Tropical Pathogen Research Articles

RT-qPCR Analysis of Inflammatory & Apoptotic Factors-Related Gene Expression in ZIKV-Infected IFNAR1-/- Mice.

ZIKV was a mosquito-borne neglected tropical pathogen until it spread into the Pacific and South America, followed by large human outbreaks related to congenital abnormalities in neonates and neurological disorders in adults. The following study used the C57BL/6 IFNAR1 receptor knockout (IFN AR1-/-) mouse model to understand the role of selected cytokines and apoptotic factors in the pathogenicity of ZIKV strain PRVABC59. Mice infected with 102 particles of Zika viruses died until 9days post infection. The brain, spleen, and lung were collected from intramuscularly infected mice on day 6 post infection (pi) to quantify the mRNA expression of targeted cytokines and apoptosis-mediated factors by RT-qPCR. Upregulation of IL-6, IL-17α, IFN-α, and IFN-β were found in the brain and lung of infected mice. IFN-γ was also significantly upregulated in the infected brain and spleen. The collective findings from our study indicate that a strong immune response was developed against ZIKV PRVABC59 in the infected mice brain.

Phylogenetic and phenotypic characterization of Burkholderia pseudomallei isolates from Ghana reveals a novel sequence type and common phenotypes.

Melioidosis is a potentially severe disease caused by the gram-negative soil-dwelling bacterium called Burkholderia pseudomallei. The true breadth of the distribution of this tropical pathogen is starting to emerge with environmental and clinical isolates frequently characterized in new countries and regions. Even so, isolates, clinical cases, and genetic data from the continent of Africa remain scant. We previously confirmed the presence of B. pseudomallei in the environment of Ghana, unmasking a new area of endemicity for this pathogen. Here, we describe the genetic characteristics of isolates obtained from that environmental survey. Twenty-one isolates were subjected to whole genome sequencing and found to represent three discrete sequence types (ST), one of which was novel, and designated ST2058. Phylogenetic analysis places this novel isolate within a B. pseudomallei clade that includes genomes derived from the Americas, although it is closely related to a sub-clade that includes isolates from Africa. Importantly, phenotypic characterization demonstrates common features including API 20NE profiles and B. pseudomallei CPS to support existing diagnostics, and susceptibility to standard of care antibiotics often used in the clinical management of melioidosis. These findings add to our knowledge about the presence and distribution of B. pseudomallei in Africa and represent the first published genomes out of Ghana.

Chandipura Virus Forms Cytoplasmic Inclusion Bodies through Phase Separation and Proviral Association of Cellular Protein Kinase R and Stress Granule Protein TIA-1.

Negative-strand RNA viruses form cytoplasmic inclusion bodies (IBs) representing virus replication foci through phase separation or biomolecular condensation of viral and cellular proteins, as a hallmark of their infection. Alternatively, mammalian cells form stalled mRNA containing antiviral stress granules (SGs), as a consequence of phosphorylation of eukaryotic initiation factor 2α (eIF2α) through condensation of several RNA-binding proteins including TIA-1. Whether and how Chandipura virus (CHPV), an emerging human pathogen causing influenza-like illness, coma and death, forms IBs and evades antiviral SGs remain unknown. By confocal imaging on CHPV-infected Vero-E6 cells, we found that CHPV infection does not induce formation of distinct canonical SGs. Instead, CHPV proteins condense and co-localize together with SG proteins to form heterogeneous IBs, which ensued independent of the activation of eIF2α and eIF2α kinase, protein kinase R (PKR). Interestingly, siRNA-mediated depletion of PKR or TIA-1 significantly decreased viral transcription and virion production. Moreover, CHPV infection also caused condensation and recruitment of PKR to IBs. Compared to SGs, IBs exhibited significant rapidity in disassembly dynamics. Altogether, our study demonstrating that CHPV replication co-optimizes with SG proteins and revealing an unprecedented proviral role of TIA-1/PKR may have implications in understanding the mechanisms regulating CHPV-IB formation and designing antiviral therapeutics. Importance: CHPV is an emerging tropical pathogen reported to cause acute influenza-like illness and encephalitis in children with a very high mortality rate of ~70%. Lack of vaccines and an effective therapy against CHPV makes it a potent pathogen for causing an epidemic in tropical parts of globe. Given these forewarnings, it is of paramount importance that CHPV biology must be understood comprehensively. Targeting of host factors offers several advantages over targeting the viral components due to the generally higher mutation rate in the viral genome. In this study, we aimed at understanding the role of SGs forming cellular RNA-binding proteins in CHPV replication. Our study helps understand participation of cellular factors in CHPV replication and could help develop effective therapeutics against the virus.

The buoyancy of cryptococcal cells and its implications for transport and persistence of Cryptococcus in aqueous environments.

