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Leishmania are intracellular protozoan parasites that cause the vectorborne disease leishmaniasis, which occurs in [approximately equal to] 88 countries (1). Human infection can result in 2 main forms of disease, [...]

After the first wave of SARS-CoV-2 infections in spring 2020, Europe experienced a resurgence of the virus starting in late summer 2020 that was deadlier and more difficult to contain.sup.1. Relaxed intervention measures and summer travel have been implicated as drivers of the second wave.sup.2. Here we build a phylogeographical model to evaluate how newly introduced lineages, as opposed to the rekindling of persistent lineages, contributed to the resurgence of COVID-19 in Europe. We inform this model using genomic, mobility and epidemiological data from 10 European countries and estimate that in many countries more than half of the lineages circulating in late summer resulted from new introductions since 15 June 2020. The success in onward transmission of newly introduced lineages was negatively associated with the local incidence of COVID-19 during this period. The pervasive spread of variants in summer 2020 highlights the threat of viral dissemination when restrictions are lifted, and this needs to be carefully considered in strategies to control the current spread of variants that are more transmissible and/or evade immunity. Our findings indicate that more effective and coordinated measures are required to contain the spread through cross-border travel even as vaccination is reducing disease burden. In many European countries, more than half of the SARS-CoV-2 lineages circulating in late summer 2020 resulted from new introductions, highlighting the threat of viral dissemination when restrictions are lifted., Author(s): Philippe Lemey [sup.1] [sup.2] , Nick Ruktanonchai [sup.3] [sup.4] , Samuel L. Hong [sup.1] , Vittoria Colizza [sup.5] , Chiara Poletto [sup.5] , Frederik Van den Broeck [sup.1] [sup.6] [...]

Introduction: Resistance against anti-Leishmania drugs (DR) has been studied for years, giving important insights into long-term adaptations of these parasites to drugs, through genetic modifications. However, microorganisms can also survive lethal drug exposure by entering into temporary quiescence, a phenomenon called drug tolerance (DT), which is rather unexplored in Leishmania. Methods: We studied a panel of nine Leishmania braziliensis strains highly susceptible to potassium antimonyl tartrate (PAT), exposed promastigotes to lethal PAT pressure, and compared several cellular and molecular parameters distinguishing DT from DR. Results and discussion: We demonstrated in vitro that a variable proportion of cells remained viable, showing all the criteria of DT and not of DR: i) signatures of quiescence, under drug pressure: reduced proliferation and significant decrease of rDNA transcription; ii) reversibility of the phenotype: return to low IC50 after removal of drug pressure; and iii) absence of significant genetic differences between exposed and unexposed lineages of each strain and absence of reported markers of DR. We found different levels of quiescence and DT among the different L. braziliensis strains. We provide here a new in-vitro model of drug-induced quiescence and DT in Leishmania. Research should be extended in vivo, but the current model could be further exploited to support R&D, for instance, to guide the screening of compounds to overcome the quiescence resilience of the parasite, thereby improving the therapy of leishmaniasis. [ABSTRACT FROM AUTHOR]

Aneuploidy is generally considered harmful, but in some microorganisms, it can act as an adaptive mechanism against environmental stress. Here, we use Leishmania—a protozoan parasite with remarkable genome plasticity—to study the early steps of aneuploidy evolution under high drug pressure (using antimony or miltefosine as stressors). By combining single‐cell genomics, lineage tracing with cellular barcodes, and longitudinal genome characterization, we reveal that aneuploidy changes under antimony pressure result from polyclonal selection of pre‐existing karyotypes, complemented by further and rapid de novo alterations in chromosome copy number along evolution. In the case of miltefosine, early parasite adaptation is associated with independent point mutations in a miltefosine transporter gene, while aneuploidy changes only emerge later, upon exposure to increased drug levels. Therefore, polyclonality and genome plasticity are hallmarks of parasite adaptation, but the scenario of aneuploidy dynamics depends on the nature and strength of the environmental stress as well as on the existence of other pre‐adaptive mechanisms. Synopsis: The dynamics of aneuploidy, a hallmark of Leishmania adaptation, depends on the nature and strength of the environmental stress as well as on the existence of other pre‐adaptive mechanisms. Mosaic aneuploidy generates multiple pre‐adapted karyotypes that can be further modulated during exposure to antimony.Karyotypes evolve in a convergent manner, indicating the adaptive role of aneuploidy changes.Adaptation to miltefosine relies on polyclonal selection of point mutations, and aneuploidy changes emerge at higher drug concentrations. [ABSTRACT FROM AUTHOR]

