Yann Gomard

Pathogenic Leptospira spp. in bats, Madagascar and Union of the Comoros.

[Extract] To the Editor: Leptospirosis is a zoonosis of global distribution; incidence rates are particularly high in tropical areas (1). Leptospirosis is a major public health problem on islands in the southwestern Indian Ocean, particularly La Reunion, Mayotte, and the Seychelles (where incidence rates are among the highest in the world) (1). In contrast, no human case has been reported on the nearby islands of Madagascar and Union of the Comoros. However, the recent demonstration of pathogenic Leptospira spp. in small mammals introduced to Madagascar suggests possible transmission from free-living animals to humans (2).

The Bacteriome of Bat Flies (Nycteribiidae) from the Malagasy Region: a Community Shaped by Host Ecology, Bacterial Transmission Mode, and Host-Vector Specificity

ABSTRACT The Nycteribiidae are obligate blood-sucking Diptera (Hippoboscoidea) flies that parasitize bats. Depending on species, these wingless flies exhibit either high specialism or generalism toward their hosts, which may in turn have important consequences in terms of their associated microbial community structure. Bats have been hypothesized to be reservoirs of numerous infectious agents, some of which have recently emerged in human populations. Thus, bat flies may be important in the epidemiology and transmission of some of these bat-borne infectious diseases, acting either directly as arthropod vectors or indirectly by shaping pathogen communities among bat populations. In addition, bat flies commonly have associations with heritable bacterial endosymbionts that inhabit insect cells and depend on maternal transmission through egg cytoplasm to ensure their transmission. Some of these heritable bacteria are likely obligate mutualists required to support bat fly development, but others are facultative symbionts with unknown effects. Here, we present bacterial community profiles that were obtained from seven bat fly species, representing five genera, parasitizing bats from the Malagasy region. The observed bacterial diversity includes Rickettsia, Wolbachia, and several Arsenophonus-like organisms, as well as other members of the Enterobacteriales and a widespread association of Bartonella bacteria from bat flies of all five genera. Using the well-described host specificity of these flies and data on community structure from selected bacterial taxa with either vertical or horizontal transmission, we show that host/vector specificity and transmission mode are important drivers of bacterial community structure.

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.

Human Leptospirosis on Reunion Island, Indian Ocean: Are Rodents the (Only) Ones to Blame?

Background Although leptospirosis is a zoonosis of major concern on tropical islands, the molecular epidemiology of the disease aiming at linking human cases to specific animal reservoirs has been rarely explored within these peculiar ecosystems. Methodology/Principal Findings Five species of wild small mammals (n = 995) as well as domestic animals (n = 101) were screened for Leptospira infection on Reunion Island; positive samples were subsequently genotyped and compared to Leptospira from clinical cases diagnosed in 2012–2013 (n = 66), using MLST analysis. We identified two pathogenic species in human cases, namely Leptospira interrogans and Leptospira borgpetersenii. Leptospira interrogans was by far dominant both in clinical samples (96.6%) and in infected animal samples (95.8%), with Rattus spp and dogs being its exclusive carriers. The genetic diversity within L. interrogans was apparently limited to two sequence types (STs): ST02, identified among most clinical samples and in all rats with complete MLST, and ST34, identified in six humans, but not in rats. Noteworthy, L. interrogans detected in two stray dogs partially matched with ST02 and ST34. Leptospira borgpetersenii was identified in two clinical samples only (3.4%), as well as in cows and mice; four haplotypes were identified, of which two seemingly identical in clinical and animal samples. Leptospira borgpetersenii haplotypes detected in human cases were clearly distinct from the lineage detected so far in the endemic bat species Mormopterus francoismoutoui, thus excluding a role for this volant mammal in the local human epidemiology of the disease. Conclusions/Significance Our data confirm rats as a major reservoir of Leptospira on Reunion Island, but also pinpoint a possible role of dogs, cows and mice in the local epidemiology of human leptospirosis. This study shows that a comprehensive molecular characterization of pathogenic Leptospira in both clinical and animal samples helps to gaining insight into leptospirosis epidemiology within a specific environmental setting.

