Next Article in Journal
Zoonotic Disease Threats: Are We Prepared?
Previous Article in Journal
The Attribution of Human Seasonal Influenza H3N2 Virus Detection to the Collector, Not Avian Sources, During the 2022 Highly Pathogenic Avian Influenza Outbreak in Pennsylvania, USA—Implications for Biosafety and Biosecurity
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Zoonotic Diseases
Volume 4
Issue 4
10.3390/zoonoticdis4040028
zoonoticdis-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Spillover of Trypanosoma lewisi and Trypanosoma musculi Allied Trypanosomes from Rodents to Bats in the Roofs of Human Dwellings: Synanthropic Bats as a Potential New Source of Human Opportunistic Trypanosomes

by
Evaristo Villalba-Alemán
1,*,
Luciana Lima
1,
Paola Andrea Ortiz
2,
Bruno Rafael Fermino
1,3,
Gladys Elena Grisante
4,
Carla Monadeli Filgueira Rodrigues
1,3,
Letícia Pereira Úngari
1,
Néstor Añez
4,
Herakles Antonio Garcia
1,* and
Marta Maria Geraldes Teixeira
1
1
Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo 05508-000, Brazil
2
Laboratorio de Investigaciones en Parasitología Tropical (LIPT), Facultad de Ciencias, Universidad del Tolima, Ibagué 730006299, Colombia
3
Instituto Federal de Rondônia (IFRO), Rondônia 76890-000, Brazil
4
Departamento de Biologia, Facultad de Ciencias, Universidad de Los Andes, Merida 5101, Venezuela
*
Authors to whom correspondence should be addressed.
Zoonotic Dis. 2024, 4(4), 320-336; https://doi.org/10.3390/zoonoticdis4040028
Submission received: 5 November 2024 / Revised: 14 December 2024 / Accepted: 19 December 2024 / Published: 22 December 2024
Browse Figures
Review Reports Versions Notes

Simple Summary

This study provides molecular evidence of Trypanosoma species typically linked to domestic rats and mice, transmitted by fleas, present in bats from Brazil and Venezuela. This unique finding suggests that trypanosomes from rodents can jump to bats, highlighting cross-species transmission. The bats carried rodent-associated parasites, which could pose an opportunistic infection risk to humans, and also harbored other trypanosomes, including the human pathogen Trypanosoma cruzi. The study emphasizes the need for a One Health approach to evaluate the potential risks these emerging trypanosomes may pose to humans.

Abstract

Bats and rodents serve as reservoirs for numerous zoonotic pathogens, including species of Trypanosoma and Leishmania. Domestic rats host the flea-transmitted Trypanosoma (Herpetosoma) lewisi, which can be associated with humans, particularly young or immunocompromised individuals. Using Fluorescent Fragment Length Barcoding (FFLB) and phylogenetic analyses based on SSU rRNA sequences, we identified two Herpetosoma species, T. lewisi-like and T. musculi-like species, in bats of different families inhabiting rooftops and peridomestic structures in Brazil (44%, 107 bats examined) and Venezuela (50%, 52 bats examined). These species are typically associated with Rattus spp. (domestic rats) and Mus musculus (house mice), respectively. Furthermore, bats were co-infected with up to five other species, including Trypanosoma dionisii, Trypanosoma cruzi marinkellei, and isolates from the Trypanosoma Neobat clade, all strongly associated with bats, and Trypanosoma cruzi and Trypanosoma rangeli, known to infect various mammals, including humans. Therefore, our findings expand the known host range of Herpetosoma to bats, marking the first report of potential spillover of Herpetosoma trypanosomes from rodents to bats and underscoring the potential for the cross-species transmission of flea-borne trypanosomes. These results highlight the need for a One Health approach to assess infection risks associated with trypanosome spillover from synanthropic rodents and bats to humans.

1. Introduction

Bats are notable reservoirs of diverse viruses, bacteria, fungi, and protistan parasites, thriving in their blood due to their unique predisposition for hosting numerous zoonotic agents, often capable of cross-species transmission. Additionally, bats can disperse a broad range of parasitic fauna due to their long lifespans, social behavior, and ability to fly long distances. Increasing anthropogenic disturbances to natural habitats are driving bats into urban areas, where they are attracted by abundant food sources and artificial shelters, such as the roofs of human dwellings. This close proximity to domestic animals and humans raises the potential for host switches, heightening the risk of emerging pathogens from synanthropic bats and rodents affecting human health [1,2,3,4].
Trypanosomes infect both domestic and wild animals, as well as humans. While most species are considered non-pathogenic to their natural hosts, some cause significant veterinary and human diseases. Currently, 11 species of trypanosomes have been confirmed to infect Chiroptera (bats). Among them, T. cruzi and T. rangeli are the only species known to infect humans and mammals of other orders, including rodents [5,6,7,8,9]. In addition, the number of trypanosome species hosted by bats, currently known only by DNA sequences, has increased due to molecular surveys in Latin America, Europe, Africa, and Asia [9,10,11,12,13,14,15].
Despite extensive molecular surveys of trypanosomes in bats worldwide, no trypanosome (nor DNA sequences) detected in bats was phylogenetically placed in the subgenus Herpetosoma. This subgenus was established to accommodate trypanosomes infecting species of the order Rodentia worldwide, including its type species Trypanosoma lewisi of Rattus spp. and Trypanosoma musculi infecting house mice (Mus musculus) [5]. Phylogenetic inferences have supported the subgenus Herpetosoma as a clade comprising trypanosomes from a diversity of domestic and wild rodents and lagomorphs [7,16,17,18,19,20]. Exclusively using the morphology of blood forms and host restrictions as taxonomic parameters, several species of Herpetosoma were reported across the world, most still requiring molecular phylogenetic validation [5]. Moreover, trypanosomes found in marsupials have been recently included in this subgenus [21], and parasites from shrews and moles (insectivorous mammals of the order Eulipotyphla), traditionally classified in the subgenus Megatrypanum, were placed at the edge of the clade comprising all species of Herpetosoma [17,19]. Trypanosomes found in bats were classified as Herpetosoma due to the morphology of their blood forms: T. lineatum from vespertilionid bats in Venezuela and T. longiflagellum from emballonurid bats in Iraq [5]. However, molecular studies of these two species are restrained by the lack of available cultures and blood smears.
The transmission of Herpetosoma spp. is cyclically mediated by fleas, which become infected by ingesting blood meals containing trypomastigotes. These trypomastigotes differentiate into proliferative epimastigotes in the flea gut and subsequently into infective metacyclic trypomastigotes, which are excreted in the flea feces. The parasites enter the host through wounds caused by flea bites, ingestion of whole fleas during grooming, licking flea feces deposited on the host’s fur, or injuries resulting from animal fighting. Trypanosoma musculi is thought to be specific to house mice (Mus musculus) and is transmitted by lice and fleas [5,22,23]. Both T. lewisi and T. musculi multiply extracellularly in the bloodstream and within the capillaries of domestic rats and house mice, respectively. Failed experimental cross-infections indicated a strong association between these trypanosomes and their respective hosts. After recovering from primary infections, hosts are resistant to reinfection. Both T. lewisi and T. musculi can induce abortions, anemia, thrombocytosis, and lethal infections in naive, undernourished, and immunocompromised rodents [24,25,26,27], and modulate host immune responses exacerbating Toxoplasma gondii infections [28].
Trypanosoma lewisi was dispersed worldwide through infected rats accompanying human movement. Invasive black rats harboring T. lewisi induced a rapid decline and the extinction of native Rattus spp. on Christmas Island, Australia [29]. Previously thought to be restricted to Rattus spp., T. lewisi and T. lewisi-like have been identified globally in both invasive Rattus spp. and native rodents [16,19,30,31,32,33,34]. A high prevalence of T. lewisi infection in rats has been reported in densely populated and poor human settlements, particularly in Asia, Africa, and South America [16,32,35,36,37,38,39,40,41]. T. lewisi can act as an opportunistic parasite in humans, particularly young, undernourished, or immuno-compromised individuals. Infected individuals generally exhibit mild and non-specific symptoms, including transient fever, lethargy, and anorexia, with most recovering naturally. However, infants and immunocompromised patients require medical intervention, as the infection can occasionally progress to death [38,42,43,44,45,46,47,48,49,50,51,52].
Human and rodent infections with T. lewisi were traditionally detected through microscopy of blood smears and, more recently, through PCR methods for parasite detection in host blood and fleas [16,31,35,37]. Sequence analysis of PCR-amplified ITS rDNA and SSU rRNA amplicons has revealed both novel trypanosome species and a breakdown of host specificity within Herpetosoma [44,47,53,54,55].
In this study, we used the highly sensitive Fluorescent Fragment Length Barcoding (FFLB) method, combined with phylogenetic analysis of SSU rRNA sequences, to detect and identify previously undetected Herpetosoma spp. in synanthropic bats cohabiting with rodents and humans in Brazil and Venezuela.

