detection of trypanosome DNA was highest for skin adipose and lowest for the blood., Conclusions: The findings reported here emphasise the complexity of trypanosome infections in a natural setting, and may indicate different tissue tropisms between the different parasite species. The results also highlight the need to include adipose and skin tissues in future diagnostic and treatment strategies., (© 2024. The Author(s).)

Background: Triatomines (kissing bugs) are natural vectors of trypanosomes, which are single-celled parasitic protozoans, such as Trypanosoma cruzi, T. conorhini and T. rangeli. The understanding of the transmission cycle of T. conorhini and Triatoma rubrofasciata in China is not fully known., Methods: The parasites in the faeces and intestinal contents of the Tr. rubrofasciata were collected, and morphology indices were measured under a microscope to determine the species. DNA was extracted from the samples, and fragments of 18S rRNA, heat shock protein 70 (HSP70) and glycosomal glyceraldehyde-3-phosphate dehydrogenase (gGAPDH) were amplified and sequenced. The obtained sequences were then identified using the BLAST search engine, followed by several phylogenetic analyses. Finally, laboratory infections were conducted to test whether Tr. rubrofasciata transmit the parasite to rats (or mice) through bites. Moreover, 135 Tr. rubrofasciata samples were collected from the Guangxi region and were used in assays to investigate the prevalence of trypanosome infection., Results: Trypanosoma sp. were found in the faeces and intestinal contents of Tr. rubrofasciata, which were collected in the Guangxi region of southern China and mostly exhibited characteristics typical of epimastigotes, such as the presence of a nucleus, a free flagellum and a kinetoplast. The body length ranged from 6.3 to 33.9 µm, the flagellum length ranged from 8.7 to 29.8 µm, the nucleus index was 0.6 and the kinetoplast length was -4.6. BLAST analysis revealed that the 18S rRNA, HSP70 and gGAPDH sequences of Trypanosoma sp. exhibited the highest degree of similarity with those of T. conorhini (99.7%, 99.0% and 99.0%, respectively) and formed a well-supported clade close to T. conorhini and T. vespertilionis but were distinct from those of T. rangeli and T. cruzi. Laboratory experiments revealed that both rats and mice developed low parasitaemia after inoculation with Trypanosoma sp. and laboratory-fed Tr. rubrofasciata became infected after feeding on trypanosome-positive rats and mice. However, the infected Tr. rubrofasciata did not transmit Trypanosoma sp. to their offspring. Moreover, our investigation revealed a high prevalence of Trypanosoma sp. infection in Tr. rubrofasciata, with up to 36.3% of specimens tested in the field being infected., Conclusions: Our study is the first to provide a solid record of T. conorhini from Tr. rubrofasciata in China with morphological and molecular evidence. This Chinese T. conorhini is unlikely to have spread through transovarial transmission in Tr. rubrofasciata, but instead, it is more likely that the parasite is transmitted between Tr. rubrofasciata and mice (or rats). However, there was a high prevalence of T. conorhini in the Tr. rubrofasciata from our collection sites and numerous human cases of Tr. rubrofasciata bites were recorded. Moreover, whether these T. conorhini strains are pathogenic to humans has not been investigated., (© 2024. The Author(s).)

Animal African trypanosomiasis, also known as nagana, is caused by Trypanosoma species, which cause significant clinical diseases and lead to losses in animal production. We carried out a cross-sectional survey to investigate the composition of vectors and parasite diversity in two districts in the eastern region of Ghana where pigs and cattle were exposed to tsetse bites. We performed cytochrome c oxidase subunit 1 polymerase chain reaction (PCR) to identify tsetse species and internal transcribed spacer 1 PCR to identify Trypanosoma species. Also, we investigated the source of tsetse blood meal based on mitochondrial cytochrome b gene sequence analysis. A total of 229 tsetse, 65 pigs, and 20 cattle were investigated for trypanosomes. An overall vector density of 4.3 tsetse/trap/day was observed. A trypanosome prevalence of 58.9% (95% CI = 52.5-65.1%), 46.2% (95% CI = 34.6-58.1%), and 0.0% (95% CI = 0.0-16.1%) in tsetse, pigs, and cattle, respectively, was detected. Trypanosoma congolense was predominant, with a prevalence of 33.3% (95% CI = 73.3-86.5%) in tsetse. There was evidence of multiple infections in tsetse and pigs. Approximately 39% of the tsetse were positive for multiple infections of T. congolense and Trypanosoma simiae. Parasite prevalence in pigs across the communities was high, with significant differences associated between locations (χ2 = 28.06, 95% CI = 0.05-0.81, P = 0.0009). Tsetse blood meal analysis revealed feeding on domestic Sus scrofa domesticus (pigs) and Phacochoerus africanus (warthogs). Infective tsetse may transmit trypanosomes to livestock and humans in the communities studied.

Vector-borne diseases (VBDs) affecting dromedary camels (Camelus dromedarius) have considerable importance in the United Arab Emirates (UAE) because of the consequences associated with production decline and economic losses. Our study aimed to determine the prevalence of selected VBDs in camels in the UAE and identify risk factors. This research is currently affected by the low number of epidemiological molecular surveys addressing this issue. Blood samples were obtained from 425 dromedary camels from different locations across the UAE. Whole genomic DNA was isolated, and PCR screening was done to detect piroplasmids (Babesia/Theileria spp.), Trypanosoma spp., and Anaplasmataceae spp. (Anaplasma, Ehrlichia, Neorickettsia and Wolbachia spp.). Amplicons were sequenced, and phylogenetic trees were constructed. Trypanosoma sequences were identified as T. brucei evansi, whereas Anaplasmataceae sequences were identified as A. platys-like. All camels were negative for Babesia/Theileria spp. (0%); however, 18 camels were positive for T. b. evansi (4%) and 52 were positive for A. platys-like (12%). Mixed infection with T. b. evansi and A. platys-like was found in one camel. Statistical analyses revealed that camels with a brown coat colour were significantly more prone to acquire the A. platys-like strain compared with those having a clearer coat. A similar finding was observed when comparing urban moving camels with desert indoor and urban indoor camels. Continuous disease surveillance is required to ensure and maintain the good health status of the camels in the UAE. Nonetheless, the risk of disease outbreak remains if the misuse of drugs continues., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Background Objectives: Vector-borne haemoprotozoan diseases comprise diverse group of single celled organism transmitted by haematophagus invertebrates. The current study was aimed at the identification of major haemoprotozoan (Babesia, Theileria and Trypanosoma) in dromedary camel of North Gujarat region in India using microscopy and Polymerase Chain Reaction (PCR)., Methods: A total of 234 blood samples were screened by the microscopic and molecular detection assays. Molecular prevalence studies of Theileria, Trypanosoma spp and Babesia was undertaken using 18s ribosomal DNA, RoTat 1.2 and SS rRNA gene respectively. The data relating to microscopic and molecular prevalence along with associated risk factors were analysed by statistical methods., Results: The overall prevalence of hamoprotozoan disease based on microscopic and molecular investigation was 23.50%. The sensitivity and specificity (95% Confidence Interval) of PCR assay was 100% in comparison to microscopy (45.45 % sensitive and 100 % specific). The kappa coefficient between PCR and microscopy indicated good level of agreement with a value of 0.704 and SE of 0.159., Interpretation Conclusion: Despite holding much significance to the animal sector, little work has been undertaken in regional parts of India regarding camel parasites. The present study offers first preliminary research data investigating haemoprotozoan disease using parasitological and molecular methods in camels in the region., (Copyright © 2024 Copyright: © 2024 Journal of Vector Borne Diseases.)

