Multilayer Networks Assisting to Untangle Direct and Indirect Pathogen Transmission in Bats.

The importance of species that connect the different types of interactions is becoming increasingly recognized, and this role may be related to specific attributes of these species. Multilayer networks have two or more layers, which represent different types of interactions, for example, between different parasites and hosts that are nonetheless connected. The understanding of the ecological relationship between bats, ectoparasites, and vector-borne bacteria could shed some light on the complex transmission cycles of these pathogens. In this study, we investigated a multilayer network in Brazil formed by interactions between bat-bacteria, bat-ectoparasite, and ectoparasite-bacteria, and asked how these interactions overlap considering different groups and transmission modes. The multilayer network was composed of 31 nodes (12 bat species, 14 ectoparasite species, and five bacteria genera) and 334 links, distributed over three layers. The multilayer network has low modularity and shows a core-periphery organization, that is, composed of a few generalist species with many interactions and many specialist species participating in few interactions in the multilayer network. The three layers were needed to accurately describe the multilayer structure, while aggregation leads to loss of information. Our findings also demonstrated that the multilayer network is influenced by a specific set of species that can easily be connected to the behavior, life cycle, and type of existing interactions of these species. Four bat species (Artibeus lituratus, A. planirostris, Phyllostomus discolor, and Platyrrhinus lineatus), one ectoparasite species (Steatonyssus) and three bacteria genera (Ehrlichia, hemotropic Mycoplasma and Neorickettsia) are the most important species for the multilayer network structure. Finally, our study brings an ecological perspective under a multilayer network approach on the interactions between bats, ectoparasites, and pathogens. By using a multilayer approach (different types of interactions), it was possible to better understand these different ecological interactions and how they affect each other, advancing our knowledge on the role of bats and ectoparasites as potential pathogen vectors and reservoirs, as well as the modes of transmission of these pathogens.

Single Cell Analysis Reveals a New Microsporidia-Host Association in a Freshwater Lake.

Microsporidia are a large group of obligate intracellular eukaryotic parasites. Recent studies suggest that their diversity can be huge in freshwater lake ecosystems especially in the < 150-µm size fraction. However, little is known about their hosts and therefore their impact on the trophic food web functioning. In this study, single cell analysis and fluorescence microscopy were used to detect new host-parasite association within rotifer communities in lake Aydat (France). Our analysis showed the existence of a potential new species belonging to the Crispospora genus able of infecting the rotifer Kellicottia with a high prevalence (42.5%) suggesting that Microsporidia could have a great impact on the rotifer populations' regulation in lakes.

Co-occurrence of Eutrombicula alfreddugesi and Oswaldofilaria chabaudi in Tropidurus torquatus and first report of microfilariae in the chigger mite: possible evidence of a lifecycle pathway?

While much attention has been paid to vector-borne filariasis, diseases that threaten millions of people in tropical and subtropical countries, the literature on host-parasite associations and transmission strategies of filarial nematodes in wildlife is scarce. Here, we report the co-occurrence of chigger mites (Eutrombicula alfreddugesi) and onchocercid nematodes (Oswaldofilaria chabaudi) parasitizing the lizard Tropidurus torquatus in the State of Minas Gerais, Brazil. Examination of chiggers established, for the first time, the occurrence of microfilariae in trombiculid mites (Trombiculidae). These larvae were morphologically similar to those recovered from adult females of O. chabaudi. The current evidence suggests that chiggers do not play a role in the transmission of filarioid nematodes, but rather act as accidental or dead-end hosts. Nevertheless, considering the polyphagous nature of trombiculid mites, similar to blood-sucking insects involved in the transmission of several infectious diseases, further studies may shed light on the potential role of chiggers as vectors of filarioids.

Molecular detection of Rickettsia species and host associations of Laelaps mites (Acari: Laelapidae) in Taiwan.