The Americas, particularly Brazil, were greatly impacted by the widespread Zika virus (ZIKV) outbreak in 2015 and 2016. Efforts were made to implement genomic surveillance of ZIKV as part of the public health responses. The accuracy of spatiotemporal reconstructions of the epidemic spread relies on the unbiased sampling of the transmission process. In the early stages of the outbreak, we recruited patients exhibiting clinical symptoms of arbovirus-like infection from Salvador and Campo Formoso, Bahia, in Northeast Brazil. Between May 2015 and June 2016, we identified 21 cases of acute ZIKV infection and subsequently recovered 14 near full-length sequences using the amplicon tiling multiplex approach with nanopore sequencing. We performed a time-calibrated discrete phylogeographic analysis to trace the spread and migration history of the ZIKV. Our phylogenetic analysis supports a consistent relationship between ZIKV migration from Northeast to Southeast Brazil and its subsequent dissemination beyond Brazil. Additionally, our analysis provides insights into the migration of ZIKV from Brazil to Haiti and the role Brazil played in the spread of ZIKV to other countries, such as Singapore, the USA, and the Dominican Republic. The data generated by this study enhances our understanding of ZIKV dynamics and supports the existing knowledge, which can aid in future surveillance efforts against the virus. [ABSTRACT FROM AUTHOR]

Salmonella enterica serovar Typhimurium strain ATCC14028s is commercially available from multiple national type culture collections, and has been widely used since 1960 for quality control of growth media and experiments on fitness ("laboratory evolution"). ATCC14028s has been implicated in multiple cross-contaminations in the laboratory, and has also caused multiple laboratory infections and one known attempt at bioterrorism. According to hierarchical clustering of 3002 core gene sequences, ATCC14028s belongs to HierCC cluster HC20_373 in which most internal branch lengths are only one to three SNPs long. Many natural Typhimurium isolates from humans, domesticated animals and the environment also belong to HC20_373, and their core genomes are almost indistinguishable from those of laboratory strains. These natural isolates have infected humans in Ireland and Taiwan for decades, and are common in the British Isles as well as the Americas. The isolation history of some of the natural isolates confirms the conclusion that they do not represent recent contamination by the laboratory strain, and 10% carry plasmids or bacteriophages which have been acquired in nature by HGT from unrelated bacteria. We propose that ATCC14028s has repeatedly escaped from the laboratory environment into nature via laboratory accidents or infections, but the escaped micro-lineages have only a limited life span. As a result, there is a genetic gap separating HC20_373 from its closest natural relatives due to a divergence between them in the late 19th century followed by repeated extinction events of escaped HC20_373. Author summary: Clades of closely related bacteria exist in nature. Individual isolates from such clades are often distinguishable by genomic sequencing because genomic sequence differences can be acquired over a few years due to neutral drift and natural selection. The evolution of laboratory strains is often largely frozen, physically due to storage conditions and genetically due to long periods of storage. Thus, laboratory strains can normally be readily distinguished from natural isolates because they show much less diversity. However, laboratory strain ATCC14028s shows modest levels of sequence diversity because it has been shipped around the world to multiple laboratories and is routinely used for analyses of laboratory evolution. Closely related natural isolates also exist, but their genetic diversity is not dramatically greater at the core genome level. Indeed, many scientists doubt that such isolates are natural, and interpret them as undetected contamination by the laboratory strain. We present data indicating that ATCC14028s has repeatedly escaped from the laboratory through inadvertent contamination of the environment, infection of technical staff and deliberate bioterrorism. The escapees survive in nature long enough that some acquire mobile genomic elements by horizontal gene transfer, but eventually they go extinct. As a result, even extensive global databases of natural isolates lack closely related isolates whose ancestors diverged from ATCC14028s within the last 100 years. [ABSTRACT FROM AUTHOR]

