Study on the relation of the characteristics of the capture sites with the Leptospira spp. occurrence in bats and rodents from Yucatan, Mexico.

This study aims to describe the natural Leptospira occurrence in small mammals from Yucatan, Mexico, and to explore the relation between the characteristics of the capture sites and the Leptospira occurrence. Bats and rodents were captured in five sites of Yucatan state, and from them, a kidney fragment was collected that was used in the genomic DNA extraction. Leptospira DNA was identified by PCR targeting the 16S-rRNA and LipL32 genes. Additionally, a bioinformatic analysis was carried out to know the Leptospira species and was corroborated with a phylogenetic tree. The assemblage of small mammals was compound of 82 (51.2 %) bats and 78 (48.8 %) rodents. A global frequency (bats plus rodents) of Leptospira occurrence of 21.2 % (34/160) was observed; in bats, it was 21.9 % (18/82), and in rodents, 20.5 % (16/78). The phylogenetic trees based on LipL32 gene showed that the recovered sequences most closely resemble the species L. borgpetersenii and L. noguchii. The ordination of the capture sites with tropical deciduous forests as original vegetation is more related to the abundance of Leptospira-infected rodents. The ordination of the capture sites with tropical sub-deciduous forests as original vegetation is more related to the diversity of Leptospira-infected bat species. The canonical ordering of the capture sites is by the original vegetation type and the diversity and abundance of Leptospira-infected bat and rodent species.

Unraveling the diversity of Trypanosoma species from Central Mexico: Molecular confirmation on the presence of Trypanosoma dionisii and novel Neobat linages.

Bats are one of the groups of mammals with the highest number of associated Trypanosoma taxa. There are 50 Trypanosoma species and genotypes infecting more than 75 species of bats across five continents. However, in Mexico, the inventory of species of the genus Trypanosoma associated with bats is limited to only two species (Trypanosoma vespertilionis and Trypanosoma cruzi) even though 140 species of bats inhabit this country. Specifically, 91 bat species have been recorded in the state of Veracruz, but records of trypanosomatids associated with this mammalian group are absent. Due to the complex Trypanosoma-bat relationship, the high diversity of bat species in Veracruz, as well as the lack of records of trypanosomatids associated with bats for this state, the aim of this work was to analyze the diversity of species of the genus Trypanosoma and their presence from a bat community in the central area of the state of Veracruz, Mexico. During the period of January to August 2022 in the Tequecholapa Environmental Management Unit where bats were collected using mist nets and blood samples were obtained from their thumbs. We extracted genetic material and amplified a fragment of 800 bp of the 18S ribosomal gene of the genus Trypanosoma by conventional PCR. The positive amplicons were sequenced, and phylogenetic reconstruction was performed to identify the parasite species. A total of 285 bats (149♀, 136♂) belonging to 13 species from 10 genera and a single family (Phyllostomidae) were collected. Twenty-three specimens from six species tested positive for the presence of Trypanosoma dionisii, Trypanosoma sp. Neobat 4, and a potential novelty species provisionally named as Trypanosoma sp. Neobat 6. The results of the present work increase the number of species of the genus Trypanosoma infecting bats in Mexico and in the Neotropical region.

Population genetics of the Ixodes affinis (Ixodida: Ixodidae) complex in America: new findings and a host-parasite review.

Ticks are hematophagous ectoparasites associated with a wide range of vertebrate hosts. Within this group, the Ixodidae family stands out, in which the Ixodes genus contains at least 245 species worldwide, from which 55 species are present in the Neotropical region. Ixodes affinis, a tick described in 1899, has a wide distribution from the Southern Cone of America to the United States. However, since its description, morphological variability has been reported among its populations. Furthermore, attempts have been made to clarify its status as a species complex using mitochondrial markers, but mainly in restricted populations of South and Central America. Thus, information related to populations of the transition region between the Neotropical and Nearctic zones is lacking. For these reasons, the objectives of the study were to evaluate the genetic diversity and structure of I. affinis across the Americas and to compile all the published records of I. affinis in America, to elucidate the host-parasite relationships and to identify their geographical distribution. For this, a phylogeny, and AMOVA analyses were performed to assess the genetic structure of samples obtained by field work in South Carolina, USA and Yucatán, Mexico. A total of 86 sequences were retrieved from a fragment of the 16S region. Phylogeny and genetic structure analysis showed four groups that were geographically and genetically related with high branch support and Fst values, all of them statistically significant. The results obtained support the hypothesis that I. affinis it corresponds to a complex of four species, which must be validated through future morphological comparisons.