2. Materials and Methods

2.1. Studied Areas, Bat Captures, and Blood Collection

Over the past two decades, we have conducted molecular surveys of trypanosomes in more than 2600 bats captured in South America and Africa. From these surveys, we have maintained extensive laboratory collections of blood samples and cultures of bat trypanosomes, along with corresponding blood smears and DNA samples. Our laboratory houses a substantial culture repository (TCC: Trypanosomatid Culture Collection) and a large databank of DNA sequences from bat trypanosomes. Based on previous surveys of bat trypanosomes conducted in our laboratories, this study focused on blood samples (bolded codes in Groups 2 and 3 of Table 1) from two collection sites: (1) bats (n = 52) captured in 2013 in the municipality of Santa Bárbara del Zulia (09°00′01.19″ N, 71°54′50.36″ W) in the state of Zulia, Venezuela, and (2) bats (n = 107) captured in 2014 in Angicos (05°39′46.56″ S, 36°36′04.92″ W) in the state of Rio Grande do Norte, Brazil. Zulia is characterized by extensive wetlands, while Rio Grande do Norte lies within the semi-arid Caatinga biome in northeastern Brazil (Figure 1). Both regions experience high temperatures year-round and are home to a diverse fauna of bats and other small mammals. The domestic and peridomestic environments where bats were captured are inhabited by an abundance of domestic rodents.
Captures of bats were performed during the day using a manual net for bats resting in roofs and at night using mist nets positioned near places used by bats to leave shelters in human buildings. Captured bats were anesthetized, and blood samples collected by heart puncture were used for microscopical searches for trypanosomes in fresh blood samples and Giemsa-stained blood smears, as well as for hemocultures and preservation in 99% ethanol (v/v) for molecular studies, as performed in our previous studies [6,56,57].
The capture of bats and ensuing manipulations were carried out in Brazil according to IBAMA (Brazilian Institute for the Environment and Renewable Natural Resources, Permit Number 10080-2) and in Venezuela according to procedures recommended by the Ethical Committee of the Los Andes University, Merida. This study was approved by the Committee on the Ethics of Animal Experimentation of the Institute of Biomedical Sciences, University of São Paulo, where all molecular characterizations were performed (Protocol nº 1038/2019 and CEP-ICB nº17/page 3/book2).

2.2. DNA Extraction and Fluorescent Fragment Length Barcoding (FFLB)

DNA was extracted from bat blood samples preserved in ethanol using the ammonium acetate precipitation protocol and submitted to Fluorescent Fragment Length Barcoding (FFLB) as described previously [58,59]. This method is based on PCR-amplification of four small regions of variable SSU rRNA genes using fluorescent-labeled primers, allowing for high sensitivity in the detection of length polymorphism among amplified fragments using an ABI 3500 DNA sequencer (Applied Biosystems®, Waltham, MA, USA). The barcodes consist of profiles formed by four DNA fragments that can differentiate genera, subgenera, species, and genotypes of trypanosomes, depending on the level of sequence conservation in rRNA genes [37,56,59,60].

2.3. Nested-PCR for the Identification of Trypanosomes in Bat Blood Samples

To characterize the species of trypanosomes of the subgenus Herpetosoma detected by FFLB in bat blood samples, we employed a nested PCR assay targeting a variable region of the SSU rRNA gene (~560 bp) as described [56,57]. Blood samples containing more than one species/genotype, indicated by FFLB profiles, were submitted to cloning of the PCR-amplified DNA fragments using the TOPO™ TA Cloning™ vector (Invitrogen™, California, USA). To assess mixed infections, 5–10 clones per sample were purified using the Wizard Plus SV Minipreps Kit (Promega, Madison, USA) and sequenced [56,57].

2.4. Phylogenetic and Single Nucleotide Polymorphism (SNP) Analyses of SSU rRNA Gene Sequences

Sequences from SSU rRNA obtained from Herpetosoma trypanosomes present in bat blood samples were submitted to Blast search and aligned with homologous sequences of closely related trypanosomes available in GenBank using CLUSTALW with subsequent manual adjustments (Table S1). Phylogenetic relationships were inferred by Maximum Likelihood (ML) and Maximum Parsimony (MP) analyses. Parsimony and bootstrap analyses were carried out using PAUP* v. 4.0b10 [61]. ML was conducted using RAxML v.7.0.0 [62], with nodal support estimated with 500 bootstrap replicates in RAxML using GTRGAMMA. Thoroughly aligned SSU rRNA sequences were analyzed using ade4, adegenet, and tidyverse packages in RStudio v. 2022.07.2 [63,64].

3. Results and Discussion

3.1. Trypanosoma (Herpetosoma) spp. and Other Trypanosomes Identified by FFLB in Synanthropic Bats

The FFLB method offers high sensitivity and allows for the simultaneous characterization of the entire repertoire of trypanosomes in large numbers of individual samples from cultures, blood, and vectors, regardless of low parasitemia or co-infections with multiple parasites [37,56,58,59,60,65]. We compared the profiles generated using DNA from cultures of T. lewisi (from Venezuelan rats), T. lewisi-like (from Brazilian rats and monkeys, as well as African rats), Trypanosoma blanchardi (from European hamsters), and Trypanosoma nabiasi (from domestic rabbits). Results showed a single profile confirming the suitability of FFLB to detect Trypanosoma (Herpetosoma) spp. The specific FFLB profile shared by species of Herpetosoma is as follows: 18S1 = 239–241, 18S3 = 216–217, 28S1 = 263–265, 28S2 = 189–192 (Figure 2). The effectiveness of the FFLB technique for field studies was demonstrated in our previous surveys for T. lewisi in the blood of R. rattus and their fleas in Venezuela [37].
In this study, we characterized the trypanosome repertoire in bats captured at two collection sites where some bats exhibited FFLB profiles of Herpetosoma. We detected these trypanosomes in 50% (26 out of 52 bats examined) of bats in Zulia, western Venezuela, and in 44% (47 of 107 bats) in Rio Grande do Norte, northeastern Brazil (Figure 3a). In addition to Herpetosoma spp., FFLB revealed a high diversity of trypanosome species in bats from both localities. Most bats carried more than one species of trypanosome, with multiple infections involving four to five different parasites (Figure 3b). FFLB showed that bats captured from the same or nearby shelters shared trypanosomes, including Trypanosoma cruzi, Trypanosoma cruzi marinkellei, Trypanosoma dionisii, Trypanosoma rangeli, and unnamed trypanosomes of the Neobat clade (Figure 3a). While each trypanosome species exhibited unique FFLB profiles, those of the Neobat clade (currently comprising at least six species of trypanosomes), widespread in neotropical bats, can share highly similar profiles, requiring DNA sequencing for species identification [56,66]. The human-infective T. cruzi and T. rangeli were highly prevalent in bats from Venezuela and Brazil, identified in various species and families (Figure 3a,b). Notably, T. cruzi, the agent of Chagas disease, and T. rangeli, a non-pathogenic parasite to humans, are widespread in Neotropical bats and other animals, producing co-infections in humans, other mammal hosts, and triatomine bugs [60].

