The hemolymph of Biomphalaria snail vectors of schistosomiasis supports a diverse microbiome.

The microbiome - the microorganism community that is found on or within an organism's body - is increasingly recognized to shape many aspects of its host biology and is a key determinant of health and disease. Microbiomes modulate the capacity of insect disease vectors (mosquitoes, tsetse flies, sandflies) to transmit parasites and disease. We investigate the diversity and abundance of microorganisms within the hemolymph (i.e. blood) of Biomphalaria snails, the intermediate host for Schistosoma mansoni, using Illumina MiSeq sequencing of the bacterial 16S V4 rDNA. We sampled hemolymph from five snails from six different laboratory populations of B. glabrata and one population of B. alexandrina. We observed 279.84 ± 0.79 amplicon sequence variants per snail. There were significant differences in microbiome composition at the level of individual snails, snail populations and species. Snail microbiomes were dominated by Proteobacteria and Bacteroidetes while water microbiomes from snail tank were dominated by Actinobacteria. We investigated the absolute bacterial load using qPCR: hemolymph samples contained 2784 ± 339 bacteria/µl. We speculate that the microbiome may represent a critical, but unexplored intermediary in the snail-schistosome interaction as hemolymph is in very close contact with the parasite at each step of its development.

Rapid phenotypic and genotypic change in a laboratory schistosome population.

Background: Genomic analysis has revealed extensive contamination among laboratory-maintained microbes including malaria parasites, Mycobacterium tuberculosis and Salmonella spp. Here, we provide direct evidence for recent contamination of a laboratory schistosome parasite population, and we investigate its genomic consequences. The Brazilian Schistosoma mansoni population SmBRE has several distinctive phenotypes, showing poor infectivity, reduced sporocysts number, low levels of cercarial shedding and low virulence in the intermediate snail host, and low worm burden and low fecundity in the vertebrate rodent host. In 2021 we observed a rapid change in SmBRE parasite phenotypes, with a~10x increase in cercarial production and ~ 4x increase in worm burden. Methods: To determine the underlying genomic cause of these changes, we sequenced pools of SmBRE adults collected during parasite maintenance between 2015 and 2023. We also sequenced another parasite population (SmLE) maintained alongside SmBRE without phenotypic changes. Results: While SmLE allele frequencies remained stable over the eight-year period, we observed sudden changes in allele frequency across the genome in SmBRE between July 2021 and February 2023, consistent with expectations of laboratory contamination. (i) SmLE-specific alleles rose in the SmBRE population from 0 to 41-46% across the genome between September and October 2021, documenting the timing and magnitude of the contamination event. (ii) After contamination, strong selection (s=~ 0.23) drove replacement of low fitness SmBRE with high fitness SmLE alleles. (iii) Allele frequency changed rapidly across the whole genome, except for a region on chromosome 4 where SmBRE alleles remained at high frequency. Conclusions: We were able to detect contamination in this case because SmBRE shows distinctive phenotypes. However, this would likely have been missed with phenotypically similar parasites. These results provide a cautionary tale about the importance of tracking the identity of parasite populations, but also showcase a simple approach to monitor changes within populations using molecular profiling of pooled population samples to characterize fixed single nucleotide polymorphisms. We also show that genetic drift results in continuous change even in the absence of contamination, causing parasites maintained in different labs (or sampled from the same lab at different times) to diverge.

Rapid phenotypic and genotypic change in a laboratory schistosome population.