DNA barcoding and new records of Ornithodoros yumatensis from Central Mexico.

Bats represent the second order of mammals with the highest number of species worldwide with over 1,616 species, and almost 10% of them are recorded in Mexico. These mammals have a great diversity of ectoparasites, in particular soft ticks of the genus Ornithodoros. Desmodus rotundus is one of the bat species that has scarcely been studied in terms of tick species richness in Mexico, with three tick species reported in five of the 32 Mexican states. For this reason, the aim of the present work was to identify ticks associated with D. rotundus from Central Mexico. Fieldwork was undertaken in the municipality El Marqués, Ejido Atongo A, Querétaro, Mexico. Bats were captured using mist nets and were visually inspected for tick presence. The ectoparasites were identified morphologically and molecularly with the use of mitochondrial markers 16SrDNA and cytochrome oxidase subunit I (COI). A total of 30 D. rotundus (1 female, 29 males) were captured, from which 20 larvae identified as Ornithodoros yumatensis were recovered. Molecular analysis confirmed the presence of this species with identity values of 99-100% with sequences of this species from the southwestern US, and the Yucatán Peninsula, Mexico. This is the first report of ticks associated with bats for the state of Querétaro, providing the first sequences of the COI gene from Mexican populations of O. yumatensis and shows an increase in the distribution of this soft tick across Central Mexico.

The Low Variability of Tc24 in Trypanosoma cruzi TcI as an Advantage for Chagas Disease Prophylaxis and Diagnosis in Mexico.

(1) Background: Chagas disease is the main neglected tropical disease in America. It is estimated that around 6 million people are currently infected with the parasite in Latin America, and 25 million live in endemic areas with active transmission. The disease causes an estimated economic loss of USD 24 billion dollars annually, with a loss of 75,200 working years per year of life; it is responsible for around ~12,000 deaths annually. Although Mexico is an endemic country that recorded 10,186 new cases of Chagas disease during the period of 1990-2017, few studies have evaluated the genetic diversity of genes that could be involved in the prophylaxis and/or diagnosis of the parasite. One of the possible candidates proposed as a vaccine target is the 24 kDa trypomastigote excretory-secretory protein, Tc24, whose protection is linked to the stimulation of T. cruzi-specific CD8+ immune responses. (2) Methods: The aim of the present study was to evaluate the fine-scale genetic diversity and structure of Tc24 in T. cruzi isolates from Mexico, and to compare them with other populations reported in the Americas with the aim to reconsider the potential role of Tc24 as a key candidate for the prophylaxis and improvement of the diagnosis of Chagas disease in Mexico. (3) Results: Of the 25 Mexican isolates analysed, 48% (12) were recovered from humans and 24% (6) recovered from Triatoma barberi and Triatoma dimidiata. Phylogenetic inferences revealed a polytomy in the T. cruzi clade with two defined subgroups, one formed by all sequences of the DTU I and the other formed by DTU II-VI; both subgroups had high branch support. Genetic population analysis detected a single (monomorphic) haplotype of TcI throughout the entire distribution across both Mexico and South America. This information was supported by Nei's pairwise distances, where the sequences of TcI showed no genetic differences. (4) Conclusions: Given that both previous studies and the findings of the present work confirmed that TcI is the only genotype detected from human isolates obtained from various states of Mexico, and that there is no significant genetic variability in any of them, it is possible to propose the development of in silico strategies for the production of antigens that optimise the diagnosis of Chagas disease, such as quantitative ELISA methods that use this region of Tc24.