3.2. Phylogenetic Analyses of Herpetosoma Trypanosomes from Bats and Rodents Using SSU rRNA Sequences

In phylogenetic analysis of the subgenus Herpetosoma, species identification relied on small polymorphisms of SSU rRNA gene sequences. We obtained sequences from Herpetosoma trypanosomes present in the blood of 7 bats, four from Venezuela and three from Brazil (Table 1). The analysis of these sequences, aligned with those retrieved from GenBank (Supplementary Table S1), positioned the trypanosomes found in bats within the subgenus Herpetosoma (Figure 4), within the major subclade formed by closely related trypanosomes allied with T. lewisi, which were distributed into Groups 1–4, while other trypanosomes from rodents of many families and lagomorphs clustered in other subclades (Figure 4 and Figure 5).
The reference strain of T. lewisi (Molteno) from R. rattus in England, well known regarding morphology, host behavior, and cross-experimental infections, clustered into group 1, along with another isolate from R. rattus from the US, Ethiopia, Indonesia, Kenya, and Venezuela (Figure 5; Table 1). T. lewisi from Venezuelan R. rattus clustered into group 1, whereas those found in Venezuelan bats (MOVE16, 44, 46, and 49) clustered into group 2, which we refer to as T. lewisi-like, as it does not include any named trypanosome species (Figure 5; Table 1). Group 2 also included T. lewisi-like from Mozambican and Brazilian Rattus spp. and from captive Neotropical Brazilian monkeys (Aotus, Alouatta, and Callithrix). Group 3 included sequences of Herpetosoma sp. found in Brazilian bats (isolates RNMO 66, 72, and 81) that are nearly identical to the sequence of reference T. musculi from house mice. This group also comprises T. rabinowitschae and T. blanchardi from European mice, as well as Trypanosoma niviventerae from native rodents in China. We refer to this cluster as the T. musculi Group (Table 1; Figure 5), as it includes T. musculi. To our knowledge, this is the first molecular report of T. musculi-like sequences in a South American host. Group 4 (Figure 4 and Figure 5) includes Trypanosoma grosi, found in Asian rodents of the genus Apodemus [67,68], as well as sequences obtained from trypanosomes detected in various African rodents, including species of genera Praomys, Acomys, Mastomys, and Mus [19].
Comparison of SSU rRNA sequences confirmed high sequence conservation among T. lewisi and its allied species, with specific Single Nucleotide Polymorphisms (SNPs) restricted to three regions between nucleotides 70–200, 390–520, and 685–790 (Figure 6). Consistent with their phylogenetic relationships, T. lewisi and its allied trypanosomes displayed high genetic homogeneity, while other Herpetosoma species exhibited notable divergences, forming groups of trypanosomes clearly distinguished by an increased number of SNPs (Figure 6).
The phylogenetic evidence from field studies for distinct groups of trypanosomes detected in Rattus spp. and classified as T. lewisi, combined with the finding that Rattus spp. can host Herpetosoma trypanosomes other than T. lewisi, challenges the use of host restriction as a taxonomic parameter for species identification [7,16,19,34]. Nevertheless, despite the proven lack of strict host restriction, overall, cross-experimental infections support notable degrees of host species constraints, with laboratory evidence supporting the restriction of T. lewisi and T. musculi for Rattus spp. and Mus musculus, respectively [5,17,18,34,37,67]. Altogether, the phylogenetic analyses from this and previous studies highlighted the need for a comprehensive taxonomical appraisal of the subgenus Herpetosoma under a phylogenetic framework, regardless of host–species of origin. It is crucial to clarify the species boundaries of the type-species T. lewisi and its closely related trypanosomes that have been found in house rats and mice, wild rodents, human and non-human primates, and, after the present study, also in bats.
Therefore, according to our analysis of the trypanosomes nested into the subclade T. lewisi of the subgenus Herpetosoma, we referred to the trypanosomes within groups 1, 2, and 3 as T. lewisi (group T. lewisi), T. lewisi-like (group T. lewisi-like), and T. musculi-like (group T. musculi), respectively (Figure 4 and Figure 5; Table 1). Any possible existence of small genetic divergences among trypanosomes within each group cannot be resolved due to the high conservation of SSU rRNA sequences (Figure 5). Despite high conservation, SSU rRNA sequences are valuable for positioning trypanosomes within this subgenus and its main groups/subclades (Figure 4). In addition, the more polymorphic gGAPDH, SL RNA, and Cathepsin L-like gene sequences have supported equal phylogenetic segregation patterns within Herpetosoma trypanosomes inferred using SSU rRNA sequences [6,16,50,69]. Unfortunately, most reports of T. lewisi from humans and rodents in Asia have been based on the morphology of blood trypomastigotes and/or ITS rDNA sequences, thus precluding a global comparison of Herpetosoma spp. [7,19,31,34,40,44,47].

3.3. Trypanosomes in Bat Blood Smears and Haemocultures

The morphological identification of Herpetosoma trypanosomes in the bloodstream of Neotropical bats presents three main challenges: very low parasitemia, questionable species-specific morphological differentiation, and frequent co-infections. These limitations hinder the identification of T. lewisi and other Herpetosoma species in Neotropical bats, as these trypanosomes are morphologically indistinguishable from one another and from T. rangeli, a common species in Neotropical bats. In fact, the shared morphological traits of blood trypomastigotes once led to T. rangeli being misclassified in the subgenus Herpetosoma [5], a mistake later corrected by molecular phylogenetic studies that justified the creation of the subgenus Aneza to accommodate T. rangeli [6,9,69,70,71].
In this study, microscopic examination of Giemsa-stained blood smears from Venezuelan bats revealed trypomastigotes in only 6.25% (2 out of 32) of the bats with blood smears. One bat exhibited a short, C-shaped trypomastigote morphologically resembling those of the subgenus Schizotrypanum [6], while the other showed a long and slender trypomastigote resembling those of both T. rangeli and T. lewisi [5,6,69]. Additionally, we found blood trypomastigotes in 6.41% (5 out of 39) of Brazilian bat blood smears: five displaying trypomastigotes typical of Schizotrypanum, and one also showing a trypomastigote resembling those of T. rangeli or T. lewisi. Furthermore, two bats exhibited larger and wider blood forms similar to those reported for T. wauwau of the Neobat clade [72]. Of the two bats exhibiting blood forms suggestive of Herpetosoma spp., T. lewisi was confirmed in one Brazilian bat by FFLB. Unfortunately, few hemocultures of bat trypanosomes were obtained from both the Venezuelan and Brazilian surveys and all isolates cryopreserved were identified as T. dionisii. However, using the same haemoculture protocol, we previously isolated T. lewisi-like trypanosomes from Rattus spp. and monkeys, regardless of whether trypanosomes were detectable in blood smears [6,7,16,56,57,72].

3.4. Biological, Ecological and Geographical Characteristics of Bats Harboring Herpetosoma spp.

Bats harboring Herpetosoma trypanosomes were predominantly synanthropic, residing in the roofs of human dwellings and peridomestic structures, whether inhabited or abandoned. In Venezuela, FFLB identified Herpetosoma in blood samples from six out of nine bat species across three families examined: 1. Noctilionidae, comprising the insectivorous Noctilio leporinus and N. albiventris; 2. Molossidae, including the insectivorous Molossus molossus and Euromops glaucinus; and 3. Phyllostomidae, which encompassed both insectivorous species (Uroderma bilobatum, Lophostoma brasiliense, and Carollia perspicillata) and insectivorous/nectarivorous species (Glossophaga soricina and Glossophaga longirostis). Herpetosoma spp. were absent in U. bilobatum, N. leporinus, and L. brasiliense (Table 1, Figure 3a). Some bats exclusively harbored Herpetosoma spp., with notable infection rates found in N. albiventris (22%), G. soricina (20%), and E. glaucinus (29%) (Figure 3a). These bats were found sheltering in roofs of human dwellings and a sports gymnasium in the populated municipality of Santa Barbara del Zulia, western Venezuela. T. lewisi-like sequences were obtained from four bats: three N. albiventris and one C. perspicillata (Table 1, Figure 5). In Brazil, FFLB identified Herpetosoma in bats from five families captured within and around a small village in the municipality of Angicos, Northeastern Brazil: Molossidae: the insectivorous M. molossus; Vespertilionidae: the insectivorous Myotis sp; Emballonuridae: the insectivorous Peropteryx macrotis; and Noctilionidae, the insectivorous N. albiventris. Additionally, from the family Phyllostomidae, the following species were identified as hosts of Herpetosoma: the omnivorous Trachops cirrhosus (which consumes insects, small reptiles, amphibians, rodents, and can eat other bats), the insectivorous/nectarivorous Lonchorhina aurita, Glossophaga soricina, Lonchophylla sp., and Carollia perspicillata (flower-visiting bats attracted by nocturnal blooms of Cactaceae abundant in the area), as well as the hematophagous Desmodus rotundus and Diphylla ecaudata, which prefer mammals and birds as blood sources, respectively. FFLB detected the highest prevalence of Herpetosoma spp. in P. macrotis (40%), T. cirrhosis (28%), D. ecaudata (22%), and M. molossus (18%). Brazilian bats of Molossidae and Vespertilionidae shared the roofs of human dwellings, while other bats, including the hematophagous, were captured near goat and chicken enclosures.
Data from the present study indicate that trypanosomes of Herpetosoma can infect a wide range of bat species with diverse feeding habits. A total of 18 bat species from five families were found to harbor Herpetosoma trypanosomes in Venezuela and Brazil (Figure 3). The significant infection rates suggest dynamic parasite transmission within bat colonies, likely mediated by fleas and grooming behavior. Domestic rats and mice, as well as the synanthropic opossum (Didelphis sp.), often shelter in the roofs of human buildings inhabited by bats, thus favoring host-switching of parasites. We previously demonstrated that Rattus sp. living in human dwellings is a common host of T. lewisi in Brazil and Venezuela [16,37]. Previous studies have suggested that synanthropic Rattus spp. are the primary source of T. lewisi for human infections worldwide. Rodents trapped in impoverished human settlements in Africa and Asia exhibit the highest infection rates from both fleas and T. lewisi compared to wild rodents [39,40,41,73,74,75].