Background: Trypanosoma evansi is the leading infectious Trypanosoma spp. in camels (Camelus dromedarius) present in the Kingdom of Saudi Arabia (KSA) that could lead to extensive economic losses. The present study was aimed to assess the prevalence rate of T. evansi in Taif governorate, Makkah province, KSA using parasitological and molecular evaluations, and analyze their genetic relationship targeting internal transcribed spacer 1 (ITS1) and variable surface glycoprotein (VSG) genes. For evaluation, we have used 102 blood samples of camels obtained from three different regions in Taif., Results: Results show a considerable prevalence rate of trypanosomosis 2/102 (2.0%) according to Giemsa-stained buffy coat smear, and 16/102 (15.7%) according to touchdown PCR. T. evansi (n = 10/102, 9.8%) was the main infectious species found in camels then T. vivax (n = 3/102, 2.9%). Mixed infections were detected in three camels with T. evansi, T. vivax, and T. congolense (n = 3/102, 2.9%). Regarding gender, the results indicate that female camels (11/66, 16.7%) show higher prevalence of Trypanosoma than males (5/36, 13.9%). Sequencing and phylogenetic analyses of ITS1 and VSG showed their relationships with T. evansi in other hosts from different countries., Conclusions: In our peer knowledge, it is the first time to report a research-based prevalence of trypanosomosis in the camels of Taif governorate, Makkah province, KSA., (© 2022. The Author(s).)

Bagrada hilaris (Burmeister) is an invasive pest of economically important crops in the United States. During physiological investigations of B. hilaris, a flagellated protozoan was discovered in the alimentary canal of many specimens. This manuscript characterizes the morphology and molecular identification of the trypanosomatid, which appears similar to trypanosomatids identified in other stink bug species. It has been identified as a species in the Blastocrithidia genus based on morphological characteristics and molecular analyses., (Published by Oxford University Press on behalf of Entomological Society of America 2022.)

African trypanosomosis, a parasitic disease caused by protozoan parasites transmitted by tsetse flies, affects both humans and animals in sub-Saharan Africa. While the human form (HAT) is now limited to foci, the animal form (AAT) is widespread and affects the majority of sub-Saharan African countries, and constitutes a real obstacle to the development of animal breeding. The control of AAT is hampered by a lack of standardized and easy-to used diagnosis tools. This study aimed to evaluate the diagnostic potential of TbLysoPLA and TbGK proteins from Trypanosoma brucei brucei for AAT serodiagnosis in indirect ELISA using experimental and field sera, individually, in combination, and associated with the BiP C-terminal domain (C25) from T. congolense. These novel proteins were characterized in silico, and their sequence analysis showed strong identities with their orthologs in other trypanosomes (more than 60% for TbLysoPLA and more than 82% for TbGK). TbLysoPLA displays a low homology with cattle (<35%) and Piroplasma (<15%). However, TbGK shares more than 58% with cattle and between 45-55% with Piroplasma. We could identify seven predicted epitopes on TbLysoPLA sequence and 14 potential epitopes on TbGK. Both proteins were recombinantly expressed in Escherichia coli. Their diagnostic potential was evaluated by ELISA with sera from cattle experimentally infected with T. congolense and with T.b. brucei, sera from cattle naturally infected with T. congolense, T. vivax and T.b. brucei. Both proteins used separately had poor diagnostic performance. However, used together with the BiP protein, they showed 60% of sensitivity and between 87-96% of specificity, comparable to reference ELISA tests. In conclusion, we showed that the performance of the protein combinations is much better than the proteins tested individually for the diagnosis of AAT., Competing Interests: The authors have declared that no competing interests exist.