Various rickettsiae have recently been detected in Laelaps mites (Acari: Laelapidae), which are common ectoparasites of rodents; however, investigations on this topic remain very scarce, particularly in Asia. In the present study, shrews and rodents were trapped from 2006 to 2010 in eight lowland regions of Taiwan (< 500 m in elevation) to collect associated Laelaps mites, from which Rickettsia-a group of emerging pathogens-were detected and identified by assaying the gltA and ompB genes. A total of 853 Laelaps mites of at least four species were collected from a sample of 1004 small mammals that included one shrew and 10 rodent species. Rattus losea was the most common species (44.9% of total hosts) and hosted the highest percentage of mites (76.6% of total mites). Laelaps nuttalli was the most abundant mite species (51.7% of total mites), followed by Laelaps echidninus (24.2%), Laelaps sedlaceki (23.1%), and Laelaps myonyssognathus (0.2%). Notably, Rickettsia species with the highest similarity to spotted fever group (SFG) rickettsiae were identified from seven of the 72 pools of Laelaps mites. The presence of SFG rickettsiae in hematophagous Laelaps mites, particularly including species that are closely associated with commensal rodents in frequent contact with humans, calls for further investigation on the competence of Laelaps mites in transmitting rickettsiae.

Ultrastructure, phylogeny and histopathology of two novel haplosporidians parasitising amphipods, and importance of crustaceans as hosts.

This study provides morphological, ultrastructural and phylogenetic characterization of 2 novel species of Haplosporidia (Haplosporidium echinogammari n. sp. and H. orchestiae n. sp.) infecting amphipods of the genera Echinogammarus and Orchestia collected in southwestern England. Both parasites infect the connective tissues associated with the digestive gland and the tegument, and eventually infect other organs causing disruption of host tissues with associated motor impairment and fitness reduction. Prevalence of infection varied with host species, provenance and season, being as high as 75% for individuals of E. marinus infected with H. echinogammari in June (n = 50). Although no spores were found in any of the infected amphipods examined (n = 82), the morphology of monokaryotic and dikaryotic unicellular stages of the parasites enabled differentiation between the 2 new species. Phylogenetic analysis of the new species based on the small subunit (SSU) rDNA gene placed H. echinogammari close to H. diporeiae in haplosporidian lineage C, and H. orchestiae in a novel branch within Haplosporidium. Genetic diversity of the haplosporidians infecting these and other amphipod species was evaluated and compared to morphological and ultrastructural changes to host tissues. The phylogenetic relationship of haplosporidian infections in other crustacean hosts is discussed after inclusion into the analysis of 25 novel SSU rDNA sequences obtained from crabs, isopods and crayfish.

Ticks infesting bats (Mammalia: Chiroptera) in the Brazilian Pantanal.

Ticks associated with bats have been poorly documented in the Neotropical Zoogeographical Region. In this study, a total of 1028 bats were sampled for tick infestations in the southern portion of the Brazilian Pantanal. A total of 368 ticks, morphologically identified as Ornithodoros hasei (n = 364) and O. mimon (n = 4), were collected from the following bat species: Artibeus planirostris, Platyrrhinus lineatus, Phyllostomus hastatus, Mimon crenulatum and Noctilio albiventris. Morphological identification of O. hasei was confirmed by molecular analysis. Regarding the most abundant bat species, only 40 (6.2%) out of 650 A. planirostris were infested by O. hasei, with a mean intensity of 7.2 ticks per infested bat, or a mean abundance of 0.44 ticks per sampled bat. Noteworthy, one single P. hastatus was infested by 55 O. hasei larvae, in contrast to the 2.5-7.2 range of mean intensity values for the whole study. As a complement to the present study, a total of 8 museum bat specimens (6 Noctilio albiventris and 2 N. leporinus), collected in the northern region of Pantanal, were examined for tick infestations. These bats contained 176 ticks, which were all morphologically identified as O. hasei larvae. Mean intensity of infestation was 22, with a range of 1-46 ticks per infested bat. Our results suggest that A. planirostris might play an important role in the natural life cycle of O. hasei in the Pantanal.