3.5. Active or Transient Infections of Bats by T. lewisi-like and T. musculi-like Species?

We detected Herpetosoma spp. in bats from only two capture sites, wetlands in Venezuela and a semi-arid region in Brazil, among various areas examined during over two decades, which included more than 2600 bats across different regions in South America and Africa. Despite numerous recent studies on bat trypanosomes employing molecular approaches, Herpetosoma spp. have not been previously identified in bats [9,10,11,12,13,14,15]. This unexpected discovery raises questions about whether the DNA of Herpetosoma spp. detected in bats reflect active or transient infections or merely remnants of non-infective trypanosomes that have promptly succumbed to the innate immune defenses of bats. The nature of infections in bats caused by T. lewisi-like and T. musculi-like trypanosomes, whether they are underestimated, opportunistic, active, or transient, remains to be investigated.
The low intensity of FFLB peaks indicated very low parasitemia of Herpetosoma spp. in all bats, even though SSU RNA sequences could be determined from blood samples of seven bats, and at least two Herpetosoma species were detected by nested-PCR sequencing. Low parasitemias in both natural hosts and experimental infections have been demonstrated for Herpetosoma spp. Following the inoculation of T. lewisi in Rattus spp., a pre-patent period of three to five days precedes exponential parasite growth, resulting in high parasitemia lasting approximately one week and terminating abruptly due to specific trypanocidal mechanisms. T. musculi exhibits a plateau phase of about ten days with a constant low number of parasites, followed by more gradual clearance of blood trypomastigotes compared with T. lewisi [5,7,25,26].
There is increasing evidence from field studies that Herpetosoma species from rodents may infect coexisting animals of different species and even other orders [16,19,34,37]. These trypanosomes can, opportunistically, spill over to humans and non-human primates [7,46,48,49]. Here, we demonstrated that Venezuelan bats harbor a T. lewisi-like trypanosome we previously reported from captive monkeys and Rattus spp. Experimental infection of Rattus sp. with this T. lewisi-like species followed the course typical of T. lewisi, with an early phase showing many trypanosomes followed by a quick decline of parasitemia). In contrast, this species of trypanosome causes, apparently, a transient infection in monkeys [7].
Parasites of bats, rats, and monkeys are known to frequently jump to humans, contributing to the emergence of zoonoses that threaten human health [1,2,4,76,77]. Understanding the diversity of trypanosomes in synanthropic bats and rodents is crucial for predicting spillovers of potential rodent- and bat-borne pathogens. Studies on host-switching of bat trypanosomes are vital for both bat conservation and the management and prevention of infections caused by T. lewisi and other potentially opportunistic trypanosomes. This framework should emphasize the potential for underdiagnosed human infections and promote the use of specific and sensitive diagnostic tools and surveillance of synanthropic rats, mice, and bats in high-risk areas of poverty worldwide.

4. Conclusions

Besides rodents, only primates (both human and non-human) and bats—identified for the first time in this study—have been recognized as natural hosts of T. lewisi and closely related trypanosomes transmitted by fleas. In the present study, we report the detection of T. lewisi-like and T. musculi-like trypanosomes in the blood of synanthropic bats from various species and families in Brazil and Venezuela. This finding highlights the potential for parasite spillover from domestic rats and mice to bats cohabiting in human dwellings and peri-domestic environments. Understanding the role of bats as active hosts for Herpetosoma trypanosomes and the associated risk of bats serving as reservoirs for human opportunistic infections requires a robust One Health approach.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/zoonoticdis4040028/s1, Table S1: Trypanosoma spp. of the subgenus Herpetosoma included in the phylogenetic analyses: host and geographic origin, and GenBank accession numbers of SSU rRNA sequences.

Author Contributions

Funding acquisition, resources, conceptualization, data curation, writing—original draft, review & editing, M.M.G.T.; data acquisition and curation, formal analysis, validation, writing—original draft, review & editing, E.V.-A., H.A.G. and L.P.Ú.; fieldwork, data acquisition, review & editing, L.L., P.A.O., B.R.F., G.E.G., C.M.F.R. and N.A. All authors have read and agreed to the published version of the manuscript.

Funding

This work was supported by the Brazilian agencies CNPq (PROSUL) (490254/2011-0) and FAPESP (016/07487-0).

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The original contributions presented in this study are included in the article/Supplementary Material Table S1. Further inquiries can be directed to the corresponding author.

Acknowledgments

The authors are grateful to many people for their invaluable assistance with bat captures, to Marta Campaner for her exceptional technical support, and to Erney P. Camargo for his encouragement, insights, and valuable discussions throughout the entire study.

Conflicts of Interest

The authors declare that they have no conflicts of interest.