Identification of the G184C, C190A and T2134A mutations in the para-sodium channel gene of the southern cattle tick Rhipicephalus (Boophilus) microplus associated with resistance to cypermethrin in northern Veracruz, Mexico.

The southern cattle tick (Rhipicephalus microplus) represents one of the ectoparasites with the greatest distribution worldwide. Infestations by this arthropod can cause a decrease in the production of meat and milk, as well as anaemia and the transmission of bacterial and parasitic agents. For this reason, several active molecules have been developed to control these arthropods. A widely used group of ixodicides are pyrethroids, especially cypermethrin, which have knockdown effects on ticks. Resistance to cypermethrin has been reported in ticks since the 2000s; it was registered for the first time in Mexico in 2009. Even though multiple studies have evaluated resistance with conventional tests, there are few studies in Mexico that have identified the presence of single nucleotide polymorphisms (SNPs) associated with resistance. Hence, the aim of this work was to monitor three mutations associated with resistance in the sodium/chlorine channel in eight populations of ticks from northern Veracruz. Engorged adult females were collected from which genomic DNA was extracted. Subsequently, three mutations in domains II and III of para­sodium channel gene were detected by conventional PCR and sequencing. Global alignments were done with the reference sequences deposited in GenBank. A total of 116 engorged females were analysed, of which 10 tested positive for G184C and C190A of domain II of the para­sodium channel gene. T2134A was present in domain III in a single production unit. This is the first work where molecular monitoring of cypermethrin resistance has been carried out in the northern zone of the state of Veracruz.

Confirmation of the presence of Rickettsia felis infecting Ornithodoros puertoricensis in Mexico.

Soft ticks are neglected competent vectors of a wide range of pathogenic microorganisms, among which bacteria of the genera Rickettsia and Borrelia stand out. In Mexico, previous studies have shown the presence of a member of the Ornithodoros talaje complex in the Virginia opossum (Didelphis virginiana Didelphimorphia: Didelphidae Kerr) from southeastern Mexico. However, its specific identification has not been achieved. Two D. virginiana were treated in a private clinic during the period of April-May 2022. Tick larvae were manually removed, DNA extraction was performed, and some genes from various bacterial and parasitic pathogens were amplified and sequenced. A total of 96 larvae were recovered, which were morphologically identified as Ornithodoros puertoricensis (Ixodida: Argasidae Fox); the 16 S sequences showed a similarity of 96.79%-99.51% with sequences of O. puertoricensis from Panama and Colombia. The presence of Rickettsia felis (Rickettsiales: Rickettsiaceae Bouyer et al.) was detected in 15 specimens from one host. The soft tick O. puertoricensis is recorded for the first time as an ectoparasite of the Virginia opossum in America and represents the second report for this soft tick in Mexico since 1963. This represents the most northern record of this tick species in its geographic distribution and brings a new soft tick-Rickettsia association.

Is Psathyromyia shannoni (Diptera: Psychodidae: Phlebotominae) a species complex? Retrospective study of genetic diversity of COI gene, pathogens and geographic distribution.

The sand fly Psathyromyia shannoni is a broadly distributed species that is relevant for the transmission of pathogens such as Leishmania, Bartonella and viruses in several countries of America. This species belongs to the Shannoni complex. Yet its identification is difficult due to morphologic intraspecific polymorphisms that make it difficult to distinguish between species, and could therefore lead to misidentification and overestimation of its distribution. The aim of this study was to perform a retrospective study on the genetic diversity of Pa. shannoni based on the Cytochrome Oxidase subunit 1 gene and considering its geographic distribution to achieve a better identification and differentiation from other species of the Shannoni complex. According to the Maximum Likelihood analysis and the data on the genetic structure, we propose a modified delimitation of Pa. shannoni species by classifying it into at least three genetic lineages, based on genetic variability and distribution. However, more genetic information on the COI gene, mainly from countries where this species has been reported, is needed to strengthen this proposal.