Whole genome sequencing (WGS) is increasingly used for molecular diagnosis and epidemiology of infectious diseases. Current Leishmania genomic studies rely on DNA extracted from cultured parasites, which might introduce sampling and biological biases into the subsequent analyses. Up to now, direct analysis of Leishmania genome in clinical samples is hampered by high levels of human DNA and large variation in parasite load in clinical samples. Here, we present a method, based on target enrichment of Leishmania donovani DNA with Agilent SureSelect technology, that allows the analysis of Leishmania genomes directly in clinical samples. We validated our protocol with a set of artificially mixed samples, followed by the analysis of 63 clinical samples (bone marrow or spleen aspirates) from visceral leishmaniasis patients in Nepal. We were able to identify genotypes using a set of diagnostic SNPs in almost all of these samples (97%) and access comprehensive genome-wide information in most (83%). This allowed us to perform phylogenomic analysis, assess chromosome copy number and identify large copy number variants (CNVs). Pairwise comparisons between the parasite genomes in clinical samples and derived in vitro cultured promastigotes showed a lower aneuploidy in amastigotes as well as genomic differences, suggesting polyclonal infections in patients. Altogether our results underline the need for sequencing parasite genomes directly in the host samples Author summary: Visceral leishmaniasis (VL) is caused by parasitic protozoa of the Leishmania donovani complex and is lethal in the absence of treatment. Whole Genome Sequencing (WGS) of L. donovani clinical isolates revealed hitherto cryptic population structure in the Indian Sub-Continent and provided insights into the epidemiology and potential mechanisms of drug resistance. However, several biases are likely introduced during the culture step. We report here the development of a method that allows determination of parasite genomes directly in clinical samples, and validate it on bone marrow and splenic aspirates of VL patients in Nepal. Our study sheds a new light on the biology of Leishmania in the human host: we found that intracellular parasites of the patients had very low levels of aneuploidy, in sharp contrast to the situation in cultivated isolates. Moreover, the observed differences in genomes between intracellular amastigotes of the patient and the derived cultured parasites suggests polyclonality of infections, with different clones dominating in clinical samples and in culture, likely due to fitness differences. We believe this method is most suitable for clinical studies and for molecular tracking in the context of elimination programs. [ABSTRACT FROM AUTHOR]

Abstract: We recently published the first genomic diversity study of Trypanosoma congolense, a major aetiological agent of Animal African Trypanosomiasis. We demonstrated striking levels of SNP and indel diversity in the Eastern province of Zambia as a consequence of hybridization between divergent trypanosome lineages. We concluded that these and earlier findings in T. congolense challenge the predominant clonal evolution (PCE) model. In a recent comment, Tibayrenc and Ayala claim that there are many features in T. congolense supporting their theory of clonality. While we can follow the reasoning of the authors, we also identify major limitations in their theory and interpretations that resulted in incorrect conclusions. First, we argue that each T. congolense subgroup should be analysed independently as they may represent different (sub)species rather than “near‐clades”. Second, the authors neglect major findings of two robust population genetic studies on Savannah T. congolense that provide clear evidence of frequent recombination. Third, we reveal additional events of introgressive hybridization in T. congolense by analysing the maxicircle coding region using next‐generation sequencing analyses. At last, we pinpoint two important misinterpretations by the authors and show that there are no spatially and temporally widespread clones in T. congolense. We stand by our earlier conclusions that the clonal framework is unlikely to accurately model the population structure of T. congolense. Other theoretical frameworks such as Maynard Smith's epidemic model may better represent the complex ancestry seen in T. congolense, where clones delimited in space and time arise against a background of recombination. [ABSTRACT FROM AUTHOR]