References

  1. Hayman, D.T.; Bowen, R.A.; Cryan, P.M.; McCracken, G.F.; O’Shea, T.J.; Peel, A.J.; Gilbert, A.; Webb, C.T.; Wood, J.L. Ecology of zoonotic infectious diseases in bats: Current knowledge and future directions. Zoonoses Public Health 2013, 60, 2–21. [Google Scholar] [CrossRef] [PubMed]
  2. Plowright, R.K.; Parrish, C.R.; McCallum, H.; Hudson, P.J.; Ko, A.I.; Graham, A.L.; Lloyd-Smith, J.O. Pathways to zoonotic spillover. Nat. Rev. Microbiol. 2017, 15, 502–510. [Google Scholar] [CrossRef] [PubMed]
  3. Blasdell, K.R.; Morand, S.; Laurance, S.G.W.; Doggett, S.L.; Hahs, A.; Trinh, K.; Perera, D.; Firth, C. Rats and the city: Implications of urbanization on zoonotic disease risk in Southeast Asia. Proc. Natl. Acad. Sci. USA 2022, 119, e2112341119. [Google Scholar] [CrossRef] [PubMed]
  4. Dhivahar, J.; Parthasarathy, A.; Krishnan, K.; Kovi, B.S.; Pandian, G.N. Bat-associated microbes: Opportunities and perils, an overview. Heliyon 2023, 9, e22351. [Google Scholar] [CrossRef]
  5. Hoare, C.A. The Trypanosomes of Mammals: A Zoological Monograph; Part 2 Systematic; Blackwell Scientific Publications: Oxford, UK, 1972; pp. 123–625. [Google Scholar]
  6. Maia da Silva, F.M.; Marcili, A.; Ortiz, P.A.; Epiphanio, S.; Campaner, M.; Catão-Dias, J.L.; Shaw, J.J.; Camargo, E.P.; Teixeira, M.M.G. Phylogenetic, morphological and behavioural analyses support host switching of Trypanosoma (Herpetosoma) lewisi from domestic rats to primates. Infect. Genet. Evol. 2010, 10, 522–529. [Google Scholar] [CrossRef]
  7. Cavazzana, M.; Marcili, A.; Lima, L.; da Silva, F.M.; Junqueira, Â.C.V.; Veludo, H.H.; Viola, L.B.; Campaner, M.; Nunes, V.L.B.; Paiva, F.; et al. Phylogeographical, ecological and biological patterns shown by nuclear (ssrRNA and gGAPDH) and mitochondrial (Cyt b) genes of trypanosomes of the subgenus Schizotrypanum parasitic in Brazilian bats. Int. J. Parasitol. 2010, 40, 345–355. [Google Scholar] [CrossRef]
  8. Hamilton, P.B.; Teixeira, M.M.G.; Stevens, J.R. The Evolution of Trypanosoma cruzi: The ’Bat Seeding’ Hypothesis. Trends Parasitol. 2012, 28, 136–141. [Google Scholar] [CrossRef]
  9. Espinosa-Álvarez, O.; Ortiz, P.A.; Lima, L.; Costa-Martins, A.G.; Serrano, M.G.; Herder, S.; Buck, G.A.; Camargo, E.P.; Hamilton, P.B.; Stevens, J.R.; et al. Trypanosoma rangeli is phylogenetically closer to Old World trypanosomes than to Trypanosoma cruzi. Int. J. Parasitol. 2018, 48, 569–584. [Google Scholar] [CrossRef]
  10. Pinto, C.M.; Ocaña-Mayorga, S.; Tapia, E.E.; Lobos, S.E.; Zurita, A.P.; Aguirre-Villacís, F.; MacDonald, A.; Villacís, A.G.; Lima, L.; Teixeira, M.M.G.; et al. Bats, trypanosomes, and triatomines in Ecuador: New insights into the diversity, transmission, and origins of Trypanosoma cruzi and Chagas disease. PLoS ONE 2015, 10, e0139999. [Google Scholar] [CrossRef]
  11. Dario, M.A.; Lisboa, C.V.; Costa, L.M.; Moratelli, R.; Nascimento, M.P.; Costa, L.P.; Reis Leite, Y.L.; Llewellyn, M.S.; Das Chagas Xavier, S.C.C.; Rodrigues Roque, A.L.; et al. High Trypanosoma spp. diversity is maintained by bats and triatomines in Espírito Santo state, Brazil. PLoS ONE 2017, 12, e0188412. [Google Scholar] [CrossRef]
  12. Cai, Y.; Wang, X.; Zhang, N.; Li, J.; Gong, P.; He, B.; Zhang, X. First report of the prevalence and genotype of Trypanosoma spp. in bats in Yunnan Province, Southwestern China. Acta Trop. 2019, 198, e105105. [Google Scholar] [CrossRef] [PubMed]
  13. Clément, L.; Dietrich, M.; Markotter, W.; Fasel, N.J.; Monadjem, A.; López-Baucells, A.; Scaravelli, D.; Théou, P.; Pigeault, R.; Ruedi, M.; et al. Out of Africa: The origins of the protozoan blood parasites of the Trypanosoma cruzi clade found in bats from Africa. Mol. Phylogenet. Evol. 2020, 145, 106705. [Google Scholar] [CrossRef] [PubMed]
  14. Thiombiano, N.G.; Boungou, M.; Chabi, B.A.M.; Oueda, A.; Werb, O.; Schaer, J. First investigation of blood parasites of bats in Burkina Faso detects Hepatocystis parasites and infections with diverse Trypanosoma spp. Parasitol. Res. 2023, 122, 3121–3129. [Google Scholar] [CrossRef] [PubMed]
  15. Torres, J.M.; de Oliveira, C.E.; Santos, F.M.; Sano, N.Y.; Martinez, É.V.; Alves, F.M.; Tavares, L.E.R.; Roque, A.L.R.; Jansen, A.M.; Herrera, H.M. Trypanosomatid diversity in a bat community of an urban area in Campo Grande, Mato Grosso do Sul, Brazil. Infect. Genet. Evol. 2024, 118, 105563. [Google Scholar] [CrossRef]
  16. Ortiz, P.A.; Garcia, H.A.; Lima, L.; da Silva, F.M.; Campaner, M.; Pereira, C.L.; Jittapalapong, S.; Neves, L.; Desquesnes, M.; Camargo, E.P.; et al. Diagnosis and Genetic Analysis of the Worldwide Distributed Rattus-borne Trypanosoma (Herpetosoma) lewis and Its Allied Species in Blood and Fleas of Rodents. Infect. Genet. Evol. 2018, 63, 380–390. [Google Scholar] [CrossRef]
  17. Mafie, E.; Saito-Ito, A.; Kasai, M.; Hatta, M.; Rivera, P.T.; Ma, X.H.; Chen, E.R.; Sato, H.; Takada, N. Integrative taxonomic approach of trypanosomes in the blood of rodents and soricids in Asian countries, with the description of three new species. Parasitol. Res. 2019, 118, 97–109. [Google Scholar] [CrossRef]
  18. Goodrich, I.; McKee, C.; Kosoy, M. Trypanosoma (Herpetosoma) diversity in rodents and lagomorphs of New Mexico with a focus on epizootological aspects of infection in Southern Plains woodrats (Neotoma micropus). PLoS ONE 2020, 15, e0244803. [Google Scholar] [CrossRef]
  19. Votýpka, J.; Stříbrná, E.; Modrý, D.; Bryja, J.; Bryjová, A.; Lukeš, J. Unexpectedly high diversity of trypanosomes in small sub-Saharan mammals. Int. J. Parasitol. 2022, 52, 647–658. [Google Scholar] [CrossRef]
  20. Wang, S.; Wang, S.; Han, X.; Hornok, S.; Wang, H.; Wang, N.; Liu, G.; Yang, M.; Wang, Y. Novel Trypanosomatid species detected in Mongolian pikas (Ochotona pallasi) and their fleas in northwestern China. Parasit. Vectors 2024, 17, 152. [Google Scholar] [CrossRef]
  21. Averis, S.; Thompson, R.C.A.; Lymbery, A.J.; Wayne, A.F.; Morris, K.D.; Smith, A. The diversity, distribution and host-parasite associations of trypanosomes in Western Australian wildlife. Parasitology 2009, 136, 1269–1279. [Google Scholar] [CrossRef]
  22. Maraghi, S.; Wallbanks, K.R.; Molyneux, D.H. Oral transmission of trypanosomes of the subgenus Herpetosoma from small mammals. Parasitol. Res. 1995, 81, 693–695. [Google Scholar] [CrossRef] [PubMed]
  23. Smith, A.; Telfer, S.; Burthe, S.; Bennett, M.; Begon, M. A role for vector-independent transmission in rodent trypanosome infection? Int. J. Parasitol. 2006, 36, 1359–1366. [Google Scholar] [CrossRef]
  24. Viens, P.; Roger, M.; Dubois, R. Influence of pregnancy on mouse immunity to Trypanosoma musculi. Trans. R. Soc. Trop. Med. Hyg. 1983, 77, 274–275. [Google Scholar] [CrossRef] [PubMed]
  25. Albright, J.; Albright, J. Rodent trypanosomes: Their conflict with the immune system of the host. Parasitol. Today 1991, 7, 137–140. [Google Scholar] [CrossRef]
  26. Monroy, F.P.; Dusanic, D.G. The kidney stage of Trypanosoma musculi: A distinct stage in the life cycle? Trends Parasitol. 2000, 16, 107–110. [Google Scholar] [CrossRef]
  27. Budovsky, A.; Prinsloo, I.; El-On, J. Pathological developments mediated by cyclophosphamide in rats infected with Trypanosoma lewisi. Parasitol. Int. 2006, 55, 237–242. [Google Scholar] [CrossRef]
  28. Gao, J.M.; Yi, S.Q.; Geng, G.Q.; Xu, Z.S.; Hide, G.; Lun, Z.R.; Lai, D.H. Infection with Trypanosoma lewisi or Trypanosoma musculi may promote the spread of Toxoplasma gondii. Parasitology 2021, 148, 703–711. [Google Scholar] [CrossRef]
  29. Wyatt, K.B.; Campos, P.F.; Gilbert, M.T.; Kolokotronis, S.O.; Hynes, W.H.; DeSalle, R.; Ball, S.J.; Daszak, P.; MacPhee, R.D.; Greenwood, A.D. Historical mammal extinction on Christmas Island (Indian Ocean) correlates with introduced infectious disease. PLoS ONE 2008, 3, e3602. [Google Scholar] [CrossRef]
  30. Desquesnes, M.; Dávila, A.M. Applications of PCR-Based Tools for Detection and Identification of Animal Trypanosomes: A Review and Perspectives. Vet. Parasitol. 2002, 109, 213–231. [Google Scholar] [CrossRef]
  31. Tang, H.J.; Lan, Y.G.; Wen, Y.Z.; Zhang, X.C.; Desquesnes, M.; Yang, T.B.; Yu, X.B.; Lun, Z.R. Detection of Trypanosoma lewisi from wild rats in Southern China and its genetic diversity based on the ITS1 and ITS2 sequences. Infect. Genet. Evol. 2012, 12, 1046–1051. [Google Scholar] [CrossRef]
  32. Cassan, C.; Diagne, C.A.; Tatard, C.; Gauthier, P.; Dalecky, A.; Bâ, K.; Kane, M.; Niang, Y.; Diallo, M.; Sow, A.; et al. Leishmania major and Trypanosoma lewisi infection in invasive and native rodents in Senegal. PLoS Negl. Trop. Dis. 2018, 12, e0006615. [Google Scholar] [CrossRef] [PubMed]
  33. Dobigny, G.; Gauthier, P.; Houéménou, G.; Dossou, H.J.; Badou, S.; Etougbétché, J.; Tatard, C.; Truc, P. Spatio-temporal survey of small mammal-borne Trypanosoma lewisi in Cotonou, Benin, and the potential risk of human infection. Infect. Genet. Evol. 2019, 75, 103967. [Google Scholar] [CrossRef] [PubMed]
  34. Babyesiza, W.S.; Katakweba, A.; Fornůsková, A.; Ssuunaf, J.; Akoth, S.; Mpagi, J.; Goüy de Bellocq, J.; Bryja, J.; Votýpka, J. Trypanosome Diversity in Small Mammals in Uganda and the Spread of Trypanosoma lewisi to Native Species. Parasitol. Res. 2024, 123, 54. [Google Scholar] [CrossRef] [PubMed]
