Analytical framework to evaluate and optimize the use of imperfect diagnostics to inform outbreak response: Application to the 2017 plague epidemic in Madagascar.

During outbreaks, the lack of diagnostic "gold standard" can mask the true burden of infection in the population and hamper the allocation of resources required for control. Here, we present an analytical framework to evaluate and optimize the use of diagnostics when multiple yet imperfect diagnostic tests are available. We apply it to laboratory results of 2,136 samples, analyzed with 3 diagnostic tests (based on up to 7 diagnostic outcomes), collected during the 2017 pneumonic (PP) and bubonic plague (BP) outbreak in Madagascar, which was unprecedented both in the number of notified cases, clinical presentation, and spatial distribution. The extent of these outbreaks has however remained unclear due to nonoptimal assays. Using latent class methods, we estimate that 7% to 15% of notified cases were Yersinia pestis-infected. Overreporting was highest during the peak of the outbreak and lowest in the rural settings endemic to Y. pestis. Molecular biology methods offered the best compromise between sensitivity and specificity. The specificity of the rapid diagnostic test was relatively low (PP: 82%, BP: 85%), particularly for use in contexts with large quantities of misclassified cases. Comparison with data from a subsequent seasonal Y. pestis outbreak in 2018 reveal better test performance (BP: specificity 99%, sensitivity: 91%), indicating that factors related to the response to a large, explosive outbreak may well have affected test performance. We used our framework to optimize the case classification and derive consolidated epidemic trends. Our approach may help reduce uncertainties in other outbreaks where diagnostics are imperfect.

Multiple Introductions of Yersinia pestis during Urban Pneumonic Plague Epidemic, Madagascar, 2017.

Pneumonic plague (PP) is characterized by high infection rate, person-to-person transmission, and rapid progression to severe disease. In 2017, a PP epidemic occurred in 2 Madagascar urban areas, Antananarivo and Toamasina. We used epidemiologic data and Yersinia pestis genomic characterization to determine the sources of this epidemic. Human plague emerged independently from environmental reservoirs in rural endemic foci >20 times during August-November 2017. Confirmed cases from 5 emergences, including 4 PP cases, were documented in urban areas. Epidemiologic and genetic analyses of cases associated with the first emergence event to reach urban areas confirmed that transmission started in August; spread to Antananarivo, Toamasina, and other locations; and persisted in Antananarivo until at least mid-November. Two other Y. pestis lineages may have caused persistent PP transmission chains in Antananarivo. Multiple Y. pestis lineages were independently introduced to urban areas from several rural foci via travel of infected persons during the epidemic.

Investigation of astrovirus, coronavirus and paramyxovirus co-infections in bats in the western Indian Ocean.

Co-infections have a key role in virus transmission in wild reservoir hosts. We investigated the simultaneous presence of astroviruses, coronaviruses, and paramyxoviruses in bats from Madagascar, Mayotte, Mozambique, and Reunion Island. A total of 871 samples from 28 bat species representing 8 families were tested by polymerase chain reactions (PCRs) targeting the RNA-dependent RNA-polymerase genes. Overall, 2.4% of bats tested positive for the presence of at least two viruses, only on Madagascar and in Mozambique. Significant variation in the proportion of co-infections was detected among bat species, and some combinations of co-infection were more common than others. Our findings support that co-infections of the three targeted viruses occur in bats in the western Indian Ocean region, although further studies are needed to assess their epidemiological consequences.

Diversity, distribution, and drivers of Polychromophilus infection in Malagasy bats.

