An update on snail and trematode communities in the Sanyati Basin of Lake Kariba: New snail and trematode species but no human schistosomes.

BACKGROUND: The construction of Lake Kariba brought about a rise in the incidence of schistosomiasis in its surrounding towns of Kariba (Zimbabwe) and Siavonga (Zambia). After extensive control programs in Kariba, schistosomiasis prevalence dropped significantly. The objective of this study was to revisit the same localities sampled by Chimbari et al. (2003), and provide an update on the snail community and prevalence of trematodes in the Northern shore of Lake Kariba while focusing on planorbid species. METHODS: Monthly sampling of snails at 16 sites along the Northern shoreline of Lake Kariba, near Kariba town, was undertaken for one year. Minimum one specimen per morphotype was identified using molecular barcoding (sequencing a fragment of cytochrome c oxidase I subunit (COI)). The infection status of snails was assessed by Rapid Diagnostic PCRs (RD-PCR), and trematode infections were genotyped by sequencing COI and 18S rDNA markers. RESULTS: We collected and identified seven snail species: Bulinus truncatus, Bulinus forskalii, Gyraulus sp., Physella acuta, Bellamya sp., Radix affinis plicatula and Pseudosuccinea columella. Physella acuta was the most abundant snail species (comprising 56.95% of the total snail count) and present at all sites. The B. truncatus population was found to be infected with the stomach fluke Carmyerius cruciformis, a Petasiger sp. and a trematode species belonging to the family Notocotylidae. No Schistosoma sp. infections were detected in our collected snail specimens. CONCLUSIONS: We report B. truncatus as an intermediate snail host for Carmyerius cruciformis, and the presence of three non-schistosome trematode species that have not been reported in Lake Kariba before. Furthermore, we detect a possible shift in the snail community when compared to the report by Chimbari et al. (2003): this is the first record of Gyraulus sp. in Lake Kariba, and we did not observe the previously reported B. pfeifferi, B. globosus and Radix natalensis. Although this shift in snail communities might have contributed to the absence of Schistosoma spp. detection in this study, further monitoring of final and intermediate hosts across the Kariba basin is essential to prove a decrease of schistosomiasis in the area.

An accident waiting to happen? Exposing the potential of urogenital schistosomiasis transmission in the Lake Albert region, Uganda.

BACKGROUND: Urogenital schistosomiasis caused by the parasitic blood fluke Schistosoma haematobium is the most common form of that constitutes a majority of over 240 million schistosomiasis cases. The enigmatic absence of urogenital schistosomiasis in Uganda has, until now, been attributed to the absence of substantial populations of suitable snail intermediate hosts. METHODS: Malacological surveys were carried out in 73 sites southeast of Lake Albert, Uganda in October and November 2020. Collected snails were transported to the laboratory for identification. The snails were identified using partial mitochondrial cytochrome c oxidase subunit one and nuclear internal transcribed spacer barcoding. Schistosome infections in snails were also assessed using cercarial shedding and rapid diagnostic PCR techniques. RESULTS: We found Bulinus globosus and Bulinus nasutus productus, the main intermediate species in the transmission of S. haematobium in mainland East Africa. In this survey, B. globosus was more common than B. nasutus productus, with the former reported at four sites (total count = 188) and the latter reported at one site (total count = 79). Molecular testing revealed a high prevalence of Schistosoma bovis in B. nasutus productus (16%), but no S. haematobium infections were found. CONCLUSIONS: Given the abundance of snail hosts and the risky human water contact behaviours observed, we highlight the potential for urogenital schistosomiasis transmission in the region.

Trematode infection affects shell shape and size in Bulinus tropicus.