  35. Milocco, C.; Kamyingkird, K.; Desquesnes, M.; Jittapalapong, S.; Herbreteau, V.; Chaval, Y.; Douangboupha, B.; Morand, S. Molecular demonstration of Trypanosoma evansi and Trypanosoma lewisi DNA in wild rodents from Cambodia, Lao PDR, and Thailand. Transbound. Emerg. Dis. 2013, 60, 17–26. [Google Scholar] [CrossRef] [PubMed]
  36. Archer, C.E.; Schoeman, M.C.; Appleton, C.C.; Mukaratirwa, S.; Hope, K.J.; Matthews, G. Predictors of Trypanosoma lewisi in Rattus norvegicus from Durban, South Africa. J. Parasitol. 2018, 104, 187–195. [Google Scholar] [CrossRef]
  37. Garcia, H.A.; Rangel, C.J.; Ortiz, P.A.; Calzadilla, C.O.; Coronado, R.A.; Silva, A.J.; Peréz, A.M.; Lecuna, J.C.; García, M.E.; Aguirre, A.M.; et al. Zoonotic trypanosomes in rats and fleas of Venezuelan slums. Ecohealth 2019, 16, 523–533. [Google Scholar] [CrossRef]
  38. Kumar, R.; Gupta, S.; Bhutia, W.D.; Vaid, R.K.; Kumar, S. Atypical human trypanosomosis: Potentially emerging disease with lack of understanding. Zoonoses Public Health 2022, 69, 259–276. [Google Scholar] [CrossRef]
  39. Nguyen, L.K.H.; Koizumi, N.; Ung, T.H.T.; Le, T.T.; Hirayama, K.; Hasebe, F.; Hoang, V.M.P.; Khong, M.T.; Le, T.Q.M.; Miura, K. Detection of Trypanosoma lewisi DNA from Rattus norvegicus and Rattus rattus in Hanoi, Vietnam. Vector Borne Zoonotic Dis. 2022, 22, 159–161. [Google Scholar] [CrossRef]
  40. Tanthanathipchai, N.; Mitsuwan, W.; Chaisiri, K.; Thaikoed, S.; Pereira, M.L.; Paul, A.K.; Saengsawang, P. Trypanosoma lewisi in blood of Rattus rattus complex residing in human settlements, Nakhon Si Thammarat, Thailand: Microscopic and molecular investigations. Comp. Immunol. Microbiol. Infect. Dis. 2023, 98, 102010. [Google Scholar] [CrossRef]
  41. Islam, M.M.; Farag, E.; Hassan, M.M.; Enan, K.A.; Mohammadi, A.; Aldiqs, A.K.; Alhussain, H.; Al Musalmani, E.; Al-Zeyara, A.A.; Al-Romaihi, H.; et al. Rodent-borne parasites in Qatar: A possible risk at the human-animal-ecosystem interface. One Health 2024, 18, 100708. [Google Scholar] [CrossRef]
  42. Johnson, P.D. A case of infection by Trypanosoma lewisi in a child. Trans. R. Soc. Trop. Med. Hyg. 1933, 26, 467–468. [Google Scholar] [CrossRef]
  43. Shrivastava, K.K.; Shrivastava, G.P. Two cases of Trypanosoma (Herpetosoma) species infection of man in India. Trans. R. Soc. Trop. Med. Hyg. 1974, 68, 3–4. [Google Scholar] [CrossRef] [PubMed]
  44. Howie, S.; Guy, M.; Fleming, L.; Bailey, W.; Noyes, H.; Faye, J.A.; Pepin, J.; Greenwood, B.; Whittle, H.; Molyneux, D.; et al. A Gambian infant with fever and an unexpected blood film. PLoS Med. 2006, 3, 1508–1512. [Google Scholar] [CrossRef] [PubMed]
  45. Sarataphan, N.; Vongpakorn, M.; Nuansrichay, B.; Autarkool, N.; Keowkarnkah, T.; Rodtian, P.; Stich, R.W.; Jittapalapong, S. Diagnosis of a Trypanosoma lewisi-like (Herpetosoma) infection in a sick infant from Thailand. J. Med. Microbiol. 2007, 56, 1118–1121. [Google Scholar] [CrossRef] [PubMed]
  46. Lun, Z.; Reid, S.A.; Lai, D.; Li, F. Atypical human trypanosomiasis: A neglected disease or just an unlucky accident? Trends Parasitol. 2009, 25, 107–108. [Google Scholar] [CrossRef]
  47. Verma, A.; Manchanda, S.; Kumar, N.; Sharma, A.; Goel, M.; Banerjee, P.S.; Garg, R.; Singh, B.P.; Balharbi, F.; Lejon, V.; et al. Trypanosoma lewisi or T. lewisi-like infection in a 37-day-old Indian infant. Am. J. Trop. Med. Hyg. 2011, 85, 221–224. [Google Scholar] [CrossRef]
  48. Truc, P.; Büscher, P.; Cuny, G.; Gonzatti, M.I.; Jannin, J.; Joshi, P.; Prayag, J.; Lun, Z.; Mittioli, R.; Pays, E.; et al. Atypical human infections by animal trypanosomes. PLoS Negl. Trop. Dis. 2013, 7, e2256. [Google Scholar] [CrossRef]
  49. Lun, Z.R.; Wen, Y.Z.; Uzureau, P.; Lecordier, L.; Lai, D.H.; Lan, Y.G.; Desquesnes, M.; Geng, G.Q.; Yang, T.B.; Zhou, W.L.; et al. Resistance to normal human serum reveals Trypanosoma lewisi as an underestimated human pathogen. Mol. Biochem. Parasitol. 2015, 199, 58–61. [Google Scholar] [CrossRef]
  50. Mafie, E.; Rupa, F.H.; Setsuda, A.; Saito-Ito, A.; Sato, H. Brief review on atypical human trypanosomiasis of Trypanosoma lewisi. Jpn. J. Vet. Res. 2016, 15, 24–33. [Google Scholar]
  51. Jain, P.; Goyal, V.K.; Agrawal, R. An atypical Trypanosoma lewisi infection in a 22-day-old neonate from India: An emergent zoonosis. Indian J. Pathol. Microbiol. 2023, 66, 199–201. [Google Scholar] [CrossRef]
  52. Desquesnes, M.; Yangtara, S.; Kunphukhieo, P.; Jittapalapong, S.; Herder, S. Zoonotic trypanosomes in Southeast Asia: Attempts to control Trypanosoma lewisi using human and animal trypanocidal drugs. Infect. Genet. Evol. 2016, 44, 514–521. [Google Scholar] [CrossRef] [PubMed]
  53. Deborggraeve, S.; Koffi, M.; Jamonneau, V.; Bonsu, F.A.; Queyson, R.; Simarro, P.P.; Herdewijn, P.; Büscher, P. Molecular analysis of archived blood slides reveals an atypical human Trypanosoma infection. Diagn. Microbiol. Infect. Dis. 2008, 61, 428–433. [Google Scholar] [CrossRef] [PubMed]
  54. Desquesnes, M.; Kamyingkird, K.; Yangtara, S.; Milocco, C.; Ravel, S.; Wang, M.H.; Lun, Z.R.; Morand, S.; Jittapalapong, S. Specific primers for PCR amplification of the ITS1 (ribosomal DNA) of Trypanosoma lewisi. Infect. Genet. Evol. 2011, 11, 1361–1367. [Google Scholar] [CrossRef]
  55. Hong, K.; Zhang, X.; Fusco, O.A.; Lan, Y.G.; Lun, Z.R.; Lai, D.H. PCR-based identification of Trypanosoma lewisi and Trypanosoma musculi using maxicircle kinetoplast DNA. Acta Trop. 2017, 171, 207–212. [Google Scholar] [CrossRef]
  56. Lima, L.; Da Silva, F.M.; Neves, L.; Attias, M.; Takata, C.S.; Campaner, M.; De Souza, W.; Hamilton, P.B.; Teixeira, M.M.G. Evolutionary insights from bat trypanosomes: Morphological, developmental and phylogenetic evidence of a new species, Trypanosoma (Schizotrypanum) erneyi sp. nov., in African bats closely related to Trypanosoma (Schizotrypanum) cruzi and allied species. Protist 2012, 163, 856–872. [Google Scholar] [CrossRef]
  57. Lima, L.; Espinosa-Álvarez, O.; Hamilton, P.B.; Neves, L.; Takata, C.S.; Campaner, M.; Attias, M.; De Souza, W.; Camargo, E.P.; Teixeira, M.M.G. Trypanosoma livingstonei: A new species from African bats supports the bat seeding hypothesis for the Trypanosoma cruzi clade. Parasites Vectors 2013, 6, 221. [Google Scholar] [CrossRef]
  58. Hamilton, P.B.; Adams, E.R.; Malele, I.I.; Gibson, W.C. A novel, high-throughput technique for species identification reveals a new species of tsetse-transmitted trypanosome related to the Trypanosoma brucei subgenus, Trypanozoon. Infect. Genet. Evol. 2008, 8, 26–33. [Google Scholar] [CrossRef]
  59. Hamilton, P.B.; Lewis, M.D.; Cruickshank, C.; Gaunt, M.W.; Yeo, M.; Llewellyn, M.S.; Silva, F.M.; Stevens, J.R.; Miles, M.A.; Teixeira, M.M.G. Identification and lineage genotyping of South American trypanosomes using fluorescent fragment length barcoding. Infect. Genet. Evol. 2011, 11, 44–51. [Google Scholar] [CrossRef]
  60. Vergara-Meza, J.G.; Brilhante, A.F.; Valente, V.d.C.; Villalba-Alemán, E.; Ortiz, P.A.; Cosmiro de Oliveira, S.; Rodrigues Cavalcante, M.; Julião, G.R.; Gonçalves, M.C.; Valente, A.S.; et al. Trypanosoma cruzi and Trypanosoma rangeli in Acre, Brazilian Amazonia: Coinfection and notable genetic diversity in an outbreak of orally acquired acute Chagas disease in a forest community, wild reservoirs, and vectors. Parasitologia 2022, 2, 350–365. [Google Scholar] [CrossRef]
  61. Swofford, D.L. PAUP: Phylogenetic Analysis Using Parsimony (and Other Methods), Version 4.0 Beta 10; Sinauer Associates: Sunderland, MA, USA, 2002.
  62. Stamatakis, A. RAxML Version 8: A Tool for Phylogenetic Analysis and Post-Analysis of Large Phylogenies. Bioinformatics 2014, 30, 1312–1313. [Google Scholar] [CrossRef]
  63. Jombart, T.; Ahmed, I. adegenet 1.3-1: New Tools for the Analysis of Genome-Wide SNP Data. Bioinformatics 2011, 27, 3070–3071. [Google Scholar] [CrossRef] [PubMed]
  64. Jombart, T. adegenet: A R Package for the Multivariate Analysis of Genetic Markers. Bioinformatics 2008, 24, 1403–1405. [Google Scholar] [CrossRef] [PubMed]
  65. Garcia, H.A.; Rodrigues, C.M.F.; Rodrigues, A.C.; Pereira, D.L.; Pereira, C.L.; Camargo, E.P.; Hamilton, P.B.; Teixeira, M.M.G. Remarkable Richness of Trypanosomes in Tsetse Flies (Glossina morsitans morsitans and Glossina pallidipes) from the Gorongosa National Park and Niassa National Reserve of Mozambique Revealed by Fluorescent Fragment Length Barcoding (FFLB). Infect. Genet. Evol. 2018, 63, 370–379. [Google Scholar] [CrossRef] [PubMed]
  66. Barros, J.H.; Lima, L.; Schubach, A.O.; Teixeira, M.M.G. Trypanosoma madeirae sp. n.: A species of the clade T. cruzi associated with the neotropical common vampire bat Desmodus rotundus. Int. J. Parasitol. Parasites Wildl. 2019, 8, 71–81. [Google Scholar] [CrossRef]
  67. Sato, H.; Osanai, A.; Kamiya, H.; Obara, Y.; Jiang, W.; Zhen, Q.; Chai, J.; Ito, M. Characterization of SSU and LSU rRNA genes of three Trypanosoma (Herpetosoma) grosi isolates maintained in Mongolian jirds. Parasitology 2005, 130, 157–167. [Google Scholar] [CrossRef]
  68. Kim, H.J.; Han, B.; Lee, H.; Ju, J.W.; Shin, H. Current Status of Trypanosoma grosi and Babesia microti in Small Mammals in the Republic of Korea. Animals 2024, 14, 989. [Google Scholar] [CrossRef]