Identification of Tenrec ecaudatus, a Wild Mammal Introduced to Mayotte Island, as a Reservoir of the Newly Identified Human Pathogenic Leptospira mayottensis

Leptospirosis is a bacterial zoonosis of major concern on tropical islands. Human populations on western Indian Ocean islands are strongly affected by the disease although each archipelago shows contrasting epidemiology. For instance, Mayotte, part of the Comoros Archipelago, differs from the other neighbouring islands by a high diversity of Leptospira species infecting humans that includes Leptospira mayottensis, a species thought to be unique to this island. Using bacterial culture, molecular detection and typing, the present study explored the wild and domestic local mammalian fauna for renal carriage of leptospires and addressed the genetic relationships of the infecting strains with local isolates obtained from acute human cases and with Leptospira strains hosted by mammal species endemic to nearby Madagascar. Tenrec (Tenrec ecaudatus, Family Tenrecidae), a terrestrial mammal introduced from Madagascar, is identified as a reservoir of L. mayottensis. All isolated L. mayottensis sequence types form a monophyletic clade that includes Leptospira strains infecting humans and tenrecs on Mayotte, as well as two other Malagasy endemic tenrecid species of the genus Microgale. The lower diversity of L. mayottensis in tenrecs from Mayotte, compared to that occurring in Madagascar, suggests that L. mayottensis has indeed a Malagasy origin. This study also showed that introduced rats (Rattus rattus) and dogs are probably the main reservoirs of Leptospira borgpetersenii and Leptospira kirschneri, both bacteria being prevalent in local clinical cases. Data emphasize the epidemiological link between the two neighbouring islands and the role of introduced small mammals in shaping the local epidemiology of leptospirosis.

Evolutionary History of Indian Ocean Nycteribiid Bat Flies Mirroring the Ecology of Their Hosts

Bats and their parasites are increasingly investigated for their role in maintenance and transmission of potentially emerging pathogens. The islands of the western Indian Ocean hold nearly 50 bat species, mostly endemic and taxonomically well studied. However, investigation of associated viral, bacterial, and external parasites has lagged behind. In the case of their ectoparasites, more detailed information should provide insights into the evolutionary history of their hosts, as well as pathogen cycles in these wild animals. Here we investigate species of Nycteribiidae, a family of obligate hematophagous wingless flies parasitizing bats. Using morphological and molecular approaches, we describe fly species diversity sampled on Madagascar and the Comoros for two cave-roosting bat genera with contrasting ecologies: Miniopterus and Rousettus. Within the sampling area, 11 endemic species of insect-feeding Miniopterus occur, two of which are common to Madagascar and Comoros, while fruit-consuming Rousettus are represented by one species endemic to each of these zones. Morphological and molecular characterization of flies reveals that nycteribiids associated with Miniopterus bats comprise three species largely shared by most host species. Flies of M. griveaudi, one of the two bats found on Madagascar and certain islands in the Comoros, belong to the same taxon, which accords with continued over-water population exchange of this bat species and the lack of inter-island genetic structuring. Flies parasitizing Rousettus belong to two distinct species, each associated with a single host species, again in accordance with the distribution of each endemic bat species.

An eco-epidemiological study of Morbilli-related paramyxovirus infection in Madagascar bats reveals host-switching as the dominant macro-evolutionary mechanism

An eco-epidemiological investigation was carried out on Madagascar bat communities to better understand the evolutionary mechanisms and environmental factors that affect virus transmission among bat species in closely related members of the genus Morbillivirus, currently referred to as Unclassified Morbilli-related paramyxoviruses (UMRVs). A total of 947 bats were investigated originating from 52 capture sites (22 caves, 18 buildings, and 12 outdoor sites) distributed over different bioclimatic zones of the island. Using RT-PCR targeting the L-polymerase gene of the Paramyxoviridae family, we found that 10.5% of sampled bats were infected, representing six out of seven families and 15 out of 31 species analyzed. Univariate analysis indicates that both abiotic and biotic factors may promote viral infection. Using generalized linear modeling of UMRV infection overlaid on biotic and abiotic variables, we demonstrate that sympatric occurrence of bats is a major factor for virus transmission. Phylogenetic analyses revealed that all paramyxoviruses infecting Malagasy bats are UMRVs and showed little host specificity. Analyses using the maximum parsimony reconciliation tool CoRe-PA, indicate that host-switching, rather than co-speciation, is the dominant macro-evolutionary mechanism of UMRVs among Malagasy bats.

Serologic evidence of leptospirosis in humans, Union of the Comoros, 2011.