Trematodes can increase intraspecific variation in the phenotype of their intermediate snail host. However, the extent of such phenotypic changes remains unclear. We investigated the influence of trematode infection on the shell morphology of Bulinus tropicus, a common host of medically important trematodes. We focused on a snail population from crater lake Kasenda (Uganda). We sampled a single homogeneous littoral habitat to minimize the influence of environmental variation on shell phenotype, and barcoded snails to document snail genotypic variation. Among the 257 adult snails analysed, 99 tested positive for trematode infection using rapid-diagnostic PCRs. Subsequently we used high-throughput amplicon sequencing to identify the trematode (co-)infections. For 86 out of the 99 positive samples trematode species delineation could discriminate among combinations of (co-)infection by 11 trematode species. To avoid confounding effects, we focused on four prevalent trematode species. We performed landmark-based geometric morphometrics to characterize shell phenotype and used regressions to examine whether shell size and shape were affected by trematode infection and the developmental stage of infection (as inferred from read counts). Snails infected by Petasiger sp. 5, Echinoparyphium sp. or Austrodiplostomum sp. 2 had larger shells than uninfected snails or than those infected by Plagiorchiida sp. Moreover, the shell shape of snails infected solely by Petasiger sp. 5 differed significantly from that of uninfected snails and snails infected with other trematodes, except from Austrodiplostomum sp. 2. Shape changes included a more protuberant apex, an inward-folded outer apertural lip and a more adapically positioned umbilicus. Size differences were more pronounced in snails with 'late' infections (>25 days) compared to earlier-stage infections. No phenotypic differences were found between snails infected by a single trematode species and those harbouring co-infections. Further work is required to assess the complex causal links between trematode infections and shell morphological alterations of snail hosts.

Simultaneous genotyping of snails and infecting trematode parasites using high-throughput amplicon sequencing.

Several methodological issues currently hamper the study of entire trematode communities within populations of their intermediate snail hosts. Here we develop a new workflow using high-throughput amplicon sequencing to simultaneously genotype snail hosts and their infecting trematode parasites. We designed primers to amplify four snail and five trematode markers in a single multiplex PCR. While also applicable to other genera, we focused on medically and economically important snail genera within the superorder Hygrophila and targeted a broad taxonomic range of parasites within the class Trematoda. We tested the workflow using 417 Biomphalaria glabrata specimens experimentally infected with Schistosoma rodhaini, two strains of Schistosoma mansoni and combinations thereof. We evaluated the reliability of infection diagnostics, the robustness of the workflow, its specificity related to host and parasite identification, and the sensitivity to detect co-infections, immature infections and changes of parasite biomass during the infection process. Finally, we investigated its applicability in wild-caught snails of other genera naturally infected with a diverse range of trematodes. After stringent quality control the workflow allows the identification of snails to species level, and of trematodes to taxonomic levels ranging from family to strain. It is sensitive to detect immature infections and changes in parasite biomass described in previous experimental studies. Co-infections were successfully identified, opening the possibility to examine parasite-parasite interactions such as interspecific competition. Together, these results demonstrate that our workflow provides a powerful tool to analyse the processes shaping trematode communities within natural snail populations.

Invasive snails, parasite spillback, and potential parasite spillover drive parasitic diseases of Hippopotamus amphibius in artificial lakes of Zimbabwe.

BACKGROUND: Humans impose a significant pressure on large herbivore populations, such as hippopotami, through hunting, poaching, and habitat destruction. Anthropogenic pressures can also occur indirectly, such as artificial lake creation and the subsequent introduction of invasive species that alter the ecosystem. These events can lead to drastic changes in parasite diversity and transmission, but generally receive little scientific attention. RESULTS: In order to document and identify trematode parasites of the common hippopotamus (Hippopotamus amphibius) in artificial water systems of Zimbabwe, we applied an integrative taxonomic approach, combining molecular diagnostics and morphometrics on archived and new samples. In doing so, we provide DNA reference sequences of the hippopotamus liver fluke Fasciola nyanzae, enabling us to construct the first complete Fasciola phylogeny. We describe parasite spillback of F. nyanzae by the invasive freshwater snail Pseudosuccinea columella, as a consequence of a cascade of biological invasions in Lake Kariba, one of the biggest artificial lakes in the world. Additionally, we report an unknown stomach fluke of the hippopotamus transmitted by the non-endemic snail Radix aff. plicatula, an Asian snail species that has not been found in Africa before, and the stomach fluke Carmyerius cruciformis transmitted by the native snail Bulinus truncatus. Finally, Biomphalaria pfeifferi and two Bulinus species were found as new snail hosts for the poorly documented hippopotamus blood fluke Schistosoma edwardiense. CONCLUSIONS: Our findings indicate that artificial lakes are breeding grounds for endemic and non-endemic snails that transmit trematode parasites of the common hippopotamus. This has important implications, as existing research links trematode parasite infections combined with other stressors to declining wild herbivore populations. Therefore, we argue that monitoring the anthropogenic impact on parasite transmission should become an integral part of wildlife conservation efforts.