BACKGROUND: Numerous studies have been undertaken to advance knowledge of apicomplexan parasites infecting vertebrates, including humans. Of these parasites, the genus Plasmodium has been most extensively studied because of the socio-economic and public health impacts of malaria. In non-human vertebrates, studies on malaria or malaria-like parasite groups have been conducted but information is far from complete. In Madagascar, recent studies on bat blood parasites indicate that three chiropteran families (Miniopteridae, Rhinonycteridae, and Vespertilionidae) are infected by the genus Polychromophilus with pronounced host specificity: Miniopterus spp. (Miniopteridae) harbour Polychromophilus melanipherus and Myotis goudoti (Vespertilionidae) is infected by Polychromophilus murinus. However, most of the individuals analysed in previous studies were sampled on the western and central portions of the island. The aims of this study are (1) to add new information on bat blood parasites in eastern Madagascar, and (2) to highlight biotic and abiotic variables driving prevalence across the island. METHODS: Fieldworks were undertaken from 2014 to 2016 in four sites in the eastern portion of Madagascar to capture bats and collect biological samples. Morphological and molecular techniques were used to identify the presence of haemosporidian parasites. Further, a MaxEnt modelling was undertaken using data from Polychromophilus melanipherus to identify variables influencing the presence of this parasite RESULTS: In total, 222 individual bats belonging to 17 species and seven families were analysed. Polychromophilus infections were identified in two families: Miniopteridae and Vespertilionidae. Molecular data showed that Polychromophilus spp. parasitizing Malagasy bats form a monophyletic group composed of three distinct clades displaying marked host specificity. In addition to P. melanipherus and P. murinus, hosted by Miniopterus spp. and Myotis goudoti, respectively, a novel Polychromophilus lineage was identified from a single individual of Scotophilus robustus. Based on the present study and the literature, different biotic and abiotic factors are shown to influence Polychromophilus infection in bats, which are correlated based on MaxEnt modelling. CONCLUSIONS: The present study improves current knowledge on Polychromophilus blood parasites infecting Malagasy bats and confirms the existence of a novel Polychromophilus lineage in Scotophilus bats. Additional studies are needed to obtain additional material of this novel lineage to resolve its taxonomic relationship with known members of the genus. Further, the transmission mode of Polychromophilus in bats as well as its potential effect on bat populations should be investigated to complement the results provided by MaxEnt modelling and eventually provide a comprehensive picture of the biology of host-parasite interactions.

Bat Astrovirus in Mozambique.

Astroviruses (AstVs) are responsible for infection of a large diversity of mammalian and avian species, including bats, aquatic birds, livestock and humans. We investigated AstVs circulation in bats in Mozambique and Mayotte, a small island in the Comoros Archipelago located between east Africa and Madagascar. Biological material was collected from 338 bats and tested for the presence of the AstV RNA-dependent RNA-polymerase gene with a pan-AstV semi-nested polymerase chain reaction assay. None of the 79 samples obtained from Mayotte bats (Pteropus seychellensis comorensis and Chaerephon pusillus) tested positive; however, 20.1% of bats sampled in Mozambique shed AstVs at the time of sampling and significant interspecific variation in the proportion of positive bats was detected. Many AstVs sequences obtained from a given bat species clustered in different phylogenetic lineages, while others seem to reflect some level of host-virus association, but also with AstVs previously reported from Malagasy bats. Our findings support active circulation of a large diversity of AstVs in bats in the western Indian Ocean islands, including the southeastern African coast, and highlight the need for more detailed assessment of its risk of zoonotic transmission to human populations.

Polychromophilus spp. (Haemosporida) in Malagasy bats: host specificity and insights on invertebrate vectors.

BACKGROUND: Bats are home to diverse haemosporidian parasites namely Plasmodium and Plasmodium-related. While information is available at a worldwide level, haemosporidian infection in bats from Madagascar is still scarce and recent changes in the taxonomy of the island's bat fauna, particularly the description of several new species, require a reassessment of previously described patterns, including blood parasite ecology and vectorial transmission. METHODS: A sample representing seven of the nine known bat families and 31 of the 46 currently recognized taxa from Madagascar and collected in the western and central portions of the island were screened by PCR for the presence of Polychromophilus. In addition, Nycteribiidae flies parasitizing Miniopteridae and Vespertilionidae were screened for parasites with the aim to better understand aspects of vector transmission. Phylogenetic reconstruction using the mitochondrial cytochrome b encoding gene was used in a Bayesian analysis to examine the relationship between Polychromophilus recovered from Malagasy bats and those identified elsewhere. RESULTS: Polychromophilus infection was restricted to Miniopterus spp. (Miniopteridae), Myotis goudoti (Vespertilionidae), and Paratriaenops furculus (Rhinonycteridae), with an overall infection rate of 13.5%. Polychromophilus melanipherus was found infecting Miniopterus spp. and P. furculus, whereas Polychromophilus murinus was only recovered from M. goudoti. These two protozoan parasites species were also detected in bat flies species known to parasitize Miniopterus spp. and M. goudoti, respectively. Generalized linear model analyses were conducted to elucidate the effect of species and sex on haemoparasites infection in Miniopterus spp., which revealed that males have higher risk of infection than females and prevalence differed according to the considered Miniopterus host. Molecular screening of nycteribiid flies revealed three positive species for Polychromophilus spp., including Penicillidia sp. (cf. fulvida), Penicillidia leptothrinax, and Nycteribia stylidiopsis. These three fly species are known to parasitize Miniopterus spp. and M. goudoti and should be considered as potential vectors of Polychromophilus spp. CONCLUSION: Phylogenetic analyses demonstrated the existence of at least four distinct clades within the genus Polychromophilus, two of which were documented in the present study. The screening of nycteribiid flies overlaid on the highly diversified genus Miniopterus, provides considerable insight into parasite transmission, with bat infection being associated with their roosting behaviour and the occurrence of specific arthropod vectors.