To the Editor: Leptospirosis is a worldwide bacterial zoonosis caused by infection with pathogenic Leptospira spp. (Spirochaetales, Leptospiraceae). Most mammals can be infected, but rats are considered the main reservoir, maintaining Leptospira spirochetes in the lumen of renal tubules and contaminating the environment with bacteria-infected urine. Transmission to humans is accidental, occurring through contact with animal secretions or with contaminated environmental materials. In temperate countries, human leptospirosis is a sporadic disease; incidence is much higher in the tropics because climate and environmental conditions are conducive to the survival of bacteria, resulting in increased exposure of humans to leptospirosis-causing pathogens (1). Among islands in the southwestern Indian Ocean, human leptospirosis is endemic to Mayotte, France, and La Reunion (2–4) and to the Seychelles, where the incidence of leptospirosis is one of the highest worldwide (5). Leptospirosis is poorly documented in other islands in the region, including Mauritius, Madagascar, and the Union of the Comoros (2,6–8). Whether the scant documentation indicates underdiagnosis or reflects local epidemiologic specificities is unknown. To improve knowledge of Leptospira infection in the region, we conducted a study in the Union of the Comoros to serologically assess the presence or absence of leptospirosis in humans. The Union of the Comoros consists of 3 islands: Grande-Comore, Moheli, and Anjouan. Together with a fourth, southern island, Mayotte, these islands form the Comoros Archipelago. For feasibility reasons, we used excess serum samples. Seventy-six samples were from healthy volunteers who gave informed consent; 318 clinical blood samples from patients had been obtained by private laboratories and by the surveillance laboratory of the National Malaria Control Programme (PNLP) during August 1–October 8, 2011. The Ministere de la Sante, de la Solidarite et de la Promotion du Genre of the Union of the Comoros, authorized the serologic investigation (authorization no. 1175/MSSPG/DNS). We used the microscopic agglutination test (MAT) to test serum samples; the MAT was based on a panel of 15 Leptospira strains, enabling the screening of all recently reported serogroups for human and animal cases on neighboring Mayotte (2,4,9). A list of the tested strains follows, shown as Genus species Serogroup/Serovar (type strain): L. borgpetersenii Ballum/Castellonis (Castellon 3), L. borgpetersenii Sejroe/Hardjobovis (Sponselee), L. borgpetersenii Sejroe/Sejroe (M 84), L. borgpetersenii Tarassovi/Tarassovi (Perepelicin), L. interrogans Australis/Australis (Ballico), L. interrogans Autumnalis/Autumnalis (Akiyami A), L. interrogans Bataviae/Bataviae (Van Tienen), L. interrogans Canicola/Canicola (Hond Utrecht IV), L. interrogans Hebdomadis/Hebdomadis (Hebdomadis), L. interrogans Icterohaemorrhagiae/Copenhageni (Wijnberg), L. interrogans Pyrogenes/Pyrogenes (Salinem), L. kirschneri Cynopteri/Cynopteri (3522C), L. kirschneri Grippotyphosa/Grippotyphosa (Moskva V), L. kirschneri Mini/Undetermined serovar (200803703) (9), L. noguchii Panama/Panama (CZ214K). Each serum sample was tested at dilutions ranging from 1:50 to 1:3,200 and considered positive when the MAT titer was >100. Our serologic findings showed evidence of Leptospira infection in humans on the 3 islands of the Union of the Comoros (MAT titers 100–1,600, geometric mean titer [GMT] 194). The positivity rate was 10.3% (95% CI 4.8–15.9) for samples from Moheli, 4.2% (95% CI 1.4–7.0) for samples from Grande-Comore, and 3.4% (95% CI 0.1–6.7) for samples from Anjouan; no significant difference was found between islands or by the age or sex of residents (p>0.05, Fisher exact test). Leptospira infection was more prevalent and MAT titers were higher among serum samples from the patient group than the healthy donor group (20 positive samples/318 total vs. 3 positive samples/76 total; GMT 207 vs. GMT 126), but the difference was not significant (p>0.05, Fisher exact test). In 78% of seropositive serum samples, antibodies reacted with serogroups Australis, Bataviae, Grippotyphosa, Panama, Pomona, Pyrogenes, Mini, and/or Sejroe. MAT titers >100, which are suggestive of more specific antibodies to Leptospira, were observed for all serogroups except Australis and Sejroe. Pyrogenes serogroup was identified in one third of positive samples from Moheli and was associated with the highest agglutination titers (Figure). Figure Microagglutination test results, showing serologic evidence of leptospirosis in humans, Union of the Comoros, 2011. The percentage of positive cases is shown for each island; the number below the percentage indicates the number of positive serum samples/total ... Our data indicate that Leptospira infections do occur in humans in the Union of the Comoros; this finding is consistent with those in studies reporting leptospirosis in persons returning from travel in the Union of the Comoros (2,8) and with the detection of pathogenic Leptospira spp. in bats sampled on these islands (10). The human leptospirosis–related serologic findings in Union of Comoros are most comparable to those from neighboring Mayotte, where leptospirosis is mainly caused by serogroups Mini/Sejroe/Hebdomadis complex, Pyrogenes, Grippotyphosa, and Pomona and where serogroup Icterohaemorraghiae is not detectable (2). These findings contrast with human leptospirosis findings from La Reunion and the Seychelles, where the Icterohaemorraghiae serogroup is most common (3). Our MAT-derived data cannot discriminate between recent and past Leptospira infections, nor can these data be used to determine the severity of the disease in the Union of the Comoros. Nonetheless, the data strongly support the presence of human leptospirosis on the 3 islands of the Union of the Comoros and emphasize the need for a proper diagnosis to ascertain the number of leptospirosis cases among the acute febrile illnesses in this country.