Epidemiological Study of the Occurrence of Typhus Group Rickettsia Natural Infection in Domiciliated Dogs from a Rural Community in South-Eastern Mexico.

The aim is to describe the Typhus group (TG) Rickettsia infection in dogs and to identify factors associated with this infection. We collected blood samples and gathered exposure and clinical data of 142 dogs from a rural community of Yucatan. The Rickettsia group was determined by semi-nested PCR. Generalized linear models with binomial error distribution were used to model the associated factors from the dog sample for risk ratio (RR) estimation. Thirty-four dogs (23.9%) showed molecular evidence of TG Rickettsia DNA. The multivariate model showed that mixed-breed dogs (RR = 0.06) and dogs that had received antiparasitic treatment (RR = 0.049) had a lower risk of getting infected, taking as reference the purebred group and the non-treated dogs, respectively. Looking at variable interactions, adult dogs without outdoor activities had a lower infection risk than puppies (RR = 0.26). Among dogs with antiparasitic treatment, females had a higher infection risk than male dogs (RR = 26.2). The results showed enzootic TG Rickettsia circulation in dogs of a rural community. The factors outdoor activities, age and previous antiparasitic treatment, as well as the clinical variables signs of hemorrhages and epistaxis, were associated with a less chance of natural infection in the studied dogs. Prevention and control of the enzootic transmission risk of TG Rickettsia should help to reduce the potential zoonotic transmission of this pathogen.

Molecular detection of Vibrio paraheamolyticus in a sperm whale (Physeter macrocephalus) stranding in northern Veracruz, Mexico.

On 25 August 2021, a single female sperm whale (Physeter macrocephalus) was found stranded dead in Playa Tuxpan, Veracruz, Mexico. Skin biopsies were obtained and screened for the detection of various potentially pathogenic bacterial genera, using conventional polymerase chain reaction and sequencing of the positive amplicons. We recorded, for the first time, the presence of Vibrio paraheamolyticus in skin samples from P. macrocephalus in the Gulf of Mexico. Additionally, we discuss 29 records reporting strandings of sperm whales from six states of the Mexican Republic. Most of the records are concentrated in the Pacific Ocean. Our findings increase the inventory of bacteria reported in P. macrocephalus worldwide, summarising the knowledge of stranding events in sperm whale populations in Mexico.

Is vertical transmission the only pathway for Rickettsia felis?

The genus Rickettsia encompasses several species grouped into two main clusters, Typhus and the Transitional groups. The latter group contains Rickettsia felis, an endosymbiont of several arthropods with an uncertain human pathogenicity and whose most efficient transmission mechanism described thus far is transovarial. The aim of this study was to evaluate whether this pathway exists using phylogenetic analysis and partial sequences of the 17kDa and gltA genes and comparing them with host phylogeny using the cytb region. This is the first study that evaluates the vertical transmission of R. felis. In general, both phylogenies of R. felis showed no polytomies, as suspected if this pathway was the only pathway occurring. When phylogenies of the invertebrates and the gltA of R. felis were compared for strong coevolutionary insight, intricate relationships were observed, suggesting that other transmission pathways must occur, such as horizontal transmission. Further studies are needed to determine which other transmission routes occur in hematophagous arthropods.

Molecular confirmation of Rickettsia parkeri in humans from Southern Mexico.

Human tick-borne rickettsioses are caused by Rickettsia africae, Rickettsia massiliae, Rickettsia parkeri, Rickettsia philipi and Rickettsia rickettsii in the American continent. Rickettsia parkeri represents the second most important agent of eschar-associated spotted fever throughout the American continent. In Mexico, the presence of three strains of this bacterium (Black Gap, Maculatum and Atlantic Rainforest) has recently been reported in four species of ticks from five states (Baja California, Campeche, Sonora, Tabasco and Veracruz) and rickettsaemia was demonstrated in a dog from the state of Yucatán. We report two human cases of R. parkeri-rickettsiosis from Yucatan, Mexico. These findings have increased the interest in R. parkeri surveillance in the human population, due to the fluctuating regional Spotted Fever morbidity and mortality in the country.