Pneumonic Plague Transmission, Moramanga, Madagascar, 2015.

During a pneumonic plague outbreak in Moramanga, Madagascar, we identified 4 confirmed, 1 presumptive, and 9 suspected plague case-patients. Human-to-human transmission among close contacts was high (reproductive number 1.44) and the case fatality rate was 71%. Phylogenetic analysis showed that the Yersinia pestis isolates belonged to group q3, different from the previous outbreak.

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

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.

Leptospira and paramyxovirus infection dynamics in a bat maternity enlightens pathogen maintenance in wildlife.

Bats are reservoirs for several zoonotic pathogens of medical importance; however, infection dynamics of pathogens in wild bat populations remain poorly understood. Here, we examine the influence of host crowding and population age structure on pathogen transmission and diversity in bat populations. Focusing on two pathogen taxa of medical importance, Leptospira bacteria and paramyxoviruses, we monitored host population and pathogen shedding dynamics within a maternity colony of the tropical bat species Mormopterus francoismoutoui, endemic to Réunion Island. Our data reveal astonishingly similar infection dynamics for Leptospira and paramyxoviruses, with infection peaks during late pregnancy and 2 months after the initial birth pulse. Furthermore, although co-infection occurs frequently during the peaks of transmission, the patterns do not suggest any interaction between the two pathogens. Partial sequencing reveals a unique bat-specific Leptospira strain contrasting with the co-circulation of four separate paramyxovirus lineages along the whole breeding period. Patterns of infection highlight the importance of host crowding in pathogen transmission and suggest that most bats developed immune response and stop excreting pathogens. Our results support that bat maternity colonies may represent hot spots of transmission for bacterial and viral infectious agents, and highlight how seasonality can be an important determinant of host-parasite interactions and disease emergence.

Highly diverse morbillivirus-related paramyxoviruses in wild fauna of the southwestern Indian Ocean Islands: evidence of exchange between introduced and endemic small mammals.

UNLABELLED: The Paramyxoviridae form an increasingly diverse viral family, infecting a wide variety of different hosts. In recent years, they have been linked to disease emergence in many different animal populations and in humans. Bats and rodents have been identified as major animal populations capable of harboring paramyxoviruses, and host shifting between these animals is likely to be an important driving factor in the underlying evolutionary processes that eventually lead to disease emergence. Here, we have studied paramyxovirus circulation within populations of endemic and introduced wild small mammals of the southwestern Indian Ocean region and belonging to four taxonomic orders: Rodentia, Afrosoricida, Soricomorpha, and Chiroptera. We report elevated infection levels as well as widespread paramyxovirus dispersal and frequent host exchange of a newly emerging genus of the Paramyxoviridae, currently referred to as the unclassified morbillivirus-related viruses (UMRVs). In contrast to other genera of the Paramyxoviridae, where bats have been shown to be a key host species, we show that rodents (and, in particular, Rattus rattus) are significant spreaders of UMRVs. We predict that the ecological particularities of the southwestern Indian Ocean, where small mammal species often live in densely packed, multispecies communities, in combination with the increasing invasion of R. rattus and perturbations of endemic animal communities by active anthropological development, will have a major influence on the dynamics of UMRV infection. IMPORTANCE: Identification of the infectious agents that circulate within wild animal reservoirs is essential for several reasons: (i) infectious disease outbreaks often originate from wild fauna; (ii) anthropological expansion increases the risk of contact between human and animal populations and, as a result, the risk of disease emergence; (iii) evaluation of pathogen reservoirs helps in elaborating preventive measures to limit the risk of disease emergence. Many paramyxoviruses for which bats and rodents serve as major reservoirs have demonstrated their potential to cause disease in humans and animals. In the context of the biodiversity hot spot of southwestern Indian Ocean islands and their rich endemic fauna, we show that highly diverse UMRVs exchange between various endemic animal species, and their dissemination likely is facilitated by the introduced Rattus rattus. Hence, many members of the Paramyxoviridae appear well adapted for the study of the viral phylodynamics that may be associated with disease emergence.