Hidden diversity of Nycteribiidae (Diptera) bat flies from the Malagasy region and insights on host-parasite interactions

BackgroundWe present information on Nycteribiidae flies parasitizing the bat families Pteropodidae, Miniopteridae and Vespertilionidae from the Malagasy Region, contributing insight into their diversity and host preference.ResultsOur phylogenetic analysis identified nine clusters of nycteribiid bat flies on Madagascar and the neighbouring Comoros Archipelago. Bat flies sampled from frugivorous bats of the family Pteropodidae are monoxenous: Eucampsipoda madagascariensis, E. theodori and Cyclopodia dubia appear wholly restricted to Rousettus madagascariensis, R. obliviosus and Eidolon dupreanum, respectively. Two different host preference patterns occurred in nycteribiids infecting insectivorous bats. Flies parasitizing bats of the genera Miniopterus (Miniopteridae) and Myotis (Vespertilionidae), namely Penicillidia leptothrinax, Penicillidia sp. and Nycteribia stylidiopsis, are polyxenous and showed little host preference, while those parasitizing the genera Pipistrellus and Scotophilus (both Vespertilionidae) and referable to Basilia spp., are monoxenous. Lastly, the inferred Bayesian phylogeny revealed that the genus Basilia, as currently configured, is paraphyletic.ConclusionThis study provides new information on the differentiation of nycteribiid taxa, including undescribed species. Host preference is either strict as exemplified by flies parasitizing fruit bats, or more relaxed as found on some insectivorous bat species, possibly because of roost site sharing. Detailed taxonomic work is needed to address three undescribed nycteribiid taxa found on Pipistrellus and Scotophilus, tentatively allocated to the genus Basilia, but possibly warranting different generic allocation.

Human leptospirosis in Seychelles: A prospective study confirms the heavy burden of the disease but suggests that rats are not the main reservoir

Background Leptospirosis is a bacterial zoonosis caused by pathogenic Leptospira for which rats are considered as the main reservoir. Disease incidence is higher in tropical countries, especially in insular ecosystems. Our objectives were to determine the current burden of leptospirosis in Seychelles, a country ranking first worldwide according to historical data, to establish epidemiological links between animal reservoirs and human disease, and to identify drivers of transmission. Methods A total of 223 patients with acute febrile symptoms of unknown origin were enrolled in a 12-months prospective study and tested for leptospirosis through real-time PCR, IgM ELISA and MAT. In addition, 739 rats trapped throughout the main island were investigated for Leptospira renal carriage. All molecularly confirmed positive samples were further genotyped. Results A total of 51 patients fulfilled the biological criteria of acute leptospirosis, corresponding to an annual incidence of 54.6 (95% CI 40.7–71.8) per 100,000 inhabitants. Leptospira carriage in Rattus spp. was overall low (7.7%) but dramatically higher in Rattus norvegicus (52.9%) than in Rattus rattus (4.4%). Leptospira interrogans was the only detected species in both humans and rats, and was represented by three distinct Sequence Types (STs). Two were novel STs identified in two thirds of acute human cases while noteworthily absent from rats. Conclusions This study shows that human leptospirosis still represents a heavy disease burden in Seychelles. Genotype data suggests that rats are actually not the main reservoir for human disease. We highlight a rather limited efficacy of preventive measures so far implemented in Seychelles. This could result from ineffective control measures of excreting animal populations, possibly due to a misidentification of the main contaminating reservoir(s). Altogether, presented data stimulate the exploration of alternative reservoir animal hosts.