The pre-mRNA splicing factor PRP19 is recruited into the spliceosome after forming the PRP19/CDC5L complex in humans and the Nineteen complex in yeast. Additionally, 'PRP19-related' proteins enter the spliceosome individually or in pre-assemblies that differ in these systems. The protistan family Trypanosomatidae, which harbors parasites such as Trypanosoma brucei, diverged early during evolution from opisthokonts. While introns are rare in these organisms, spliced leader trans splicing is an obligatory step in mRNA maturation. So far, ∼70 proteins have been identified as homologs of human and yeast splicing factors. Moreover, few proteins of unknown function have recurrently co-purified with splicing proteins. Here we silenced the gene of one of these proteins, termed PRC5, and found it to be essential for cell viability and pre-mRNA splicing. Purification of PRC5 combined with sucrose gradient sedimentation revealed a complex of PRC5 with a second trypanosomatid-specific protein, PRC3, and PRP19-related proteins SYF1, SYF3 and ISY1, which we named PRP19-related complex (PRC). Importantly, PRC and the previously described PRP19 complex are distinct from each other because PRC, unlike PRP19, co-precipitates U4 snRNA, which indicates that PRC enters the spliceosome prior to PRP19 and uncovers a unique pre-organization of these proteins in trypanosomes., (© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Background: African Trypanosomiases threaten the life of both humans and animals. Trypanosomes are transmitted by tsetse and other biting flies. In Rwanda, the African Animal Trypanosomiasis (AAT) endemic area is mainly around the tsetse-infested Akagera National Park (NP). The study aimed to identify Trypanosoma species circulating in cattle, their genetic diversity and distribution around the Akagera NP., Methodology: A cross-sectional study was carried out in four districts, where 1,037 cattle blood samples were collected. The presence of trypanosomes was determined by microscopy, immunological rapid test VerY Diag and PCR coupled with High-Resolution Melt (HRM) analysis. A parametric test (ANOVA) was used to compare the mean Packed cell Volume (PCV) and trypanosomes occurrence. The Cohen Kappa test was used to compare the level of agreement between the diagnostic methods., Findings: The overall prevalence of trypanosome infections was 5.6%, 7.1% and 18.7% by thin smear, Buffy coat technique and PCR/HRM respectively. Microscopy showed a low sensitivity while a low specificity was shown by the rapid test (VerY Diag). Trypanosoma (T.) congolense was found at a prevalence of 10.7%, T. vivax 5.2%, T. brucei brucei 2% and T. evansi 0.7% by PCR/HRM. This is the first report of T.evansi in cattle in Rwanda. The non-pathogenic T. theileri was also detected. Lower trypanosome infections were observed in Ankole x Friesian breeds than indigenous Ankole. No human-infective T. brucei rhodesiense was detected. There was no significant difference between the mean PCV of infected and non-infected animals (p>0.162)., Conclusions: Our study sheds light on the species of animal infective trypanosomes around the Akagera NP, including both pathogenic and non-pathogenic trypanosomes. The PCV estimation is not always an indication of trypanosome infection and the mechanical transmission should not be overlooked. The study confirms that the area around the Akagera NP is affected by AAT, and should, therefore, be targeted by the control activities. AAT impact assessment on cattle production and information on the use of trypanocides are needed to help policymakers prioritise target areas and optimize intervention strategies. Ultimately, these studies will allow Rwanda to advance in the Progressive Control Pathway (PCP) to reduce or eliminate the burden of AAT., Competing Interests: The authors have declared that no competing interests exist.

Background: African trypanosomiasis, caused by protozoa of the genus Trypanosoma and transmitted by the tsetse fly, is a serious parasitic disease of humans and animals. Reliable data on the vector distribution, feeding preference and the trypanosome species they carry is pertinent to planning sustainable control strategies., Methodology: We deployed 109 biconical traps in 10 villages in two districts of northwestern Uganda to obtain information on the apparent density, trypanosome infection status and blood meal sources of tsetse flies. A subset (272) of the collected samples was analyzed for detection of trypanosomes species and sub-species using a nested PCR protocol based on primers amplifying the Internal Transcribed Spacer (ITS) region of ribosomal DNA. 34 blood-engorged adult tsetse midguts were analyzed for blood meal sources by sequencing of the mitochondrial cytochrome c oxidase 1 (COI) and cytochrome b (cytb) genes., Results: We captured a total of 622 Glossina fuscipes fuscipes tsetse flies (269 males and 353 females) in the two districts with apparent density (AD) ranging from 0.6 to 3.7 flies/trap/day (FTD). 10.7% (29/272) of the flies were infected with one or more trypanosome species. Infection rate was not significantly associated with district of origin (Generalized linear model (GLM), χ 2  = 0.018, P = 0.895, df = 1, n = 272) and sex of the fly (χ2 = 1.723, P = 0.189, df = 1, n = 272). However, trypanosome infection was highly significantly associated with the fly's age based on wing fray category (χ 2  = 22.374, P < 0.001, df = 1, n = 272), being higher among the very old than the young tsetse. Nested PCR revealed several species of trypanosomes: T. vivax (6.62%), T. congolense (2.57%), T. brucei and T. simiae each at 0.73%. Blood meal analyses revealed five principal vertebrate hosts, namely, cattle (Bos taurus), humans (Homo sapiens), Nile monitor lizard (Varanus niloticus), African mud turtle (Pelusios chapini) and the African Savanna elephant (Loxodonta africana)., Conclusion: We found an infection rate of 10.8% in the tsetse sampled, with all infections attributed to trypanosome species that are causative agents for AAT. However, more verification of this finding using large-scale passive and active screening of human and tsetse samples should be done. Cattle and humans appear to be the most important tsetse hosts in the region and should be considered in the design of control interventions., (© 2021. The Author(s).)

Trypanosomatid infections are an important public health threat affecting many low-income countries across the tropics, particularly in the Americas. Trypanosomatids can infect many vertebrate, invertebrate, and plant species and play an important role as human pathogens. Among these clinically relevant pathogens are species from the genera Leishmania and Trypanosoma . Mixed trypanosomatid infections remain a largely unexplored phenomenon. Herein, we describe the application of an amplicon-based next-generation sequencing (NGS) assay to detect and identify trypanosomatid species in mammalian reservoirs, human patients, and sand fly vectors throughout regions of Leishmania endemicity. Sixty-five samples from different departments of Colombia, including two samples from Venezuela, were analyzed: 49 samples from cutaneous leishmaniasis (CL) patients, 8 from sand flies, 2 from domestic reservoirs (Canis familiaris), and 6 from wild reservoirs (Phyllostomus hastatus). DNA from each sample served to identify the presence of trypanosomatids through conventional PCR using heat shock protein 70 (HSP70) gene as the target. PCR products underwent sequencing by Sanger sequencing and NGS, and trypanosomatid species were identified by using BLASTn against a reference database built from trypanosomatid-derived HSP70 sequences. The alpha and beta diversity indexes of amplicon sequence variants were calculated for each group. The results revealed the presence of mixed infections with more than two Leishmania species in 34% of CL samples analyzed. Trypanosoma cruzi was identified in samples from wild reservoirs, as well as in sand fly vectors. Coinfection events with three different Leishmania species were identified in domestic reservoirs. These findings depose the traditional paradigm of leishmaniasis as being a single-species-driven infection and redraw the choreography of host-pathogen interaction in the context of multiparasitism. Further research is needed to decipher how coinfections may influence disease progression. This knowledge is key to developing an integrated approach for diagnosis and treatment. IMPORTANCE Traditionally, there has been a frequent, yet incorrect assumption that phlebotomine vectors, animal reservoirs, and human hosts are susceptible to Leishmania infection by a single parasite species. However, current evidence supports that these new vector-parasite-reservoir associations lend vectors and reservoirs greater permissiveness to certain Leishmania species, thus promoting the appearance of coinfection events, particularly in disease-endemic regions. The present study describes the application of an amplicon-based next-generation sequencing (NGS) assay to detect and identify trypanosomatid species in mammalian reservoirs, human patients, and sand fly vectors from regions of endemicity for leishmaniasis. This changes our understanding of the clinical course of leishmaniasis in areas of endemicity.