  69. Maia da Silva, F.; Marcili, A.; Lima, L.; Cavazzana, M., Jr.; Ortiz, P.A.; Campaner, M.; Takeda, G.F.; Paiva, F.; Nunes, V.L.B.; Camargo, E.P.; et al. Trypanosoma rangeli isolates of bats from Central Brazil: Genotyping and phylogenetic analysis enable description of a new lineage using spliced-leader gene sequences. Acta Trop. 2008, 109, 199–207. [Google Scholar] [CrossRef]
  70. Maia da Silva, F.; Naiff, R.D.; Marcili, A.; Gordo, M.; D’Affonseca, J.A.; Naiff, M.F.; Franco, A.M.R.; Campaner, M.; Valente, V.; Valente, S.A.; et al. Infection rates and genotypes of Trypanosoma rangeli and Trypanosoma cruzi infecting free-ranging Saguinus bicolor (Callitrichidae), a critically endangered primate of the Amazon Rainforest. Acta Trop. 2008, 107, 168–173. [Google Scholar] [CrossRef]
  71. Molinari, J.; Moreno, S.A. The correct subgeneric name of Trypanosoma rangeli Tejera, 1920 (Euglenozoa: Trypanosomatidae), a human-infective endoparasite of neotropical mammals. Zootaxa. 2018, 4418, 98–100. [Google Scholar] [CrossRef]
  72. Lima, L.; Espinosa-Álvarez, O.; Pinto, C.M.; Cavazzana Jr, M.; Pavan, A.C.; Carranza, J.C.; Lim, B.K.; Campaner, M.; Takata, C.S.A.; Camargo, E.P.; et al. New insights into the evolution of the Trypanosoma cruzi clade provided by a new trypanosome species tightly linked to Neotropical Pteronotus bats and related to an Australian lineage of trypanosomes. Parasites Vectors 2015, 8, 1255. [Google Scholar] [CrossRef]
  73. Jittapalapong, S.; Inpankaew, T.; Sarataphan, N.; Herbreteau, V.; Hugot, J.P. Molecular detection of divergent trypanosomes among rodents of Thailand. Infect. Genet. Evol. 2008, 8, 445–449. [Google Scholar] [CrossRef] [PubMed]
  74. Pumhom, P.; Pognon, D.; Yangtara, S.; Thaprathorn, N.; Milocco, C.; Douangboupha, B.; Herder, S.; Chaval, Y.; Morand, S.; Jittapalapong, S.; et al. Molecular prevalence of Trypanosoma spp. in wild rodents of Southeast Asia: Influence of human settlement habitat. Epidemiol. Infect. 2014, 142, 1221–1230. [Google Scholar] [CrossRef] [PubMed]
  75. Salzer, J.S.; Pinto, C.M.; Grippi, D.C.; Williams-Newkirk, A.J.; Peterhans, J.K.; Rwego, I.B.; Carroll, D.S.; Gillespie, T.R. Impact of anthropogenic disturbance on native and invasive trypanosomes of rodents in forested Uganda. Ecohealth 2016, 13, 698–707. [Google Scholar] [CrossRef] [PubMed]
  76. Ellwanger, J.H.; Chies, J.A.B. Zoonotic spillover: Understanding basic aspects for better prevention. Genet. Mol. Biol. 2021, 44, e20200355. [Google Scholar] [CrossRef]
  77. Austen, J.M.; Barbosa, A.D. Diversity and Epidemiology of Bat Trypanosomes: A One Health Perspective. Pathogens 2021, 10, 1148. [Google Scholar] [CrossRef]
Zoonoticdis 04 00028 g001
Figure 1. Map of South America indicating Brazilian and Venezuelan capture sites of bats found harboring T. lewisi-like and T. musculi-like trypanosomes.
Figure 1. Map of South America indicating Brazilian and Venezuelan capture sites of bats found harboring T. lewisi-like and T. musculi-like trypanosomes.
Zoonoticdis 04 00028 g001
Zoonoticdis 04 00028 g002
Figure 2. Fluorescent Fragment Length Barcoding (FFLB) profile of trypanosomes of the subgenus Herpetosoma. x axis, size of fragment (bp); y axis, fluorescence intensity. Blue, green, light blue, and black lines: fluorochrome used for each molecular marker.
Figure 2. Fluorescent Fragment Length Barcoding (FFLB) profile of trypanosomes of the subgenus Herpetosoma. x axis, size of fragment (bp); y axis, fluorescence intensity. Blue, green, light blue, and black lines: fluorochrome used for each molecular marker.
Zoonoticdis 04 00028 g002
Zoonoticdis 04 00028 g003
Figure 3. (a) Trypanosomes of the subgenus Herpetosoma identified using FFLB in blood samples from 18 bat species across five families in Venezuela and Brazil. (b) Bats were found co-infected with up to five different Trypanosoma species: HEP: Herpetosoma, Neo: Neobat clade, TCM: T. c. marinkellei, TCZ: T. cruzi, TDI: T. dionisii, TRA: T. rangeli. The numbers in (a) (within bars) and (b) indicate the number of bats (nº bats) found infected by each parasite, either isolated or mixed in different combinations of trypanosomes.
Figure 3. (a) Trypanosomes of the subgenus Herpetosoma identified using FFLB in blood samples from 18 bat species across five families in Venezuela and Brazil. (b) Bats were found co-infected with up to five different Trypanosoma species: HEP: Herpetosoma, Neo: Neobat clade, TCM: T. c. marinkellei, TCZ: T. cruzi, TDI: T. dionisii, TRA: T. rangeli. The numbers in (a) (within bars) and (b) indicate the number of bats (nº bats) found infected by each parasite, either isolated or mixed in different combinations of trypanosomes.
Zoonoticdis 04 00028 g003
Zoonoticdis 04 00028 g004
Figure 4. Phylogenetic tree inferred by Maximum Likelihood (ML) based on SSU rRNA sequences from 73 trypanosomes of the subgenus Herpetosoma (clade T. lewisi) and species representative of other major clades/subgenera (collapsed) of Trypanosoma. The numbers at nodes refer to ML support values derived from 500 replicates.
Figure 4. Phylogenetic tree inferred by Maximum Likelihood (ML) based on SSU rRNA sequences from 73 trypanosomes of the subgenus Herpetosoma (clade T. lewisi) and species representative of other major clades/subgenera (collapsed) of Trypanosoma. The numbers at nodes refer to ML support values derived from 500 replicates.
Zoonoticdis 04 00028 g004
Zoonoticdis 04 00028 g005
Figure 5. Phylogenetic analysis of Herpetosoma trypanosomes found in bats, rodents, and primates positioned the bat trypanosomes (bold) into Group 2 (T. lewisi-like) and Group 3 (T. musculi-like) of the major subclade within Herpetosoma comprising T. lewisi (Group 1) and T. grosi (Group 4). The numbers at nodes refer to ML support values derived from 500 replicates.
Figure 5. Phylogenetic analysis of Herpetosoma trypanosomes found in bats, rodents, and primates positioned the bat trypanosomes (bold) into Group 2 (T. lewisi-like) and Group 3 (T. musculi-like) of the major subclade within Herpetosoma comprising T. lewisi (Group 1) and T. grosi (Group 4). The numbers at nodes refer to ML support values derived from 500 replicates.
Zoonoticdis 04 00028 g005
Zoonoticdis 04 00028 g006
Figure 6. Single Nucleotide Polymorphisms (SNP) in the variable regions of SSU rRNA gene sequences of T. lewisi, T. lewisi-like, T. musculi, and other species of the subgenus Herpetosoma.
Figure 6. Single Nucleotide Polymorphisms (SNP) in the variable regions of SSU rRNA gene sequences of T. lewisi, T. lewisi-like, T. musculi, and other species of the subgenus Herpetosoma.
Zoonoticdis 04 00028 g006
Table 1. Host species and geographic origins of Trypanosoma (Herpetosoma) spp. from Venezuelan and Brazilian bats and their closest related trypanosomes included in molecular analyses.
Table 1. Host species and geographic origins of Trypanosoma (Herpetosoma) spp. from Venezuelan and Brazilian bats and their closest related trypanosomes included in molecular analyses.
GroupSpeciesIsolate CodeOrder/FamilyHost Origin SpeciesGeographic Origin
Group 1T. lewisiMolteno B3Rodentia/MuridaeRattus sp.UK
TryBiIDN203Rodentia/MuridaeBandicota indicaID
ATCC30085Rodentia/MuridaeRattus norvegicusUS
ROVE61Rodentia/MuridaeRattus norvegicusVE
AF05b_KE836Rodentia/MuridaeMus tritonKE
AF05c_ETH1492Rodentia/MuridaeStenocephalemys albipesET
AF05g_KE675Rodentia/MuridaeLemniscomys striatusKE
AF05e_RS0805Rodentia/MuridaePraomys minorZB
AF05a_KE670Rodentia/MuridaePraomys jacksoniKE
AF05f_TA190Rodentia/MuridaePraomys jacksoniTZ
AF05i_KE452Rodentia/MuridaeMastomys natalensisKE
Group 2T. lewisi-like 4470Rodentia/MuridaeRattus RattusBR
M11871Rodentia/MuridaeRattus RattusBR
M11872Rodentia/MuridaeRattus RattusBR
GORO52Rodentia/MuridaeRattus RattusMZ
GORO53Rodentia/MuridaeRattus RattusMZ
1148Rodentia/MuridaeRattus norvegicusBR
M11227Primates/AotidaeAotus sp.BR
95Primates/CebidaeCallithrix jacchusBR
AfPrimates/AtelidaeAlouatta fuscaBR
MOVE16Chiroptera/NoctilionidaeNoctilio albiventrisVE
MOVE44Chiroptera/PhyllostomidaeCarollia perspicillattaVE
MOVE46Chiroptera/PhyllostomidaeGlossophaga soricinaVE
MOVE49Chiroptera/PhyllostomidaeGlossophaga soricinaVE
Group 3T. musculiLUM343Rodentia/MuridaeMus musculusDE
T. rabinowitschaeLV422Rodentia/CricetidaeCricetus cricetusFR
T. blanchardiLV421Rodentia/CricetidaeEliomys quercinusFR
T. niviventeraeTryNcCHN503Rodentia/MuridaeNiviventer confucianusCN
T. lewisiWC365Rodentia/MuridaeRattus loseaCN
Trypanosoma sp.BRA3Rodentia/MuridaeRattus fuscipesAU
Trypanosoma sp.BRA1Rodentia/MuridaeRattus fuscipesAU
Trypanosoma sp.AF05d_TZ28179Rodentia/MuridaeMus minutoidesTZ
Trypanosoma sp.RNMO66Chiroptera/PhyllostomidaeTrachops cirrhosusBR
Trypanosoma sp.RNMO72Chiroptera/PhyllostomidaeTrachops cirrhosusBR
Trypanosoma sp.RNMO81Chiroptera/PhyllostomidaeDiphylla ecaudataBR
Geographic origin countries: AU: Australia, BR: Brazil, CN: China, DE: Germany, ET: Ethiopia, FR: France, ID: Indonesia, KE: Kenya, MZ: Mozambique, TZ: Tanzania, UK: United Kingdom, US: United States of America, VE: Venezuela, VN: Vietnam, ZB: Zambia.
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