Liolaemus lizards (Squamata: Liolaemidae) as hosts for the nymph of Amblyomma parvitarsum (Acari: Ixodidae), with notes on Rickettsia infection.

Adults of Amblyomma parvitarsum are common ectoparasites of South American camelids of the genera Lama and Vicugna, occuring in highlands of Argentina, Bolivia, Chile, Peru and also in Argentinean Patagonia. Whereas larval stages of this tick are known to feed on small lizards, host records for the nymphal instar have remained unreported. Supported by morphological and molecular analyses, herein we report A. parvitarsum nymphs parasitizing two Liolaemus species (Reptilia: Squamata) in the Andean Plateau of Argentina and Chile. Additionally, by a PCR screening targetting gltA and ompA genes, DNA of Rickettsia was detected in one of the collected nymphs. Obtained sequences of this agent were identical to a recent Rickettsia sp. described infecting adults of this tick species in Chile and Argentina.

Superinfection reconciles host-parasite association and cross-species transmission.

Parasites are either dedicated to a narrow host range, or capable of exploiting a wide host range. Understanding how host ranges are determined is very important for public health, as well as wildlife, plant, livestock and agricultural diseases. Our current understanding of host-parasite associations hinges on co-evolution, which assumes evolved host preferences (host specialization) of the parasite. Despite the explanatory power of this framework, we have only a vague understanding of why many parasites routinely cross the host species' barrier. Here we introduce a simple model demonstrating how superinfection (in a heterogeneous community) can promote host-parasite association. Strikingly, the model illustrates that strong host-parasite association occurs in the absence of host specialization, while still permitting cross-species transmission. For decades, host specialization has been foundational in explaining the maintenance of distinct parasites/strains in host species. We argue that host specializations may be exaggerated, and can occur as a byproduct (not necessarily the cause) of host-parasite associations.

Assessment of Associations between Malaria Parasites and Avian Hosts-A Combination of Classic System and Modern Molecular Approach.

Avian malaria and related haemosporidian parasites are responsible for fitness loss and mortality in susceptible bird species. This group of globally distributed parasites has long been used as a classical system for investigating host-parasite associations. The association between a parasite and its hosts can be assessed by the prevalence in the host population and infection intensity in a host individual, which, respectively, reflect the ability of the parasite to infect the host and reproduce within the host. However, the latter has long been poorly investigated due to numerous challenges, such as lack of general molecular markers and limited sensitivity of traditional methods, especially when analysing naturally infected birds. The recent development of genetic databases, together with novel molecular methodologies, has shed light on this long-standing problem. Real-time quantitative PCR has enabled more accurate quantification of avian haemosporidian parasites, and digital droplet PCR further improved experimental sensitivity and repeatability of quantification. In recent decades, parallel studies have been carried out all over the world, providing great opportunities for exploring the adaptation of haemosporidian parasites to different hosts and the variations across time and space, and further investigating the coevolutionary history between parasites and their hosts. I hereby review the most important milestones in diagnosis techniques of avian haemosporidian parasites and illustrate how they provide new insights for understanding host-parasite associations.

Wide geographic distribution of overlooked parasites: Rare Microsporidia in Gammarus balcanicus, a species complex with a high rate of endemism.