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., (© 2021. The Author(s).)

Background: Bovine trypanosomosis transmitted by tsetse flies is a major constraint to cattle health and productivity in all sub-Saharan countries, including Uganda. The objectives of this study were to determine the prevalence of bovine trypanosomosis and identify its associated risk factors and the species of trypanosomes associated with the disease., Methodology: A cross-sectional study was conducted around Murchison Falls National Park, Uganda from January 2020 to April 2020. Trypanosomes were detected in blood samples by PCR analysis targeting the internal transcribed spacer 1 (ITS-PCR assays), and trypanosomes in positive blood samples were sequenced., Results: Of 460 blood samples collected and tested, 136 (29.6%) were positive for trypanosome infections and 324 (70.4%) were negative. The overall trypanosome prevalence was 29.6% (95% confidence interval 25.4-33.8%), attributed to three trypanosome species. Of these three species, Trypanosoma vivax was the most prevalent (n = 130, 28.3%) while the others were detected as mixed infections: T. vivax + Trypanosoma congolense (n = 2, 0.4%) and T. vivax + Trypanosoma evansi (n = 1, 0.2%). There were significant differences in trypanosome prevalence according to sex (χ 2  = 62, df = 1, P < 0.05), age (χ 2  = 6.28, df = 2, P = 0.0043) and cattle breed (χ 2  = 10.61, df = 1, P = 0.001)., Conclusions: Trypanosomosis remains a major limitation to cattle production around Murchison Falls National Park and interventions are urgently needed. In our study, the prevalence of trypanosome infections was high, with T. vivax identified as the most prevalent species. Age, sex and breed of cattle were risk factors for trypanosome infection., (© 2021. The Author(s).)

The closest relative of human pathogen Leishmania , the trypanosomatid Novymonas esmeraldas , harbors a bacterial endosymbiont " Candidatus Pandoraea novymonadis." Based on genomic data, we performed a detailed characterization of the metabolic interactions of both partners. While in many respects the metabolism of N. esmeraldas resembles that of other Leishmaniinae, the endosymbiont provides the trypanosomatid with heme, essential amino acids, purines, some coenzymes, and vitamins. In return, N. esmeraldas shares with the bacterium several nonessential amino acids and phospholipids. Moreover, it complements its carbohydrate metabolism and urea cycle with enzymes missing from the " Ca. Pandoraea novymonadis" genome. The removal of the endosymbiont from N. esmeraldas results in a significant reduction of the overall translation rate, reduced expression of genes involved in lipid metabolism and mitochondrial respiratory activity, and downregulation of several aminoacyl-tRNA synthetases, enzymes involved in the synthesis of some amino acids, as well as proteins associated with autophagy. At the same time, the genes responsible for protection against reactive oxygen species and DNA repair become significantly upregulated in the aposymbiotic strain of this trypanosomatid. By knocking out a component of its flagellum, we turned N. esmeraldas into a new model trypanosomatid that is amenable to genetic manipulation using both conventional and CRISPR-Cas9-mediated approaches. IMPORTANCE Novymonas esmeraldas is a parasitic flagellate of the family Trypanosomatidae representing the closest insect-restricted relative of the human pathogen Leishmania . It bears symbiotic bacteria in its cytoplasm, the relationship with which has been established relatively recently and independently from other known endosymbioses in protists. Here, using the genome analysis and comparison of transcriptomic profiles of N. esmeraldas with and without the endosymbionts, we describe a uniquely complex cooperation between both partners on the biochemical level. We demonstrate that the removal of bacteria leads to a decelerated growth of N. esmeraldas , substantial suppression of many metabolic pathways, and increased oxidative stress. Our success with the genetic transformation of this flagellate makes it a new model trypanosomatid species that can be used for the dissection of mechanisms underlying the symbiotic relationships between protists and bacteria.

Trypanosoma brucei evansi and T. brucei equiperdum are animal infective trypanosomes conventionally classified by their clinical disease presentation, mode of transmission, host range, kinetoplast DNA (kDNA) composition and geographical distribution. Unlike other members of the subgenus Trypanozoon , they are non-tsetse transmitted and predominantly morphologically uniform (monomorphic) in their mammalian host. Their classification as independent species or subspecies has been long debated and genomic studies have found that isolates within T. brucei evansi and T. brucei equiperdum have polyphyletic origins. Since current taxonomy does not fully acknowledge these polyphyletic relationships, we re-analysed publicly available genomic data to carefully define each clade of monomorphic trypanosome. This allowed us to identify, and account for, lineage-specific variation. We included a recently published isolate, IVM-t1, which was originally isolated from the genital mucosa of a horse with dourine and typed as T. equiperdum . Our analyses corroborate previous studies in identifying at least four distinct monomorphic T. brucei clades. We also found clear lineage-specific variation in the selection efficacy and heterozygosity of the monomorphic lineages, supporting their distinct evolutionary histories. The inferred evolutionary position of IVM-t1 suggests its reassignment to the T. brucei evansi type B clade, challenging the relationship between the Trypanozoon species, the infected host, mode of transmission and the associated pathological phenotype. The analysis of IVM-t1 also provides, to our knowledge, the first evidence of the expansion of T. brucei evansi type B, or a fifth monomorphic lineage represented by IVM-t1, outside of Africa, with important possible implications for disease diagnosis.