Past connections with the mainland structure patterns of insular species richness in a continental-shelf archipelago (Aegean Sea, Greece).

Recent research in island biogeography has highlighted the important role of late Quaternary sea-level fluctuations in shaping biogeographic patterns in insular systems but focused on oceanic systems. Through this study, we aim investigate how late Quaternary sea-level fluctuations shaped species richness patterns in continental-shelf island systems. Focusing on the Aegean archipelago, we first compiled maps of the area's geography using published data, under three sea-level stands: (a) current; (b) median sea-level over the last nine glacial-interglacial cycles (MSL); and (c) Last Glacial Maximum (LGM). We gathered taxon-island occurrences for multiple chorotypes of angiosperms, butterflies, centipedes, and reptiles. We investigated the impact of present-day and past geographic settings on chorological groups by analyzing island species-area relationships (ISARs) and using generalized linear mixed models (GLMMs) selection based on multiple metrics of goodness of fit. Our results confirm that the Aegean's geography has changed dramatically since the LGM, whereas the MSL only modestly differs from the present configuration. Apart for centipedes, paleogeographic changes affected both native and endemic species diversity through altering connections between land-bridge islands and the mainland. On land-bridge islands, we detected over-representation of native species and under-representation of endemics. Unlike oceanic islands, sea-level-driven increase of isolation and area contraction did not strongly shape patterns of species richness. Furthermore, the LGM configurations rather than the MSL configuration shaped patterns of endemic species richness. This suggests that even short episodes of increased connectivity with continental populations are sufficient to counteract the genetic differentiation of insular populations. On the other hand, the over-representation of native nonendemic species on land-bridge islands reflected MSL rather than LGM mainland connections. Our study shows that in terms of processes affecting species richness patterns, continental archipelagos differ fundamentally from oceanic systems because episodic connections with the mainland have profound effects on the biota of land-bridge islands.

Exposing the Barcoding Void: An Integrative Approach to Study Snail-Borne Parasites in a One Health Context.

Trematodes are snail-borne parasites of major zoonotic importance that infect millions of people and animals worldwide and frequently hybridize with closely related species. Therefore, it is desirable to study trematodiases in a One Health framework, where human and animal trematodes are considered equally important. It is within this framework that we set out to study the snail and trematode communities in four artificial lakes and an abattoir in Zimbabwe. Trematode infections in snails were detected through multiplex PCR protocols. Subsequently, we identified snails by sequencing a partial mitochondrial cytochrome c oxidase subunit I (COI) fragment, and trematodes (adults from the abattoir and larval stages detected in snails) using COI and nuclear rDNA markers. Of the 1,674 collected snails, 699 were molecularly analyzed, in which we identified 12 snail and 19 trematode species. Additionally, three parasite species were sampled from the abattoir. Merely four trematode species were identified to species level through COI-based barcoding. Moreover, identification of members of the superfamilies Opisthorchioidea and Plagiorchioidea required a phylogenetic inference using the highly conserved 18S rDNA marker, as no related COI reference sequences were present in public databases. These barcoding challenges demonstrate a severe barcoding void in the available databases, which can be attributed to the neglected status of trematodiases. Adding to this, many available sequences cannot be used as different studies use different markers. To fill this gap, more studies on African trematodes, using a standardized COI barcoding region, are desperately needed.

Molecular identification of Bulinus spp. intermediate host snails of Schistosoma spp. in crater lakes of western Uganda with implications for the transmission of the Schistosoma haematobium group parasites.