Hybrid populations and introgressive hybridization remain poorly documented in pathogenic micro-organisms, as such that genetic exchange has been argued to play a minor role in their evolution. Recent work demonstrated the existence of hybrid microsatellite profiles in Trypanosoma congolense, a parasitic protozoan with detrimental effects on livestock productivity in sub-Saharan Africa. Here, we present the first population genomic study of T. congolense, revealing a remarkable number of single nucleotide polymorphisms ( SNPs), small insertions/deletions (indels) and gene deletions among 56 parasite genomes from ten African countries. One group of parasites from Zambia was particularly diverse, displaying a substantial number of heterozygous SNP and indel sites compared to T. congolense parasites from the nine other sub-Saharan countries. Genomewide 5-kb phylogenetic analyses based on phased SNP data revealed that these parasites were the product of hybridization between phylogenetically distinct T. congolense lineages. Other parasites within the same region in Zambia presented a mosaic of haplotypic ancestry and genetic variability, indicating that hybrid parasites persisted and recombined beyond the initial hybridization event. Our observations challenge traditional views of trypanosome population biology and encourage future research on the role of hybridization in spreading genes for drug resistance, pathogenicity and virulence. [ABSTRACT FROM AUTHOR]

Background: Anthropogenic environmental changes may lead to ecosystem destabilization and the unintentional colonization of new habitats by parasite populations. A remarkable example is the outbreak of intestinal schistosomiasis in Northwest Senegal following the construction of two dams in the ‘80s. While many studies have investigated the epidemiological, immunological and geographical patterns of Schistosoma mansoni infections in this region, little is known about its colonization history. Methodology/Principal Findings: Parasites were collected at several time points after the disease outbreak and genotyped using a 420 bp fragment of the mitochondrial cytochrome c oxidase subunit 1 gene (cox1) and nine nuclear DNA microsatellite markers. Phylogeographic and population genetic analyses revealed the presence of (i) many genetically different haplotypes at the non-recombining mitochondrial marker and (ii) one homogenous S. mansoni genetic group at the recombining microsatellite markers. These results suggest that the S. mansoni population in Northwest Senegal was triggered by intraspecific hybridization (i.e. admixture) between parasites that were introduced from different regions. This would comply with the extensive immigration of infected seasonal agricultural workers from neighboring regions in Senegal, Mauritania and Mali. The spatial and temporal stability of the established S. mansoni population suggests a swift local adaptation of the parasite to the local intermediate snail host Biomphalaria pfeifferi at the onset of the epidemic. Conclusions/Significance: Our results show that S. mansoni parasites are very successful in colonizing new areas without significant loss of genetic diversity. Maintaining high levels of diversity guarantees the adaptive potential of these parasites to cope with selective pressures such as drug treatment, which might complicate efforts to control the disease. [ABSTRACT FROM AUTHOR]

Drug-resistant pathogens emerge faster than new drugs come out of drug discovery pipelines. Current and future drug options should therefore be better protected, requiring a clear understanding of the factors that contribute to the natural history of drug resistance. Although many of these factors are relatively well understood for most bacteria, this proves to be more complex for vectorborne parasites. In this review, we discuss considering three key models ( Plasmodium, Leishmania and Schistosoma) how drug resistance can emerge, spread and persist. We demonstrate a multiplicity of scenarios, clearly resulting from the biological diversity of the different organisms, but also from the different modes of action of the drugs used, the specific within- and between-host ecology of the parasites, and environmental factors that may have direct or indirect effects. We conclude that integrated control of drug-resistant vectorborne parasites is not dependent upon chemotherapy only, but also requires a better insight into the ecology of these parasites and how their transmission can be impaired. [ABSTRACT FROM AUTHOR]