Population dynamics of plague vector fleas in an endemic focus: implications for plague surveillance.

Plague is a zoonotic vector-borne disease caused by the bacterium Yersinia pestis. In Madagascar, it persists in identified foci, where it is a threat to public health generally from September to April. A more complete understanding of how the disease persists could guide control strategies. Fleas are the main vector for transmission between small mammal hosts and humans, and fleas likely play a role in the maintenance of plague. This study characterized the dynamics of flea populations in plague foci alongside the occurrence of human cases. From 2018 to 2020, small mammals were trapped at sites in the central Highlands of Madagascar. A total of 2,762 small mammals were captured and 5,295 fleas were collected. The analysis examines 2 plague vector species in Madagascar (Synopsyllus fonquerniei and Xenopsylla cheopis). Generalized linear models were used to relate flea abundance to abiotic factors, with adjustments for trap location and flea species. We observed significant effects of abiotic factors on the abundance, intensity, and infestation rate by the outdoor-associated flea species, S. fonquerniei, but weak seasonality for the indoor-associated flea species, X. cheopis. A difference in the timing of peak abundance was observed between the 2 flea species during and outside the plague season. While the present study did not identify a clear link between flea population dynamics and plague maintenance, as only one collected X. cheopis was infected, the results presented herein can be used by local health authorities to improve monitoring and control strategies of plague vector fleas in Madagascar.

High Rickettsial Diversity in Rodents and Their Ectoparasites From the Central Highlands of Madagascar.

Rickettsioses are among emerging infectious diseases around the world. In Madagascar, little information is available regarding Rickettsia (Rickettsiales: Rickettsiaceae) diversity and their potential impacts on public health. In fact, molecular screening of ectoparasites of mammals reported the presence of three species, Rickettsia africae, Rickettsia typhi, and Rickettsia felis. The present study aims to investigate the diversity of Rickettsia in small mammals and associated ectoparasites (fleas and ticks) using a molecular approach. In September and December 2016, fieldworks were undertaken in two districts of Madagascar to capture small mammals using standard traps (Tomahawk and Sherman traps) and collect associated ectoparasites. In total, 12 taxa of ectoparasites (5 flea and 7 tick species) were collected from 89 individuals of four species of terrestrial small mammals. Rickettsia spp. were molecularly identified in one specimen of Rattus rattus (Rodentia: Muridae), one specimen of Pulex irritans (Siphonaptera: Pulicidae) as well as four specimens of Ixodes cf. colasbelcouri (Ixodida: Ixodidae). This study showed the presence of three phylogenetically distinct taxa of Rickettsia in small mammals and their ectoparasites. The current study broadens our knowledge on the diversity of Rickettsia in the Central Highlands of Madagascar and highlights for the first time the presence of Ri. felis in R. rattus and in tick, I. cf. colasbelcouri in Madagascar. Additional studies are needed to have exhaustive information on Rickettsia in small mammals and their ectoparasites, to determine their pathogenicity as well as their potential effects on public health in order to update the national policy for the control of emerging infectious diseases in Madagascar.

Local-scale diversity of Yersinia pestis: A case study from Ambohitromby, Ankazobe District, Madagascar.

Plague is a re-emerging zoonotic disease and a major public health concern in several portions of the world, especially in Madagascar. We report on the presence of different subtypes of Yersinia pestis co-occurring in the same locality. After confirmation of a human plague case in Ambohitromby Commune (Ankazobe District) via isolation of Y. pestis, we undertook small mammal trapping to identify the circulation of Y. pestis amongst rodents in this locality; blood samples were collected from rodents for seroprevalence analysis. Of the 60 individuals of Rattus rattus captured, one yielded an isolate of Y. pestis, 13 others were positive for F1 antigen of Y. pestis using a rapid diagnostic test, and 4 were PCR positive targeting the caf1 and pla genes; 28/60 (46.7%) of the captured R. rattus were seropositive for Y. pestis. Whole-genome SNP analyses revealed that the two isolates obtained from the human case, and the R. rattus belonged to two different subtypes of Y. pestis (s05 and s13, respectively) that were circulating concurrently in Ambohitromby in 2016. Three Y. pestis subtypes (s03, s05 and s13) have now been isolated from Ambohitromby. Subtype s05 had been persisting there for >10 years but one or both of the other subtypes may have been introduced from the Central Highlands region as they were not observed in previous years (s13) or only observed once previously (s03). High seroprevalence against Y. pestis in R. rattus suggests that a portion of the local murine population may have acquired resistance to Y. pestis. Future research should focus on genomically characterizing Y. pestis strains circulating in Ankazobe District and other plague-endemic regions of Madagascar to better understand the overall phylogeography of Y. pestis.