Background: African trypanosomes are parasites mainly transmitted by tsetse flies. They cause trypanosomiasis in humans (HAT) and animals (AAT). In Chad, HAT/AAT are endemic. This study investigates the diversity and distribution of trypanosomes in Mandoul, an isolated area where a tsetse control campaign is ongoing, and Maro, an area bordering the Central African Republic (CAR) where the control had not started., Methods: 717 human and 540 cattle blood samples were collected, and 177 tsetse flies were caught. Trypanosomal DNA was detected using PCR targeting internal transcribed spacer 1 (ITS1) and glycosomal glyceraldehyde-3 phosphate dehydrogenase (gGAPDH), followed by amplicon sequencing., Results: Trypanosomal DNA was identified in 14 human samples, 227 cattle samples, and in tsetse. Besides T. b. gambiense, T. congolense was detected in human in Maro. In Mandoul, DNA from an unknown Trypanosoma sp.-129-H was detected in a human with a history of a cured HAT infection and persisting symptoms. In cattle and tsetse samples from Maro, T. godfreyi and T. grayi were detected besides the known animal pathogens, in addition to T. theileri (in cattle) and T. simiae (in tsetse). Furthermore, in Maro, evidence for additional unknown trypanosomes was obtained in tsetse. In contrast, in the Mandoul area, only T. theileri, T. simiae, and T. vivax DNA was identified in cattle. Genetic diversity was most prominent in T. vivax and T. theileri., Conclusion: Tsetse control activities in Mandoul reduced the tsetse population and thus the pathogenic parasites. Nevertheless, T. theileri, T. vivax, and T. simiae are frequent in cattle suggesting transmission by other insect vectors. In contrast, in Maro, transhumance to/from Central African Republic and no tsetse control may have led to the high diversity and frequency of trypanosomes observed including HAT/AAT pathogenic species. Active HAT infections stress the need to enforce monitoring and control campaigns. Additionally, the diverse trypanosome species in humans and cattle indicate the necessity to investigate the infectivity of the unknown trypanosomes regarding their zoonotic potential. Finally, this study should be widened to other trypanosome hosts to capture the whole diversity of circulating trypanosomes., Competing Interests: The authors have declared that no competing interests exist.

Background: African trypanosomiases are vector-borne diseases that affect humans and livestock in sub-Saharan Africa. Although data have been collected on tsetse fauna as well as trypanosome infections in tsetse flies and mammals in foci of sleeping sickness in Chad, the situation of tsetse fly-transmitted trypanosomes remains unknown in several tsetse-infested areas of Chad. This study was designed to fill this epidemiological knowledge gap by determining the tsetse fauna as well as the trypanosomes infecting tsetse flies in the area of Lake Iro in southeastern Chad., Methods: Tsetse flies were trapped along the Salamat River using biconical traps. The proboscis and tsetse body were removed from each fly. DNA was extracted from the proboscis using proteinase K and phosphate buffer and from the tsetse body using Chelex 5%. Tsetse flies were identified by amplifying and sequencing the cytochrome c oxydase I gene of each tsetse fly. Trypanosome species were detected by amplifying and sequencing the internal transcribed spacer 1 of infecting trypanosomes., Results: A total of 617 tsetse flies were trapped; the apparent density of flies per trap per day was 2. 6. Of the trapped flies, 359 were randomly selected for the molecular identification and for the detection of infecting trypanosomes. Glossina morsitans submorsitans (96.1%) was the dominant tsetse fly species followed by G. fuscipes fuscipes (3.1%) and G. tachinoides (0.8%). Four trypanosome species, including Trypanosoma vivax, T. simiae, T. godfreyi and T. congolense savannah, were detected. Both single infection (56.7%) and mixed infections of trypanosomes (4.6%) were detected in G. m. submorsitans. The single infection included T. simiae (20.5%), T. congolense savannah (16.43%), T. vivax (11.7%) and T. godfreyi (9.8%). The trypanosome infection rate was 61.4% in G. m. submorsitans, 72.7% in G. f. fuscipes and 66.6% in G. tachinoides. Trypanosome infections were more prevalent in tsetse bodies (40.6%) than in the proboscis (16.3%)., Conclusion: This study revealed the presence of different tsetse species and a diversity of trypanosomes pathogenic to livestock in the area of Lake Iro. The results highlight the risks and constraints that animal African trypanosomiasis pose to livestock breeding and the importance of assessing trypanosome infections in livestock in this area.

Toxoplasma gondii can infect almost all warm-blooded vertebrates with pathogensis being largely influenced by the host immune status. As important epidemiological hosts, rodents are globally distributed and are also commonly found infected with haemoflagellates, such as those in the genus Trypanosoma. We here address whether and how co-infection with trypanosomes can influence T. gondii infection in laboratory models. Rats of five strains, co-infected with T. lewisi and mice of four strains, co-infected with T. musculi, were found to be more or less susceptible to T. gondii infection, respectively, with corresponding increased or decreased brain cyst burdens. Downregulation of iNOS expression and decreased NO production or reverse were observed in the peritoneal macrophages of rats or mice, infected with trypanosomes, respectively. Trypanosoma lewisi and T. musculi can modulate host immune responses, either by enhancement or suppression and influence the outcome of Toxoplasma infection.

Differentiation is a central aspect of the parasite life cycle and encompasses adaptation to both host and environment. If we accept that evolution cannot anticipate an organism's needs as it enters a new environment, how do parasite differentiation pathways arise? The transition between vertebrate and insect stage African trypanosomes is probably one of the better studied and involves a cell-cycle arrested or 'stumpy' form that activates metabolic pathways advantageous to the parasite in the insect host. However, a range of stimuli and stress conditions can trigger similar changes, leading to formation of stumpy-like cellular states. We propose that the origin and optimisation of this differentiation program represents repurposing of a generic stress response to gain considerable gain-of-fitness associated with parasite transmission., (Copyright © 2020 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: In the 1980s and 1990s, great strides were taken towards the elimination of tsetse and animal African trypanosomiasis (AAT) in Zimbabwe. However, advances in recent years have been limited. Previously freed areas have been at risk of reinvasion, and the disease in tsetse-infested areas remains a constraint to food security. As part of ongoing control activities, monitoring of tsetse and AAT is performed regularly in the main areas at risk. However, a centralized digital archive is missing. To fill this gap, a spatially explicit, national-level database of tsetse and AAT (i.e. atlas) was established through systematic data collation, harmonization and geo-referencing for the period 2000-2019., Methods: The atlas covers an area of approximately 70,000 km 2 , located mostly in the at-risk areas in the north of the country. In the tsetse component, a total of 33,872 entomological records were assembled for 4894 distinct trap locations. For the AAT component, 82,051 samples (mainly dry blood smears from clinically suspicious animals) were collected at 280 diptanks and examined for trypanosomal infection by microscopy., Results: Glossina pallidipes (82.7% of the total catches) and Glossina morsitans morsitans (17.3%) were the two tsetse species recorded in the north and northwest parts of the country. No fly was captured in the northeast. The distribution of AAT follows broadly that of tsetse, although sporadic AAT cases were also reported from the northeast, apparently because of transboundary animal movement. Three trypanosome species were reported, namely Trypanosoma brucei (61.7% of recorded infections), Trypanosoma congolense (28.1%) and Trypanosoma vivax (10.2%). The respective prevalences, as estimated in sentinel herds by random sampling, were 2.22, 0.43 and 0.30%, respectively., Discussion: The patterns of tsetse and AAT distributions in Zimbabwe are shaped by a combination of bioclimatic factors, historical events such as the rinderpest epizootic at the turn of the twentieth century and extensive and sustained tsetse control that is aimed at progressively eliminating tsetse and trypanosomiasis from the entire country. The comprehensive dataset assembled in the atlas will improve the spatial targeting of surveillance and control activities. It will also represent a valuable tool for research, by enabling large-scale geo-spatial analyses.