Background: Schistosoma mansoni and S. haematobium are co-endemic in many areas in Africa. Yet, little is known about the micro-geographical distribution of these two infections or associated disease within such foci. Such knowledge could give important insights into the drivers of infection and disease and as such better tailor schistosomiasis control and elimination efforts. Methodology: In a co-endemic farming community in northern Senegal (346 children (0–19 y) and 253 adults (20–85 y); n = 599 in total), we studied the spatial distribution of S. mansoni and S. haematobium single and mixed infections (by microscopy), S. mansoni-specific hepatic fibrosis, S. haematobium-specific urinary tract morbidity (by ultrasound) and water contact behavior (by questionnaire). The Kulldorff's scan statistic was used to detect spatial clusters of infection and morbidity, adjusted for the spatial distribution of gender and age. Principal Findings: Schistosoma mansoni and S. haematobium infection densities clustered in different sections of the community (p = 0.002 and p = 0.023, respectively), possibly related to heterogeneities in the use of different water contact sites. While the distribution of urinary tract morbidity was homogeneous, a strong geospatial cluster was found for severe hepatic fibrosis (p = 0.001). Particularly those people living adjacent to the most frequently used water contact site were more at risk for more advanced morbidity (RR = 6.3; p = 0.043). Conclusions/Significance: Schistosoma infection and associated disease showed important micro-geographical heterogeneities with divergent patterns for S. mansoni and S. haematobium in this Senegalese community. Further in depth investigations are needed to confirm and explain our observations. The present study indicates that local geospatial patterns should be taken into account in both research and control of schistosomiasis. The observed extreme focality of schistosomiasis even at community level, suggests that current strategies may not suffice to move from morbidity control to elimination of schistosomiasis, and calls for less uniform measures at a finer scale. Author Summary: In the developing world, over 230 million people are infected with parasitic Schistosoma worms. Schistosoma mansoni and S. haematobium are the most abundant species in Africa, affecting the liver and urinary tract, respectively. Both parasites are spread through infested freshwater. Although it is known that the disease occurs focally within countries or regions, little is known on its geographic spread on a smaller scale. Here, we examined 599 people from a community in northern Senegal for S. mansoni and S. haematobium infections and related abnormalities of the liver and urinary tract. We recorded where they lived and where they had water contact and visualized this information in geographical maps. The study showed that each Schistosoma species clustered in a different section of the community, and that liver abnormalities were more severe near the mostly used water contact site. So far, this is the first study to investigate the geographical spread of both species in a single community, and the first to map schistosomal disease on such a small scale. Further studies are needed to confirm and explain these results. They could contribute to a better understanding of schistosomiasis and have important consequences for the control and elimination of this disease. [ABSTRACT FROM AUTHOR]

Background: Schistosoma mansoni and S. haematobium are co-endemic in many areas in Africa. Yet, little is known about the micro-geographical distribution of these two infections or associated disease within such foci. Such knowledge could give important insights into the drivers of infection and disease and as such better tailor schistosomiasis control and elimination efforts. Methodology: In a co-endemic farming community in northern Senegal (346 children (0–19 y) and 253 adults (20–85 y); n = 599 in total), we studied the spatial distribution of S. mansoni and S. haematobium single and mixed infections (by microscopy), S. mansoni-specific hepatic fibrosis, S. haematobium-specific urinary tract morbidity (by ultrasound) and water contact behavior (by questionnaire). The Kulldorff's scan statistic was used to detect spatial clusters of infection and morbidity, adjusted for the spatial distribution of gender and age. Principal Findings: Schistosoma mansoni and S. haematobium infection densities clustered in different sections of the community (p = 0.002 and p = 0.023, respectively), possibly related to heterogeneities in the use of different water contact sites. While the distribution of urinary tract morbidity was homogeneous, a strong geospatial cluster was found for severe hepatic fibrosis (p = 0.001). Particularly those people living adjacent to the most frequently used water contact site were more at risk for more advanced morbidity (RR = 6.3; p = 0.043). Conclusions/Significance: Schistosoma infection and associated disease showed important micro-geographical heterogeneities with divergent patterns for S. mansoni and S. haematobium in this Senegalese community. Further in depth investigations are needed to confirm and explain our observations. The present study indicates that local geospatial patterns should be taken into account in both research and control of schistosomiasis. The observed extreme focality of schistosomiasis even at community level, suggests that current strategies may not suffice to move from morbidity control to elimination of schistosomiasis, and calls for less uniform measures at a finer scale. [ABSTRACT FROM AUTHOR]

In total, 245 Cryptosporidium parvum specimens obtained from calves in 205 Irish herds between 2003 and 2005 were subtyped by sequencing the glycoprotein gene gp60 and performing multilocus analysis of seven markers. The transmission dynamics of C. parvum and the influence of temporal, spatial, parasitic, and host-related factors on the parasite (sub)populations were studied. The relationship of those factors to the risk of cryptosporidiosis was also investigated using results from 1,368 fecal specimens submitted to the veterinary laboratories for routine diagnosis during 2005. The prevalence was greatest in the northwest and midwest of the country and on farms that bought in calves. The panmixia (random mating) detected in the C. parvum population may relate to its high prevalence, the cattle density, and the frequent movement of cattle. However, local variations in these factors were reflected in the C. parvumsubpopulations. This study demonstrated the importance of biosecurity in the control of bovine cryptosporidiosis (e.g., isolation and testing of calves before introduction into a herd). Furthermore, the zoonotic risk of C. parvum was confirmed, as most specimens possessed GP60 and MS1 subtypes previously described in humans. [ABSTRACT FROM AUTHOR]