Rodent control to fight plague: field assessment of methods based on rat density reduction.

Rodents represent a serious threat to food security and public health. The extent to which rodent control can mitigate the risk from rodent-borne disease depends on both the effectiveness of control in reducing rodent abundance and the impact on disease epidemiology. Focusing on a plague-endemic region of Madagascar, this study compared the effectiveness of 3 methods: live-traps, snap-traps, and rodenticides. Control interventions were implemented inside houses between May and October 2019. Tracking tiles monitored rodent abundance. Rodent fleas, the vector involved in plague transmission, were collected. Rodent populations consisted of Rattus rattus and Mus musculus. In terms of trap success, we found that our live-trap regime was more effective than snap-traps. While all 3 control strategies appeared to reduce in-house rodent activity in the short term, we found no evidence of a longer-term effect, with in-house rodent abundance in treated sites comparable to non-treatment sites by the following month. Endemic flea, Synopsyllus fonquerniei, is a key plague vector usually found on rats living outdoors. Although we found no evidence that its abundance inside houses increased following control, this may have been due to a lack of power caused by significant variation in S. fonquerniei abundance. The presence of S. fonquerniei in houses was more likely when S. fonquerniei abundance on outdoor rats was higher, which in turn correlated with high rat abundance. Our results emphasize that control strategies need to consider this connectivity between in-house rat-flea populations and the outdoor populations, and any potential consequences for plague transmission.

Plague reservoir species throughout the world.

Plague has been known since ancient times as a re-emerging infectious disease, causing considerable socioeconomic burden in regional hotspots. To better understand the epidemiological cycle of the causative agent of the plague, its potential occurrence, and possible future dispersion, one must carefully consider the taxonomy, distribution, and ecological requirements of reservoir-species in relation either to natural or human-driven changes (e.g. climate change or urbanization). In recent years, the depth of knowledge on species taxonomy and species composition in different landscapes has undergone a dramatic expansion, driven by modern taxonomic methods such as synthetic surveys that take into consideration morphology, genetics, and the ecological setting of captured animals to establish their species identities. Here, we consider the recent taxonomic changes of the rodent species in known plague reservoirs and detail their distribution across the world, with a particular focus on those rodents considered to be keystone host species. A complete checklist of all known plague-infectable vertebrates living in plague foci is provided as a Supporting Information table.

Bat coronavirus phylogeography in the Western Indian Ocean.

Bats provide key ecosystem services such as crop pest regulation, pollination, seed dispersal, and soil fertilization. Bats are also major hosts for biological agents responsible for zoonoses, such as coronaviruses (CoVs). The islands of the Western Indian Ocean are identified as a major biodiversity hotspot, with more than 50 bat species. In this study, we tested 1,013 bats belonging to 36 species from Mozambique, Madagascar, Mauritius, Mayotte, Reunion Island and Seychelles, based on molecular screening and partial sequencing of the RNA-dependent RNA polymerase gene. In total, 88 bats (8.7%) tested positive for coronaviruses, with higher prevalence in Mozambican bats (20.5% ± 4.9%) as compared to those sampled on islands (4.5% ± 1.5%). Phylogenetic analyses revealed a large diversity of α- and ß-CoVs and a strong signal of co-evolution between CoVs and their bat host species, with limited evidence for host-switching, except for bat species sharing day roost sites. These results highlight that strong variation between islands does exist and is associated with the composition of the bat species community on each island. Future studies should investigate whether CoVs detected in these bats have a potential for spillover in other hosts.

Human plague: An old scourge that needs new answers.

Yersinia pestis, the bacterial causative agent of plague, remains an important threat to human health. Plague is a rodent-borne disease that has historically shown an outstanding ability to colonize and persist across different species, habitats, and environments while provoking sporadic cases, outbreaks, and deadly global epidemics among humans. Between September and November 2017, an outbreak of urban pneumonic plague was declared in Madagascar, which refocused the attention of the scientific community on this ancient human scourge. Given recent trends and plague's resilience to control in the wild, its high fatality rate in humans without early treatment, and its capacity to disrupt social and healthcare systems, human plague should be considered as a neglected threat. A workshop was held in Paris in July 2018 to review current knowledge about plague and to identify the scientific research priorities to eradicate plague as a human threat. It was concluded that an urgent commitment is needed to develop and fund a strong research agenda aiming to fill the current knowledge gaps structured around 4 main axes: (i) an improved understanding of the ecological interactions among the reservoir, vector, pathogen, and environment; (ii) human and societal responses; (iii) improved diagnostic tools and case management; and (iv) vaccine development. These axes should be cross-cutting, translational, and focused on delivering context-specific strategies. Results of this research should feed a global control and prevention strategy within a "One Health" approach.