Specimens of a Trypanosoma sp. were found in a haemolymph sample of Rhipicephalus microplus from Argentina. Polymerase chain reaction (PCR) was done targeting the SSU rRNA gene of Trypanosoma spp. and a fragment of 2300 base pairs (bp) was amplified, subsequently a phylogenetic analysis was conducted, based on an alignment of 905 bp, containing the sequence of the Argentina isolate and sequences of different Trypanosoma species retrieved from GenBank. Phylogenetic analysis revealed that this trypanosome is not related to Trypanosoma theileri as was previously thought, instead the strain of Trypanosoma detected in this study can be provisionally determined as belonging to the recently described organism Trypanosoma rhipicephalis. Furthermore, phylogenetic analysis performed in this work revealed that T. rhipicephalis belongs to a novel clade of tick-related trypanosomes, most with limited genetic data, for which essential aspects of both the vertebrate and invertebrate life cycles are lacking. The lack of basic information restricts the inferences that can be done from the present finding and, in addition, points out a clear knowledge gap in the biology of this group of trypanosomes., (Copyright © 2020 Elsevier GmbH. All rights reserved.)

Background: Trypanosomes are single-celled eukaryotic parasites characterised by the unique biology of their mitochondrial DNA. African livestock trypanosomes impose a major burden on agriculture across sub-Saharan Africa, but are poorly understood compared to those that cause sleeping sickness and Chagas disease in humans. Here we explore the potential of the maxicircle, a component of trypanosome mitochondrial DNA to study the evolutionary history of trypanosomes., Results: We used long-read sequencing to completely assemble maxicircle mitochondrial DNA from four previously uncharacterized African trypanosomes, and leveraged these assemblies to scaffold and assemble a further 103 trypanosome maxicircle gene coding regions from published short-read data. While synteny was largely conserved, there were repeated, independent losses of Complex I genes. Comparison of pre-edited and non-edited genes revealed the impact of RNA editing on nucleotide composition, with non-edited genes approaching the limits of GC loss. African tsetse-transmitted trypanosomes showed high levels of RNA editing compared to other trypanosomes. The gene coding regions of maxicircle mitochondrial DNAs were used to construct time-resolved phylogenetic trees, revealing deep divergence events among isolates of the pathogens Trypanosoma brucei and T. congolense., Conclusions: Our data represents a new resource for experimental and evolutionary analyses of trypanosome phylogeny, molecular evolution and function. Molecular clock analyses yielded a timescale for trypanosome evolution congruent with major biogeographical events in Africa and revealed the recent emergence of Trypanosoma brucei gambiense and T. equiperdum, major human and animal pathogens.

The eukaryotic blood parasite genus Trypanosoma includes several important pathogens of humans and livestock, but has been understudied in wildlife broadly. The trypanosomes that infect birds are in particular need of increased attention, as these parasites are abundant and globally distributed, yet few studies have addressed their evolutionary origins and diversity using modern molecular and analytical approaches. Of specific interest are the deep evolutionary relationships of the avian trypanosomes relative to the trypanosome species that are pathogenic in humans, as well as their species level diversity in regions where they have been understudied such as North America. Here, we address these unresolved areas of study using phylogenomic data for two species of avian trypanosomes that were isolated as "bycatch" from host transcriptome assemblies, as well as a large 18S DNA barcode sequence dataset that includes 143 novel avian Trypanosoma 18S sequences from North America. Using a phylogenomic approach, we find that the avian trypanosomes are nested within a clade of primarily mammalian trypanosomes that includes the human pathogen Trypanosoma cruzi, and are paraphyletic with respect to the ruminant trypanosome Trypanosoma theileri. DNA barcode sequences showed that T. avium and an unidentified small, non-striated trypanosome that was morphologically similar to T. everetti are each represented by highly abundant and divergent 18S haplotypes in North America. Community-level sampling revealed that additional species-level Trypanosoma lineages exist in this region. We compared the newly sequenced DNA barcodes from North America to a global database, and found that avian Trypanosoma 18S haplotypes generally exhibited a marked lack of host specificity with at least one T. avium haplotype having an intercontinental distribution. This highly abundant T. avium haplotype appears to have a remarkably high dispersal ability and cosmopolitan capacity to evade avian host immune defenses, which warrant further study., Competing Interests: The authors have declared that no competing interests exist.

The Indonesian island of Sulawesi is a globally significant biodiversity hotspot with substantial undescribed biota, particularly blood-borne parasites of endemic wildlife. Documenting the blood parasites of Sulawesi's murine rodents is the first fundamental step towards the discovery of pathogens likely to be of concern for the health and conservation of Sulawesi's endemic murines. We screened liver samples from 441 specimens belonging to 20 different species of murine rodents from 2 mountain ranges on Sulawesi, using polymerase chin reaction (PCR) primers targeting the conserved 18S rDNA region across the protozoan class Kinetoplastea. We detected infections in 156 specimens (10 host species) with a mean prevalence of 35.4% (95% confidence interval [CI] = 30.9-39.8%). Sequences from these samples identified 4 infections to the genus Parabodo, 1 to Blechomonas, and the remaining 151 to the genus Trypanosoma. Within Trypanosoma, we recovered 17 haplotypes nested within the Trypanosoma theileri clade infecting 117 specimens (8 host species) and 4 haplotypes nested within the Trypanosoma lewisi clade infecting 34 specimens (6 host species). Haplotypes within the T. theileri clade were related to regional Indo-Australian endemic trypanosomes, displayed geographic structuring but with evidence of long-term connectivity between mountains, and had substantial phylogenetic diversity. These results suggest T. theileri clade parasites are native to Sulawesi. Conversely, T. lewisi clade haplotypes were recovered from both endemic and introduced rodents, demonstrated complete geographic separation between clades, and had low genetic diversity. These results suggest that the T. lewisi clade parasites invaded Sulawesi recently and likely in 2 separate invasion events. Our results provide the first records of metakinetoplastids in Sulawesi's rodents and highlight the need for more extensive sampling for pathogens in this biodiversity hotspot., (© American Society of Parasitologists 2020.)