Parasites and other symbionts deeply influence host organisms, and no living organism can be considered to have evolved independent of its symbionts. The first step towards understanding symbiotic influences upon host organisms is a strong supporting knowledge of parasite/symbiont diversity. Parasites of freshwater amphipods are diverse, with Microsporidia being a major group. These intracellular parasites impact gammarid fitness in different ways, ranging from reduced fitness to increased fecundity. Many Microsporidia have been recorded using molecular data, with multiple taxa pending formal taxonomic description. While some parasites are common, others are known only through sporadic records of single infections. In this study, we focus on rare/sporadic microsporidian infections within Gammarus balcanicus, a host species complex with a high level of endemism. In addition to enriching our knowledge on Microsporidia parasite diversity in amphipod hosts, we test whether these symbionts are specific to G. balcanicus or if they are the same taxa infecting other gammarid species. Of 2231 hosts from 87 sites, we catalogued 29 sequences of "rare" Microsporidia clustering into 19 haplogroups. These haplogroups cluster into 11 lineages: four pre-described taxa (Cucumispora roeselum, C. ornata, C. dikerogammari and Enterocytospora artemiae) and seven 'Molecular Operational Taxonomic Units', which are known from previously published studies to infect other European amphipod species. Our study significantly widens the geographic range of these Microsporidia and expands the known spectrum of hosts infected. Our results suggest that these parasites are ancient infections of European gammarids. For some host-parasite systems, we hypothesize that the common parasite ancestors that infected the hosts' common ancestors, diversified alongside host diversification. For others, we observe Microsporidia taxa with wide host ranges that do not follow host phylogeny.

Pinworms of the red howler monkey (Alouatta seniculus) in Colombia: Gathering the pieces of the pinworm-primate puzzle.

Pinworms of primates are believed to be highly host specific parasites, forming co-evolutionary associations with their hosts. In order to assess the strength and reach of such evolutionary links, we need to have a broad understanding of the pinworm diversity associated with primates. Here, we employed an integrative taxonomic approach to assess pinworm diversity in red howler monkeys in Colombia. Molecular and morphological evidence validate the presence of at least four different species of Trypanoxyuris occurring in red howler monkeys: T. minutus, a widely distributed species, and three new species, T. seunimiii n. sp., T. kemuimae n. sp. and T. kotudoi n. sp. The mitochondrial COI gene and the 28S ribosomal gene were used for phylogenetic assessments through Bayesian inference. The three new species were morphologically distinct and formed reciprocally monophyletic lineages. Further molecular lineage subdivision in T. minutus and T. kotudoi n. sp. without morphological correspondence, suggests the potential scenario for the existence of cryptic species. Phylogenetic relationships imply that the different species of Trypanoxyuris occurring in each howler monkey species were acquired through independent colonization events. On-going efforts to uncover pinworm diversity will allow us to test the degree of host specificity and the co-phylogenetic hypothesis, as well as to further unravel the primate-pinworm evolutionary history puzzle.

Norileca indica (Crustacea: Isopoda, Cymothoidae) Infects Rastrelliger kanagurta Along the Malabar Coast of India - Seasonal Variation in the Prevalence and Aspects of Host-parasite Interactions.

This paper reports seasonal variations in the prevalence of host-parasite associations between Norileca indica, a cymothoid, and the Indian mackerel Rastrelliger kanagurta along the Malabar Coast of India. Eighty-eight marine fish species belonging to diverse families were examined, and only R. kanagurta was shown to be parasitized by N. indica, indicating a narrow host specificity. The prevalence, mean intensity, and abundance were 30.70, 1.71, and 0.52%, respectively. Different life cycle of this parasite, from larva to adult, infect the host fish. In most instances, a pair of N. indica infected the host, and in these instances a male-female combination was the most common (89.17%). The monthly occurrence of N. indica was charted for a period of 38 months (July 2012 to July 2014; March 2017 to March 2018), and statistical comparison of the data showed a significant difference (p < 0.001) among seasons. A positive correlation (r = 0.40) was observed between the size of female parasites and that of their respective host fish. There was a positive correlation (r = 0.78) between the size of female parasites and their fecundity. In all instances, adult N. indica individuals were found to specifically attach to the mucus membrane of branchial operculum, causing visible physical damage, including atrophy (reduced length) of the gill filaments and overall loss of gill normalcy. Furthermore, permanent occupancy by female N. indica resulted in the formation of a deep pit in the gill chamber floor and also caused atrophy of gill filaments. Overall, our findings yielded a greater understanding of the occurrence, season-wise prevalence, and potential host- parasite associations of N. indica.