Rhipicephalus sanguineus Complex in the Americas: Systematic, Genetic Diversity, and Geographic Insights.

The Rhipicephalus sanguineus group encompasses at least 12 validated species of Palearctic and Afrotropical hard ticks, which are relevant in veterinary medicine and public health. The taxonomy of R. sanguineus s.s., has been particularly intensely debated, due to its wide geographic distribution, morphological variants, parasite-host associations, and its capacity and vectorial competence for the transmission of several pathogens. By sequencing mitochondrial markers, it was possible to identify the existence of multiple lineages, among which the Tropical and the Temperate lineages stand out, particularly in America. However, the northern limit between these lineages is not clear due to the lack of extensive sampling across Mexico. For this reason, the aim of the present study was to determine the genetic diversity and structure of the R. sanguineus group in Mexico and to compare it with the populations reported in the Americas, in order to propose the northern limit of the R. sanguineus Tropical lineage and the potential regions of sympatry with R. sanguineus s.s. The findings of this study now confirm the presence of R. sanguineus s.s. in Mexico, showing a subtle genetic structure and high genetic diversity throughout its distribution in the Americas. In contrast, the Tropical lineage seems to be genetically less diverse in its overall distribution in the Americas. The genetic diversity of these two independent lineages could have important epidemiological implications in the transmission of tick pathogens.

Rediscovering an old friend: the case of Ixodes dampfi in a protected natural reserve in central Mexico.

The tick Ixodes dampfi Cooley 1943, a member of the Pholeoixodes subgenus, was first described from gophers (Geomys sp.) collected in the state of Mexico. However, information on the origin and parasite-host association of I. dampfi is lacking. Here, new records of the occurrence of this tick species were provided nearly 80 years since its original description, in addition to new localities, genetic data, and host-parasite records. A total of four hosts (one Bassariscus astutus, two Didelphis virginiana, and one Peromyscus gratus) from three orders (Carnivora, Didelphimorphia, and Rodentia) were parasitised by seven I. dampfi females. Sequences of the 16S rDNA gene from the ticks exhibited a similarity ranging from 97 (389/403 bp) to 100% (403/403 bp), with the unique sequence of I. dampfi available on GenBank (AF549837). Additionally, Rickettsia-specific 16S rDNA, htrA, and ompA gene assays generated sequences in four of the seven I. dampfi specimens (57.14%). Partial sequences revealed 99-100% genetic identities with Candidatus Rickettsia angustus (GenBank accession HF935069, HF935072, HF935078) and Candidatus Rickettsia kingi (GenBank accession HF935068, HF935071, HF935077) from Canada. Our results represent the first record of this elusive ectoparasite in a natural protected area, which is a triumph for conservation, as accelerated defaunation processes pose the risk of coextinction for many groups of vertebrates and their ectoparasites.

Molecular detection of Rickettsia sp. cf. Rickettsia monacensis in Ixodes sp. cf. Ixodes affinis collected from white-tailed deer in Campeche, Mexico.

Deer encompass a group of large-sized vertebrates that serve as hosts for a wide variety of ectoparasites, mainly ticks. In Mexico, ticks have relevance as vectors of pathogenic microorganisms, and 20 species of hard ticks are associated with four species of deer, although only a single study has been conducted to detect bacterial agents associated with ticks from deer in the country. In February, 2019 three white-tailed deers (Odocoileus virginianus) were hunted from the locality of Chiná from the municipality of Campeche, Mexico. The sampled deers were parasitized by 26 ticks belonged to three species: Amblyomma mixtum (5♀, 1♂), Amblyomma ovale (2♀, 1♂), and Ixodes sp. cf. Ixodes affinis (15♀, 2♂). Specimens were screened individually for Anaplasma, Borrelia, Ehrlichia, and Rickettsia DNA by the amplification of several fragments of 16S rRNA, gltA, 17-kDa, and flaB genes. This study report for the first time the presence of Rickettsia sp. cf. Rickettsia monacensis in Ixodes sp. cf. Ixodes affinis in Mexico.