Background: Genomic analysis has revealed extensive contamination among laboratory-maintained microbes including malaria parasites, Mycobacterium tuberculosis and Salmonella spp. Here, we provide direct evidence for recent contamination of a laboratory schistosome parasite population, and we investigate its genomic consequences. The Brazilian Schistosoma mansoni population SmBRE has several distinctive phenotypes, showing poor infectivity, reduced sporocysts number, low levels of cercarial shedding and low virulence in the intermediate snail host, and low worm burden and low fecundity in the vertebrate rodent host. In 2021 we observed a rapid change in SmBRE parasite phenotypes, with a ~10x increase in cercarial production and ~4x increase in worm burden. Methods: To determine the underlying genomic cause of these changes, we sequenced pools of SmBRE adults collected during parasite maintenance between 2015 and 2023. We also sequenced another parasite population (SmLE) maintained alongside SmBRE without phenotypic changes. Results: While SmLE allele frequencies remained stable over the eight-year period, we observed sudden changes in allele frequency across the genome in SmBRE between July 2021 and February 2023, consistent with expectations of laboratory contamination. (i) SmLE-specific alleles rose in the SmBRE population from 0 to 41-46% across the genome between September and October 2021, documenting the timing and magnitude of the contamination event. (ii) After contamination, strong selection (s = ~0.23) drove replacement of low fitness SmBRE with high fitness SmLE alleles. (iii) Allele frequency changed rapidly across the whole genome, except for a region on chromosome 4 where SmBRE alleles remained at high frequency. Conclusions: We were able to detect contamination in this case because SmBRE shows distinctive phenotypes. However, this would likely have been missed with phenotypically similar parasites. These results provide a cautionary tale about the importance of tracking the identity of parasite populations, but also showcase a simple approach to monitor changes within populations using molecular profiling of pooled population samples to characterize fixed single nucleotide polymorphisms. We also show that genetic drift results in continuous change even in the absence of contamination, causing parasites maintained in different labs (or sampled from the same lab at different times) to diverge.

Abundant genetic variation is retained in many laboratory schistosome populations.

Schistosomes are obligately sexual blood flukes that can be maintained in the laboratory using freshwater snails as intermediate and rodents as definitive hosts. The genetic composition of laboratory schistosome populations is poorly understood: whether genetic variation has been purged due to serial inbreeding or retained is unclear. We sequenced 19 - 24 parasites from each of five laboratory Schistosoma mansoni populations and compared their genomes with published exome data from four S. mansoni field populations. We found abundant genomic variation (0.897 - 1.22 million variants) within laboratory populations: these retained on average 49% (π = 3.27e-04 - 8.94e-04) of the nucleotide diversity observed in the four field parasite populations (π = 1.08e-03 - 2.2e-03). However, the pattern of variation was very different in laboratory and field populations. Tajima's D was positive in all laboratory populations except SmBRE, indicative of recent population bottlenecks, but negative in all field populations. Current effective population size estimates of laboratory populations were lower (2 - 258) compared to field populations (3,174 - infinity). The distance between markers at which linkage disequilibrium (LD) decayed to 0.5 was longer in laboratory populations (59 bp - 180 kb) compared to field populations (9 bp - 9.5 kb). SmBRE was the least variable; this parasite also shows low fitness across the lifecycle, consistent with inbreeding depression. The abundant genetic variation present in most laboratory schistosome populations has several important implications: (i) measurement of parasite phenotypes, such as drug resistance, using laboratory parasite populations will determine average values and underestimate trait variation; (ii) genome-wide association studies (GWAS) can be conducted in laboratory schistosome populations by measuring phenotypes and genotypes of individual worms; (iii) genetic drift may lead to divergence in schistosome populations maintained in different laboratories. We conclude that the abundant genetic variation retained within many laboratory schistosome populations can provide valuable, untapped opportunities for schistosome research.

Genomic data reveal a north-south split and introgression history of blood fluke (Schistosoma haematobium) populations from across Africa.

The human parasitic fluke, Schistosoma haematobium hybridizes with the livestock parasite S. bovis in the laboratory, but the extent of hybridization in nature is unclear. We analyzed 34.6 million single nucleotide variants in 162 samples from 18 African countries, revealing a sharp genetic discontinuity between northern and southern S. haematobium. We found no evidence for recent hybridization. Instead the data reveal admixture events that occurred 257-879 generations ago in northern S. haematobium populations. Fifteen introgressed S. bovis genes are approaching fixation in northern S. haematobium with four genes potentially driving adaptation. We identified 19 regions that were resistant to introgression; these were enriched on the sex chromosomes. These results (i) demonstrate strong barriers to gene flow between these species, (ii) indicate that hybridization may be less common than currently envisaged, but (iii) reveal profound genomic consequences of interspecific hybridization between schistosomes of medical and veterinary importance.