Polo-like kinases (Plks) are evolutionarily conserved serine/threonine protein kinases playing crucial roles during multiple stages of mitosis and cytokinesis in yeast and animals. Plks are characterized by a unique Polo-box domain, which plays regulatory roles in controlling Plk activation, interacting with substrates and targeting Plk to specific subcellular locations. Plk activity and protein abundance are subject to temporal and spatial control through transcription, phosphorylation and proteolysis. In the early branching protists, Plk orthologues are present in some taxa, such as kinetoplastids and Giardia , but are lost in apicomplexans, such as Plasmodium . Works from characterizing a Plk orthologue in Trypanosoma brucei , a kinetoplastid protozoan, discover its essential roles in regulating the inheritance of flagellum-associated cytoskeleton and the initiation of cytokinesis, but not any stage of mitosis. These studies reveal evolutionarily conserved and species-specific features in the control of Plk activation, substrate recognition and protein abundance, and suggest the divergence of Plk function and regulation for specialized needs in this flagellated unicellular eukaryote.

Background: Species of the Tabanidae are potent vectors of human and animal diseases, but they have not been thoroughly investigated to date. In Senegal (West Africa), little information is available on these dipterans. Our objective in this study was to investigate Senegalese tabanids and their diversity by using molecular and proteomics approaches, as well as their associated pathogens., Methods: A total of 171 female tabanids were collected, including 143 from Casamance and 28 from Niokolo-Koba. The samples were identified morphologically by PCR sequencing and by MALDI-TOF MS, and PCR analysis was employed for pathogen detection and blood-meal characterization., Results: The morphological identification revealed four species concordantly with the molecular identification: Atylotus fuscipes (79.5%), Tabanus guineensis (16.4%), Chrysops distinctipennis (3.5%) and Tabanus taeniola (0.6%) (not identified by PCR). The molecular investigation of pathogens revealed the presence of Trypanosoma theileri (6.6%), Leishmania donovani (6.6%), Setaria digitata (1.5%), Rickettsia spp. (5.1%) and Anaplasmataceae bacteria (0.7%) in A. fuscipes. Tabanus guineensis was positive for L. donovani (35.7%), S. digitata (3.6%) and Anaplasmataceae (17.8%). Leishmania donovani has been detected in 50% of C. distinctipennis specimens and the only T. taeniola specimen. No Piroplasmida, Mansonella spp. or Coxeilla burnetii DNA was detected. In addition to humans (96.43%), Chlorocebus sabeus, a non-human primate, has been identified as a host of (3.57%) analysed tabanids. MALDI-TOF MS enabled us to correctly identify all tabanid species that had good quality spectra and to create a database for future identification., Conclusions: Tabanids in Senegal could be vectors of several pathogens threatening animal and public health. To fully characterize these dipterans, it is therefore necessary that researchers in entomology and infectiology employ molecular characterization and mass spectrometric techniques such as MALDI-TOF MS to analyse these dipterans in Senegal and West Africa.

Background: Wildlife species carry a remarkable diversity of trypanosomes. The detection of trypanosome infection in native Australian fauna is central to understanding their diversity and host-parasite associations. The implementation of total RNA sequencing (meta-transcriptomics) in trypanosome surveillance and diagnosis provides a powerful methodological approach to better understand the host species distribution of this important group of parasites., Methods: We implemented a meta-transcriptomic approach to detect trypanosomes in a variety of tissues (brain, liver, lung, skin, gonads) sampled from native Australian wildlife, comprising four marsupials (koala, Phascolarctos cinereus; southern brown bandicoot, Isoodon obesulus; swamp wallaby, Wallabia bicolor; bare-nosed wombat, Vombatus ursinus), one bird (regent honeyeater, Anthochaera phrygia) and one amphibian (eastern dwarf tree frog, Litoria fallax). Samples corresponded to both clinically healthy and diseased individuals. Sequencing reads were de novo assembled into contigs and annotated. The evolutionary relationships among the trypanosomatid sequences identified were determined through phylogenetic analysis of 18S rRNA sequences., Results: We detected trypanosome sequences in all six species of vertebrates sampled, with positive samples in multiple organs and tissues confirmed by PCR. Phylogenetic analysis indicated that the trypanosomes infecting marsupials were related to those previously detected in placental and marsupial mammals, while the trypanosome in the regent honeyeater grouped with avian trypanosomes. In contrast, we provide the first evidence for a trypanosome in the eastern dwarf tree frog that was phylogenetically distinct from those described in other amphibians., Conclusions: To our knowledge, this is the first meta-transcriptomic analysis of trypanosomes in native Australian wildlife, expanding the known genetic diversity of these important parasites. We demonstrated that RNA sequencing is sufficiently sensitive to detect low numbers of Trypanosoma transcripts and from diverse hosts and tissues types, thereby representing an effective means to detect trypanosomes that are divergent in genome sequence.

Trypanosomes have long been recognised as being amongst the most important protozoan parasites of vertebrates, from both medical and veterinary perspectives. Whilst numerous insect species have been identified as vectors, the role of ticks is less well understood. Here we report the isolation and partial molecular characterisation of a novel trypanosome from questing Ixodes ricinus ticks collected in Slovakia. The trypanosome was isolated in tick cell culture and then partially characterised by microscopy and amplification of fragments of the 18S rRNA and 24Sα rDNA genes. Analysis of the resultant sequences suggests that the trypanosome designated as Trypanosoma sp. Bratislava1 may be a new species closely related to several species or strains of trypanosomes isolated from, or detected in, ticks in South America and Asia, and to Trypanosoma caninum isolated from dogs in Brazil. This study highlights the potential involvement of ixodid ticks in the epidemiology of trypanosomes, as well as the use of tick cell lines for isolation of such tick-borne protozoa. Further studies are required to investigate the epidemiology, transmission and life cycle of this putative novel species., (Copyright © 2020 The Authors. Published by Elsevier GmbH.. All rights reserved.)

