Leptospires, which cause the zoonotic disease leptospirosis, persist in soil and aqueous environments. Several factors, including rainfall, the presence of reservoir animals, and various abiotic and biotic components interact to influence leptospiral survival, persistence, and pathogenicity in the environment. However, how these factors modulate the risk of infection is poorly understood. Here we developed an approach using environmental DNA (eDNA) metabarcoding for detecting the microbiome, vertebrates, and pathogenic Leptospira in aquatic samples. Specifically, we combined 4 sets of primers to generate PCR products for high-throughput sequencing of multiple amplicons through next-generation sequencing. Using our method to analyze the eDNA of leptospirosis-endemic areas in northern Okinawa, Japan, we found that the microbiota in each river shifted over time. Operating taxonomic units corresponding to pathogenic L . alstonii , L . kmetyi , and L . interrogans were detected in association with 12 nonpathogenic bacterial species. In addition, the frequencies of 11 of these species correlated with the amount of rainfall. Furthermore, 10 vertebrate species, including Sus scrofa , Pteropus dasymallus , and Cynops ensicauda , showed high correlation with leptospiral eDNA detection. Our eDNA metabarcoding method is a powerful tool for understanding the environmental phase of Leptospira and predicting human infection risk.

SummaryLeptospira interrogans is responsible for the zoonotic disease leptospirosis. The pathogenic mechanisms of this spirochaete remain poorly understood; however, virulence has been correlated with increased phagocytic uptake and survival within macrophages. Leptospiral outer membrane proteins are thought to be responsible for persistence in vivo via interaction with specific host components. In this study, we analysed the transcriptional profile of a virulent strain and its culture-attenuated derivative strain to identify bacterial factors that may be involved in pathogenesis. Two outer membrane proteins, LMB216 and LigB (leptospiral immunoglobulin-like protein B) were downregulated more than 10-fold in the culture-attenuated strain. We show that both proteins play a role in leptospiral uptake by macrophages and that LMB216, as well as LigB, enhances the binding of leptospires to fibronectin. Taken together, our results indicate that LMB216 plays a role in pathogen interaction with host molecule/s, which may contribute to pathogenesis of leptospirosis.

Little is known about effects of large storm systems on mesophotic reefs. This study reports on how Typhoon 17 (Jelawat) affected Ryugu Reef on Okinawa-jima, Japan in September 2012. Benthic communities were surveyed before and after the typhoon using line intercept transect method. Comparison of the benthic assemblages showed highly significant differences in coral coverage at depths of 25–32 m before and after Typhoon 17. A large deep stand of Pachyseris foliosa was apparently less resistant to the storm than the shallower high diversity area of this reef. Contradictory to common perception, this research shows that large foliose corals at deeper depths are just as susceptible to typhoon damage as shallower branching corals. However, descriptive functional group analyses resulted in only minor changes after the disturbance, suggesting the high likelihood of recovery and the high resilience capacity of this mesophotic reef.

Okinawa, Japan is known for its high marine biodiversity, yet little work has been performed on examining impacts of numerous large-scale coastal development projects on its marine ecosystems. Here, we examine apparent impacts of the construction of the Kaichu-Doro causeway, which was built over 40 years ago. The causeway is a 4.75 km long embankment that divides a large tidal flat and has only two points of water exchange along its entire length. We employed quadrats, transects, sampling, visual surveys, and microbial community analyses combined with environmental, water quality data, and 1m cores, at five stations of two paired sites each (one on each side of Kaichu-Doro) to investigate how the environment and biota have changed since the Kaichu-Doro was built. Results indicate reduction in water flow, and site S1 was particularly heavily impacted by poor water quality, with low diversity and disturbed biotic communities.

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