Genetic analysis of praziquantel response in schistosome parasites implicates a transient receptor potential channel.

Mass drug administration with praziquantel (PZQ) monotherapy is considered the mainstay for control and elimination of the parasites causing schistosomiasis in humans. This drug shows imperfect cure rates in the field, and parasites showing reduced PZQ response can be selected in the laboratory, but the extent of resistance in Schistosoma mansoni populations is unknown. We examined the genetic basis of the variation in response in a PZQ-selected S. mansoni population (SmLE-PZQ-R) in which 35% of the parasitic worms survive high-dose PZQ (73 micrograms per milliliter) treatment. We used genome-wide association to map loci underlying PZQ response and identified a transient receptor potential (Sm.TRPMPZQ) channel (Smp_246790) within the major chromosome 3 peak that is activated by nanomolar concentrations of PZQ. The PZQ response showed recessive inheritance and marker-assisted selection of parasites at a single Sm.TRPMPZQ SNP that produced populations of PZQ-enriched resistant (PZQ-ER) and PZQ-enriched sensitive (PZQ-ES) parasites, exhibiting >377-fold difference in PZQ response. The PZQ-ER parasites survived treatment in rodents at higher frequencies compared with PZQ-ES, and resistant parasites exhibited 2.25-fold lower expression of Sm.TRPMPZQ relative to sensitive parasites. Specific chemical blockers of Sm.TRPMPZQ enhanced PZQ resistance, whereas Sm.TRPMPZQ activators increased sensitivity. We surveyed Sm.TRPMPZQ sequence variations in 259 parasites from different global sites and identified one nonsense mutation that resulted in a truncated protein with no PZQ binding site. Our results demonstrate that Sm.TRPMPZQ underlies variation in PZQ responses in S. mansoni and provides an approach for monitoring emerging PZQ-resistant alleles in schistosome elimination programs.

Striking differences in virulence, transmission and sporocyst growth dynamics between two schistosome populations.

BACKGROUND: Parasite traits associated with transmission success, such as the number of infective stages released from the host, are expected to be optimized by natural selection. However, in the trematode parasite Schistosoma mansoni, a key transmission trait, i.e. the number of cercariae larvae shed from infected Biomphalaria spp. snails, varies significantly within and between different parasite populations and selection experiments demonstrate that this variation has a strong genetic basis. In this study, we compared the transmission strategies of two laboratory schistosome population and their consequences for their snail host. METHODS: We infected inbred Biomphalaria glabrata snails using two S. mansoni parasite populations (SmBRE and SmLE), both isolated from Brazil and maintained in the laboratory for decades. We compared life history traits of these two parasite populations by quantifying sporocyst growth within infected snails (assayed using qPCR), output of cercaria larvae and impact on snail host physiological response (i.e. hemoglobin rate, laccase-like activity) and survival. RESULTS: We identified striking differences in virulence and transmission between the two studied parasite populations. SmBRE (low shedder (LS) parasite population) sheds very low numbers of cercariae and causes minimal impact on the snail physiological response (i.e. laccase-like activity, hemoglobin rate and snail survival). In contrast, SmLE (high shedder (HS) parasite population) sheds 8-fold more cercariae (mean ± SE cercariae per shedding: 284 ± 19 vs 2352 ± 113), causes high snail mortality and has strong impact on snail physiology. We found that HS sporocysts grow more rapidly inside the snail host, comprising up to 60% of cells within infected snails, compared to LS sporocysts, which comprised up to 31%. Cercarial production is strongly correlated to the number of S. mansoni sporocyst cells present within the snail host tissue, although the proportion of sporocyst cells alone does not explain the low cercarial shedding of SmBRE. CONCLUSIONS: We demonstrated the existence of alternative transmission strategies in the S. mansoni parasite consistent with trade-offs between parasite transmission and host survival: a "boom-bust" strategy characterized by high virulence, high transmission and short duration infections and a "slow and steady" strategy with low virulence, low transmission but long duration of snail host infections.