Epidemiological characteristics of an urban plague epidemic in Madagascar, August-November, 2017: an outbreak report.

BACKGROUND: Madagascar accounts for 75% of global plague cases reported to WHO, with an annual incidence of 200-700 suspected cases (mainly bubonic plague). In 2017, a pneumonic plague epidemic of unusual size occurred. The extent of this epidemic provides a unique opportunity to better understand the epidemiology of pneumonic plagues, particularly in urban settings. METHODS: Clinically suspected plague cases were notified to the Central Laboratory for Plague at Institut Pasteur de Madagascar (Antananarivo, Madagascar), where biological samples were tested. Based on cases recorded between Aug 1, and Nov 26, 2017, we assessed the epidemiological characteristics of this epidemic. Cases were classified as suspected, probable, or confirmed based on the results of three types of diagnostic tests (rapid diagnostic test, molecular methods, and culture) according to 2006 WHO recommendations. FINDINGS: 2414 clinically suspected plague cases were reported, including 1878 (78%) pneumonic plague cases, 395 (16%) bubonic plague cases, one (<1%) septicaemic case, and 140 (6%) cases with unspecified clinical form. 386 (21%) of 1878 notified pneumonic plague cases were probable and 32 (2%) were confirmed. 73 (18%) of 395 notified bubonic plague cases were probable and 66 (17%) were confirmed. The case fatality ratio was higher among confirmed cases (eight [25%] of 32 cases) than probable (27 [8%] of 360 cases) or suspected pneumonic plague cases (74 [5%] of 1358 cases) and a similar trend was seen for bubonic plague cases (16 [24%] of 66 confirmed cases, four [6%] of 68 probable cases, and six [2%] of 243 suspected cases). 351 (84%) of 418 confirmed or probable pneumonic plague cases were concentrated in Antananarivo, the capital city, and Toamasina, the main seaport. All 50 isolated Yersinia pestis strains were susceptible to the tested antibiotics. INTERPRETATION: This predominantly urban plague epidemic was characterised by a large number of notifications in two major urban areas and an unusually high proportion of pneumonic forms, with only 23% having one or more positive laboratory tests. Lessons about clinical and biological diagnosis, case definition, surveillance, and the logistical management of the response identified in this epidemic are crucial to improve the response to future plague outbreaks. FUNDING: US Agency for International Development, WHO, Institut Pasteur, US Department of Health and Human Services, Laboratoire d'Excellence Integrative Biology of Emerging Infectious Diseases, Models of Infectious Disease Agent Study of the National Institute of General Medical Sciences, AXA Research Fund, and the INCEPTION programme.

Biogeography of Leptospira in wild animal communities inhabiting the insular ecosystem of the western Indian Ocean islands and neighboring Africa.

Understanding the processes driving parasite assemblages is particularly important in the context of zoonotic infectious diseases. Leptospirosis is a widespread zoonotic bacterial infection caused by pathogenic species of the genus Leptospira. Despite a wide range of animal hosts, information is still lacking on the factors shaping Leptospira diversity in wild animal communities, especially in regions, such as tropical insular ecosystems, with high host species richness and complex biogeographical patterns. Using a large dataset (34 mammal species) and a multilocus approach at a regional scale, we analyzed the role of both host species diversity and geography in Leptospira genetic diversity in terrestrial small mammals (rodents, tenrecs, and shrews) and bats from 10 different islands/countries in the western Indian Ocean (WIO) and neighboring Africa. At least four Leptospira spp. (L. interrogans, L. borgpetersenii, L. kirschneri, and L. mayottensis) and several yet-unidentified genetic clades contributed to a remarkable regional Leptospira diversity, which was generally related to the local occurrence of the host species rather than the geography. In addition, the genetic structure patterns varied between Leptospira spp., suggesting different evolutionary histories in the region, which might reflect both in situ diversification of native mammals (for L. borgpetersenii) and the more recent introduction of non-native host species (for L. interrogans). Our data also suggested that host shifts occurred between bats and rodents, but further investigations are needed to determine how host ecology may influence these events.