Rickettsial agents detected in the genus Psathyromyia (Diptera:Phlebotominae) from a Biosphere Reserve of Veracruz, Mexico.

Phlebotomine sand flies are considered the main vectors of Leishmania, the causal agents of leishmaniasis, which is a serious emerging public health problem worldwide. The use of biological control alternatives, like endosymbiotic bacteria (Wolbachia and Rickettsia), have been proposed to decrease sand fly populations and reduce Leishmania transmissions, yet only few records on the detection of Wolbachia or Rickettsia in sand flies are available worldwide. The aim of this study was to perform the molecular detection of Rickettsial agents associated with sand flies from the last patch of a rainforest in south-eastern Mexico, where a high prevalence of Leishmania infantum has been reported. Sampling effort of sand flies covered 300 trap-nights between 2011 and 2013, and a total of 925 specimens from twelve species were morphologically identified. Using PCR techniques, we identified a new lineage of the endosymbionts Rickettsia in Psathyromyia aclydifera (prevalence of 19.54%), and Wolbachia in Psathyromyia shannoni and Lutzomyia sp. (prevalence of 25%). The detected Wolbachia lineage was similar to the wWhi strain found in Pa. shannoni from Colombia and Nyssomyia whitmani from Brazil; whereas the identified Rickettsia represents a new lineage worldwide. This is the first record of Rickettsial agents associated to sand flies from this region, yet it remains for analysed if these bacteria possibly play a role as vector control agents, capable of reducing the sand fly populations in Mexico.

Borrelia in neotropical bats: Detection of two new phylogenetic lineages.

The genus Borrelia encompasses 50 spirochetal species, several of which are pathogenic and have been detected in a wide range of mammals, especially rodents and cervids. Although the order Chiroptera is the second most diverse mammalian order, and borreliosis represents a human and veterinary health problem in endemic countries, few studies have previously reported infections of Borrelia in these flying mammals. For this reason, the aim of the present study was to detect the presence of, and to analyze the diversity of Borrelia species in several bat species from Mexico. A total of 69 bats belonging to 11 species were collected and molecular detection of Borrelia was performed by amplifying three genes using specific primers. Only five individuals of four bat species (Saccopteryxbilineata, Choeroniscus godmani, Sturnira parvidens and Lasiurus cinereus) tested positive for Borrelia DNA. We now show the first Borrelia record in Mexican bats from two different ecosystems, where previously several potential vector species of the genus Ixodes and Ornithodoros had been reported. The Borrelia sequences obtained from the bats revealed two new putative lineages, one from the relapsing fever group and the second one belonging to the Borrelia burgdorferi s.l. complex, both of which are related to zoonotic species. These results highlight the importance of bats as potential hosts of Borrelia, and the imperative need of active surveillance in flying mammals in order to understand their potential role in the life cycle of this bacteria genus.

The genus Rickettsia in Mexico: Current knowledge and perspectives.

The genus Rickettsia encompasses 35 valid species of intracellular, coccobacilli bacteria that can infect several eukaryotic taxa, causing multiple emerging and re-emerging diseases worldwide. This work aimed to gather and summarise the current knowledge about the genus Rickettsia in Mexico, updating the taxonomy of the bacteria and their hosts by including all the records available until 2020, to elucidate host-parasite relationships and determine the geographical distribution of each Rickettsia species present in the country. Until now, 14 species of Rickettsia belonging to four groups have been recorded in Mexico. These species have been associated with 26 arthropod species (14 hard ticks, three soft ticks, two sucking lice, and seven fleas) and 17 mammal species distributed over 30 states in Mexico. This work highlights the high biological inventory of rickettsias for Mexico and reinforces the need to approach the study of this group from a One Health perspective.