Trypanosomes are ubiquitous blood parasites of fishes and at least 16 species were originally described infecting African freshwater fishes. This number was later reduced to six and in the late 1990s it was proposed that most records of freshwater fish trypanosomes across Africa are Trypanosoma mukasai Hoare, 1932. Recently, results from a molecular analysis of fish trypanosomes from the Okavango Delta, Botswana, reported the presence of at least two genotypic groups and concluded that the identification of T. mukasai remains problematic. The aims of the present study were thus to elucidate the life cycle of a freshwater fish trypanosome from southern Africa and to do a morphological and molecular characterization of this parasite from both the fish host and leech vector. To locate trypanosome stages, leeches were removed from fishes captured in the Phongolo River, South Africa, and fish blood films and leech squashes were Giemsa-stained and screened. To determine whether trypanosome stages in fishes and leeches were of the same genotype, DNA was extracted and fragments of the 18S rDNA gene were amplified and sequenced. Trypanosomes were detected in the fish families Cichlidae, Clariidae, Mochokidae and Schilbeidae. Sequence data showed that the trypanosome from one of the leeches, identified as Batracobdelloides tricarinata (Blanchard, 1897), was highly similar to those obtained from the plain squeaker, Synodontis zambezensis, with 0.7% difference recorded between them. From morphological and molecular data presented here, it is clear that the trypanosomes from Phongolo are closely related to those of the Okavango and should be considered as a single diverse species with genetic differentiation between 0.4-2.9%, under the 3-5% differences expected to be seen between true distinct species within the rRNA. Developmental stages of the trypanosome found in the leech B. tricarinata supports its status as the vector and the molecular evidence shows the relationship between the trypanosome in the fish and leech, but also illustrates the exceptional genetic and morphological diversity of a single species of trypanosome between host species. The work presented here provides us with clear information to take further steps in resolving the taxonomy and systematics of African freshwater fish trypanosomes., (Copyright © 2020 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.)

This study investigated the occurrence and phylogenetic relationship of protozoan parasites and Ehrlichia infecting domestic animals from three municipalities in uMkhanyakude district of KwaZulu-Natal province, South Africa. A total of 208 blood samples collected from clinically healthy cattle, sheep, goats and dogs from uMkhanyakude district were examined by polymerase chain reaction (PCR) assays, using either genus or species-specific primers to determine the occurrence and phylogenetic relationship of various protozoan parasites and Ehrlichia of veterinary importance. A total of 5/109 (4.6%) cattle were PCR-positive for the presence of Toxoplasma gondii, 33/109 (30.3%) for Babesia bovis, 24/109 (22.02%) for Babesia bigemina and 20/109 (18.3%) for Trypanosoma sp., while 3/10 (30%) of sheep were PCR-positive for Theileria ovis and none of the goats were positive for any of the detected pathogens. The co-infection of 4/109 (3.7%) B. bovis and B. bigemina was detected in cattle. Only Ehrlichia canis was detected in dogs with infection rate of 20/48 (41.7%). Sequences of PCR-positive isolates (B. bovis, B. bigemina, E. canis, T. ovis and T. gondii) showed that they were closely related to their relevant species from various countries. These findings have expanded our knowledge about the prevalence and phylogenetic similarity between protozoan parasites and Ehrlichia isolates of South African origin. To date, this is the first study in South Africa to detect T. gondii infections from cattle blood using PCR.

Detection of trypanosomes in tsetse or domestic livestock is a basic requirement for epidemiological studies as well as for planning and implementing control measures against tsetse and trypanosomiasis. This epidemiological study aimed at assessing the prevalence of trypanosomes in pigs and tsetse flies in the Jomoro district of the western region of Ghana using molecular techniques. Blood was collected from pigs and biconical traps were used to collect tsetse flies. DNA was isolated from 300 pig blood samples and 300 flies for trypanosome detection and identification by PCR. Packed Cell Volume (PCV) of blood samples from 300 pigs was measured using a micro-haematocrit reader. Glossina palpalis palpalis was the only tsetse species found in the area with fly apparent density of 18.4 fly/trap/day. An overall prevalence of trypanosomes in the study area was 4.3% and 0.8% in pigs and tsetse flies respectively. Mixed infection with Trypanosoma (T.) congolense forest and T. vivax was most prevalent 46.2% followed by single infection of T. vivax 15.4%, T. congolense and a mixed infection of T. congolense, T. vivax and T. brucei sl. were the least with 7.7% each. There were no significant differences in trypanosome prevalence among different age groups and between both sexes of the studied pigs (p > 0.05). Trypanosome prevalence was lower in healthy looking 1.9% than the sick looking 20%, pigs (P < 0.05). Mean PCV of parasitaemic pigs 29.3% was significantly lower than that of aparasitaemic pigs 37.8%. Two out of the five species-specific primers used could not identify any trypanosome species from the total blood samples examined. This could possibly mean that those species are not found in the present study area. These results provide useful background information for further study and justification to extend tsetse control to the Jomoro district., Competing Interests: Declaration of Competing Interest All authors declare that they have no Conflict of Interest for the publication of this Manuscript., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The increasing number and severity of surra outbreaks in the Philippines led the government to consider it as the second most important disease of livestock in the country. It is one of the most economically important animal parasitic diseases and has been reported in several animal species, including water buffaloes, cattle, and horses in different regions of the Philippines. However, it has not yet been reported in Cebu, the usual gateway of livestock trade in the area that raises 6% of the 3.75 million goats in the country. In the current study, a PCR-based assay was conducted for the molecular detection and characterization of Trypanosoma evansi in goats in Cebu. A total of 251 goats were randomly sampled from four farms. DNA was extracted and ITS1-PCR was applied to detect different trypanosomes in goats. Eighty-five out of the 251 (33.9%) samples tested positive for T. evansi, two of which were also positive for T. theileri-like trypanosome. The detection rate of T. evansi was slightly higher in male goats (38.3%) than in females (32.5%), and in younger goats (34.5%) than in adults (33.5%). The findings, however, did not differ significantly to suggest any association between sex and age with T. evansi infection in goats. The detection of T. evansi and T. theileri-like trypanosome in goats was confirmed by sequence analysis of ITS1 region. To our knowledge, this is the first report on the molecular detection and identification of caprine T. evansi infection in Cebu, Philippines., Competing Interests: Declaration of Competing Interest The authors declare no competing interests in association with this study., (Copyright © 2020. Published by Elsevier B.V.)