BACKGROUND: Human schistosomiasis is the second most important tropical disease and occurs in two forms in Africa (intestinal and urogenital) caused by the digenetic trematodes Schistosoma mansoni and Schistosoma haematobium, respectively. A proposed recent shift of schistosomiasis above a previously established altitudinal threshold of 1400 m above sea level in western Ugandan crater lakes has triggered more research interest there. METHODS: Based on extensive field sampling in western Uganda and beyond and employing an approach using sequences of the mitochondrial barcoding gene cytochrome c oxidase subunit 1 (cox1) this study aims were: (i) identification and establishment of the phylogenetic affinities of Bulinus species as potential hosts for Schistosoma spp.; (ii) determining diversity, frequency and distribution patterns of Bulinus spp.; and (iii) establishing genetic variability and phylogeographical patterns using Bayesian inference and parsimony network analyses. RESULTS: Out of the 58 crater lakes surveyed, three species of Bulinus snails were found in 34 crater lakes. Bulinus tropicus was dominating, Bulinus forskalii was found in two lakes and Bulinus truncatus in one. The latter two species are unconfirmed potential hosts for S. haematobium in this region. However, Bulinus tropicus is an important species for schistosomiasis transmission in ruminants. Bulinus tropicus comprised 31 haplotypes while both B. forskalii and B. truncatus exhibited only a single haplotype in the crater lakes. All species clustered with most of the haplotypes from surrounding lake systems forming source regions for the colonization of the crater lakes. CONCLUSIONS: This first detailed malacological study of the crater lakes systems in western Uganda revealed presence of Bulinus species that are either not known or not regionally known to be hosts for S. haematobium, the causing agent of human urogenital schistosomiasis. Though this disease risk is almost negligible, the observed dominance of B. tropicus in the crater lakes shows that there is a likelihood of a high risk of infections with Schistosoma bovis. Thus, extra attention should be accorded to safeguard wild and domestic ruminants in this region as the population benefits from these animals.

A rapid diagnostic multiplex PCR approach for xenomonitoring of human and animal schistosomiasis in a 'One Health' context.

Studying the epidemiology of schistosomiasis-the most prevalent gastropod-borne human disease and an economic burden for the livestock industry-relies on adequate monitoring tools. Here we describe a molecular assay for detecting human and animal African schistosome species in their planorbid gastropod host (xenomonitoring) using a two-step approach. First, schistosome infections are detected and discriminated from other trematode infections using a multiplex polymerase chain reaction (PCR) that includes a trematode-specific marker (in 18S rDNA), a Schistosoma genus-specific marker (in internal transcribed spacer 2 [ITS2]) and a general gastropod marker (in 18S rDNA) as an internal control. Upon Schistosoma sp. detection, a second multiplex PCR is performed to discriminate among Schistosoma haematobium, Schistosoma mansoni, Schistosoma mattheei and Schistosoma bovis/Schistosoma curassoni/Schistosoma guineensis using markers of differential lengths in the cytochrome c oxidase subunit 1 (COX1) gene. The specificity of these assays was validated with adult worms, naturally infected gastropods and human urine and stool samples. Sensitivity was tested on experimentally infected snail specimens that were sacrificed 10 and 40 days post-infection in order to mimic a natural prepatent and mature infection, respectively. The assay provides a diagnostic tool to support the xenomonitoring of planorbid gastropods for trematode infections in a One Health context, with a focus on the transmission monitoring of schistosomiasis.

A cascade of biological invasions and parasite spillback in man-made Lake Kariba.

Parasite spillback, the infection of a non-indigenous organism by a native parasite, is a highly important although understudied component of ecological invasion dynamics. Here, through the first analysis of the parasite fauna of lymnaeid gastropods of Lake Kariba (Zimbabwe). We illustrate how the creation of an artificial lake may lead to a cascade of biological invasions in which an invasive aquatic plant promotes the proliferation of invasive gastropods, which in turn alters the epidemiology of trematodiases of potential medical and veterinary importance. Using a new multiplex Rapid Diagnostic PCR assay, we assessed the prevalence of Fasciola sp. infections in the gastropod populations. Both gastropod hosts and trematode parasites were identified using DNA barcoding. We provide the first record of the invasive North-American gastropod Pseudosuccinea columella in Lake Kariba. This species was found at 14 out of 16 sampled sites and its abundance was strongly positively correlated with the abundance of the invasive South-American water hyacinth (Eichhornia crassipes). About 65% of the P. columella specimens analysed were infected with a hitherto unknown Fasciola species. Phylogenetic analyses indicate close affinity to Fasciola hepatica and F. gigantica, which cause fasciolosis, an important liver disease affecting both ruminants and humans. In addition, another non-native Lymnaeid species was found: a Radix sp. that clustered closely with a Vietnamese Radix species. Radix sp. hosted both amphistome and Fasciola trematodes. By linking an invasion cascade and parasite spillback, this study shows how both processes can act in combination to lead to potentially important epidemiological changes.