Malagasy bats shelter a considerable genetic diversity of pathogenic Leptospira suggesting notable host-specificity patterns.

Pathogenic Leptospira are the causative agents of leptospirosis, a disease of global concern with major impact in tropical regions. Despite the importance of this zoonosis for human health, the evolutionary and ecological drivers shaping bacterial communities in host reservoirs remain poorly investigated. Here, we describe Leptospira communities hosted by Malagasy bats, composed of mostly endemic species, in order to characterize host-pathogen associations and investigate their evolutionary histories. We screened 947 individual bats (representing 31 species, 18 genera and seven families) for Leptospira infection and subsequently genotyped positive samples using three different bacterial loci. Molecular identification showed that these Leptospira are notably diverse and include several distinct lineages mostly belonging to Leptospira borgpetersenii and L. kirschneri. The exploration of the most probable host-pathogen evolutionary scenarios suggests that bacterial genetic diversity results from a combination of events related to the ecology and the evolutionary history of their hosts. Importantly, based on the data set presented herein, the notable host-specificity we have uncovered, together with a lack of geographical structuration of bacterial genetic diversity, indicates that the Leptospira community at a given site depends on the co-occurring bat species assemblage. The implications of such tight host-specificity on the epidemiology of leptospirosis are discussed.

Experimental infection of the digeneans to some congeneric snail species radiated in a single water system: Relative importance of local evolution and phylogenetic constraint.

To determine the relative importance of local adaptation caused by host-parasite coevolution and resource tracking by the parasites, the susceptibility of the freshwater snail genus Semisulcospira to the digenean parasite genus Genarchopsis was investigated experimentally. Four snail species endemic to the Lake Biwa system in Japan and two non-endemic species were investigated. All but one species was also tested for local variation in susceptibility. Parasites were obtained from Takashima (mix population of Genarchopsis gigi and Genarchopsis chubuensis) and Nagahama (G. chubuensis). In endemic Semisulcospira, closely related specie pairs (Semisulcospira habei and Semisulcospira niponica, Semisulcospira decipiens and Semisulcospira nakasekoae) showed similar susceptibilities to parasites from both localities. S. habei and S. niponica were highly susceptible to parasites from Takashima, but were resistant to parasites from Nagahama. S. decipiens and S. nakasekoae showed moderate susceptibility to parasites from both localities. None of the endemic snail species showed a clear local variation in susceptibility. These results show that the susceptibility of endemic Semisulcospira to Genarchopsis is conservative and can be regarded as an example of resource-tracking. One of the non-endemic snails, Semisulcospira libertina, showed local variation in susceptibility. This variation was not related to the sympatry of the parasites used for the experimental infection, suggesting that it was not the result of local adaptation by parasites.

A new genus of Strepsiptera, Rozenia gen. n. (Stylopidae), a parasite of bee genera Acamptopoeum and Calliopsis (Andrenidae, Panurginae, Calliopsini).

A new Strepsiptera genus from South America is described, Rozenia gen. n., with three new species: Rozeniacalliopsidis sp. n. (type species), Rozeniaperuana sp. n. and Rozeniaplaticephala sp. n. These three new species are parasites of bees belonging to the tribe Calliopsini (Andrenidae, Panurginae). Rozeniacalliopsidis sp. n. is a parasite of the bee genus Calliopsis Smith, 1853 and Rozeniaperuana sp. n. and Rozeniaplaticephala sp. n. are parasites of the bee genus Acamptopoeum Cockerell, 1905. Diagnoses and descriptions of female puparia are presented for all three species. Diagnoses and descriptions of first instars (triungulinids) are presented for Rozeniacalliopsidis sp. n. and Rozeniaplaticephala sp. n. The first case of increased number of setae on the body of the first instars and augmentation of chaetotaxy of Strepsiptera are discussed.