Share and Cite

MDPI and ACS Style

Villalba-Alemán, E.; Lima, L.; Ortiz, P.A.; Fermino, B.R.; Grisante, G.E.; Rodrigues, C.M.F.; Úngari, L.P.; Añez, N.; Garcia, H.A.; Teixeira, M.M.G. Spillover of Trypanosoma lewisi and Trypanosoma musculi Allied Trypanosomes from Rodents to Bats in the Roofs of Human Dwellings: Synanthropic Bats as a Potential New Source of Human Opportunistic Trypanosomes. Zoonotic Dis. 2024, 4, 320-336. https://doi.org/10.3390/zoonoticdis4040028

AMA Style

Villalba-Alemán E, Lima L, Ortiz PA, Fermino BR, Grisante GE, Rodrigues CMF, Úngari LP, Añez N, Garcia HA, Teixeira MMG. Spillover of Trypanosoma lewisi and Trypanosoma musculi Allied Trypanosomes from Rodents to Bats in the Roofs of Human Dwellings: Synanthropic Bats as a Potential New Source of Human Opportunistic Trypanosomes. Zoonotic Diseases. 2024; 4(4):320-336. https://doi.org/10.3390/zoonoticdis4040028

Chicago/Turabian Style

Villalba-Alemán, Evaristo, Luciana Lima, Paola Andrea Ortiz, Bruno Rafael Fermino, Gladys Elena Grisante, Carla Monadeli Filgueira Rodrigues, Letícia Pereira Úngari, Néstor Añez, Herakles Antonio Garcia, and Marta Maria Geraldes Teixeira. 2024. "Spillover of Trypanosoma lewisi and Trypanosoma musculi Allied Trypanosomes from Rodents to Bats in the Roofs of Human Dwellings: Synanthropic Bats as a Potential New Source of Human Opportunistic Trypanosomes" Zoonotic Diseases 4, no. 4: 320-336. https://doi.org/10.3390/zoonoticdis4040028

APA Style

Villalba-Alemán, E., Lima, L., Ortiz, P. A., Fermino, B. R., Grisante, G. E., Rodrigues, C. M. F., Úngari, L. P., Añez, N., Garcia, H. A., & Teixeira, M. M. G. (2024). Spillover of Trypanosoma lewisi and Trypanosoma musculi Allied Trypanosomes from Rodents to Bats in the Roofs of Human Dwellings: Synanthropic Bats as a Potential New Source of Human Opportunistic Trypanosomes. Zoonotic Diseases, 4(4), 320-336. https://doi.org/10.3390/zoonoticdis4040028

Article Metrics

Back to TopTop