Quantifying the contribution of the various processes that influence population genetic structure is important, but difficult. One of the reasons is that no single measure appropriately quantifies all aspects of genetic structure. An increasing number of studies is analysing population structure using the statistic D, which measures genetic differentiation, next to GST, which quantifies the standardized variance in allele frequencies among populations. Few studies have evaluated which statistic is most appropriate in particular situations. In this study, we evaluated which index is more suitable in quantifying postglacial divergence between three-spined stickleback ( Gasterosteus aculeatus) populations from Western Europe. Population structure on this short timescale (10 000 generations) is probably shaped by colonization history, followed by migration and drift. Using microsatellite markers and anticipating that D and GST might have different capacities to reveal these processes, we evaluated population structure at two levels: (i) between lowland and upland populations, aiming to infer historical processes; and (ii) among upland populations, aiming to quantify contemporary processes. In the first case, only D revealed clear clusters of populations, putatively indicative of population ancestry. In the second case, only GST was indicative for the balance between migration and drift. Simulations of colonization and subsequent divergence in a hierarchical stepping stone model confirmed this discrepancy, which becomes particularly strong for markers with moderate to high mutation rates. We conclude that on short timescales, and across strong clines in population size and connectivity, D is useful to infer colonization history, whereas GST is sensitive to more recent demographic events. [ABSTRACT FROM AUTHOR]

The tropical freshwater snail Bulinus truncatus serves as an important intermediate host of several human and cattle Schistosoma species in many African regions. Despite some ecological and malacological studies, there is no information on the genetic diversity of B. truncatus in Egypt. Here, we sampled 70–100 snails in ten localities in Upper Egypt and the Nile Delta. Per locality, we sequenced 10 snails at a partial fragment of the cytochrome c oxidase subunit 1 gene ( cox 1) and we genotyped 25–30 snails at six microsatellite markers. A total of nine mitochondrial haplotypes were detected, of which five were unique to the Nile Delta and three were unique to Upper Egypt, indicating that snail populations may have evolved independently in both regions. Bayesian clustering and hierarchical F -statistics using microsatellite markers further revealed strong population genetic structure at the level of locality. Observed heterozygosity was much lower compared to what is expected under random mating, which could be explained by high selfing rates, population size reductions and to a lesser extent by the Wahlund effect. Despite these observations, we found signatures of gene flow and cross-fertilization, even between snails from the Nile Delta and Upper Egypt, indicating that B. truncatus can travel across large distances in Egypt. These observations could have serious consequences for disease epidemiology, as it means that infected snails from one region could rapidly and unexpectedly spark a new epidemic in another distant region. This could be one of the factors explaining the rebound of human Schistosoma infections in the Nile Delta, despite decades of sustained schistosomiasis control. [ABSTRACT FROM AUTHOR]

Abstract: In order to understand the epidemiological dynamics of antimonial (SbV) resistance in zoonotic tegumentary leishmaniasis and its link with treatment outcome, we analyzed the population structure of 24 Peruvian Leishmania braziliensis clinical isolates with known in vitro antimony susceptibility and clinical phenotype by multilocus microsatellite typing (14 microsatellite loci). The genetic variability in the Peruvian isolates was high and the multilocus genotypes were strongly differentiated from each other. No correlation was found between the genotypes and in vitro drug susceptibility or clinical treatment outcome. The finding of a polyphyletic pattern among the SbV-resistant L. braziliensis might be explained by (i) independent events of drug resistance emergence, (ii) sexual recombination and/or (iii) other phenomena mimicking recombination signals. Interestingly, the polyphyletic pattern observed here is very similar to the one we observed in the anthroponotic Leishmania donovani (), hereby questioning the role of transmission and/or chemotherapeutic drug pressure in the observed population structure. [Copyright &y& Elsevier]