Cryptococcus is a genus of saprophytic fungi with global distribution. Two species complexes, C. neoformans and C. gattii, pose health risks to humans and animals. Cryptococcal infections result from inhalation of aerosolized spores and/or desiccated yeasts from terrestrial reservoirs such as soil, trees, and avian guano. More recently, C. gattii has been implicated in infections in marine mammals, suggesting that inhalation of liquid droplets or aerosols from the air-water interface is also an important, yet understudied, mode of respiratory exposure. Water transport has also been suggested to play a role in the spread of C. gattii from tropical to temperate environments. However, the dynamics of fungal survival, persistence, and transport via water have not been fully studied. The size of the cryptococcal capsule was previously shown to reduce cell density and increase buoyancy. Here, we demonstrate that cell buoyancy is also impacted by the salinity of the media in which cells are suspended, with formation of a halocline interface significantly slowing the rate of settling of cryptococcal cells through water, resulting in persistence of C. neoformans within 1 cm of the air-water interface for over 60 min and C. gattii for 4-6 h. Our data also showed that during culture in yeast peptone dextrose media (YPD), polysaccharide accumulating in the supernatant formed a raft that augmented buoyancy and further slowed settling of cryptococcal cells. These findings illustrate new mechanisms by which cryptococcal cells may persist in aquatic environments, with important implications for aqueous transport and pathogen exposure.

Host proteins Alpha-2-Macroglobulin and LRP1 associate with Chandipura virus

Chandipura Virus is an emerging tropical pathogen with a high mortality rate among children. No mode of treatment or antivirals exists against CHPV infection, due to little information regarding its host interaction. Studying viral pathogen interaction with its host can not only provide valuable information regarding its propagation strategy, but also on which host proteins interact with the virus. Identifying these proteins and understanding their role in the infection process can provide more stable anti-viral targets. In this study, we focused on identifying host factors that interact with CHPV and may play a critical role in CHPV infection. We are the first to report the successful identification of Alpha-2-Macroglobulin (A2M), a secretory protein of the host that interacts with CHPV. We also established that LRP1 (Low-density lipoprotein receptor-related protein 1) and GRP78 (Glucose regulated protein 78), receptors of A2M, also interact with CHPV. Furthermore, we could also demonstrate that knocking out A2M has a severe effect on viral infection. We conclusively show the interaction of these host proteins with CHPV. Our findings also indicate that these host proteins could play a role in viral entry into the host cell.

Design of a multi-epitope subunit vaccine candidate for chikungunya virus using computational methodology.

Chikungunya virus (CHIKV) is a neglected tropical pathogen that causes fever and long-lasting severe arthralgia. Despite its high morbidity, there is still no licensed specific therapeutic option for it. This study proposes a multi-epitope subunit vaccine candidate for CHIKV, designed using computational methods. It was based on the E2 spike glycoprotein in CHIKV, from which T- and B-cell epitopes were predicted and then refined. The pan HLA DR-binding epitope (PADRE) was added to this refined construct, then simulated compared with the native protein, where it was predicted to elicit more than twice the number of antibody titers. Thus, this construct is potentially effective against CHIKV, which further experimentation using live models would be able to verify. This study also demonstrates the feasibility of using rational tools in the future to further optimize vaccine design.

A novel lytic phage potentially effective for phage therapy against Burkholderia pseudomallei in the tropics

BackgroundBurkholderia pseudomallei is a tropical pathogen that causes melioidosis. Its intrinsic drug-resistance is a leading cause of treatment failure, and the few available antibiotics require prolonged use to be effective. This study aimed to assess the clinical potential of B. pseudomallei phages isolated from Hainan, China.MethodsBurkholderia pseudomallei strain (HNBP001) was used as the isolation host, and phages were recovered from domestic environmental sources, which were submitted to the host range determination, lytic property assays, and stability tests. The best candidate was examined via the transmission electron microscope for classification. With its genome sequenced and analyzed, its protective efficacy against B. pseudomallei infection in A549 cells and Caenorhabditis elegans was evaluated, in which cell viability and survival rates were compared using the one-way ANOVA method and the log-rank test.ResultsA phage able to lyse 24/25 clinical isolates was recovered. It was classified in the Podoviridae family and was found to be amenable to propagation. Under the optimal multiplicity of infection (MOI) of 0.1, an eclipse period of around 20 min and a high titer (1012 PFU/ml) produced within 1 h were demonstrated. This phage was found stabile at a wide range of temperatures (24, 37, 40, 50, and 60 °C) and pH values (3–12). After being designated as vB_BpP_HN01, it was fully sequenced, and the 71,398 bp linear genome, containing 93 open reading frames and a tRNA-Asn, displayed a low sequence similarity with known viruses. Additionally, protective effects of applications of vB_BpP_HN01 (MOI = 0.1 and MOI = 1) alone or in combination with antibiotics were found to improve viability of infected cells (70.6 ± 6.8%, 85.8 ± 5.7%, 91.9 ± 1.8%, and 96.8 ± 1.8%, respectively). A significantly reduced mortality (10%) and a decreased pathogen load were demonstrated in infected C. elegans following the addition of this phage.ConclusionsAs the first B. pseudomallei phage was isolated in Hainan, China, phage vB_BpP_HN01 was characterized by promising lytic property, stability, and efficiency of bacterial elimination during the in vitro/vivo experiments. Therefore, we can conclude that it is a potential alternative agent for combating melioidosis.Graphical

Differential Excretory/Secretory Proteome of the Adult Female and Male Stages of the Human Blood Fluke, Schistosoma mansoni

Intricate molecular communication between schistosome flatworms and their mammalian host, as well as between paired male and female schistosomes has shaped the secreted proteome of these flatworms. Whereas the schistosome egg is responsible for the disease manifestations of chronic schistosomiasis, the long lived, adult female and male stages also release different mediators including glycans, lipids, proteins and small molecules, known as excretory/secretory products (ESPs), that facilitate their survival. Given their importance, deeper analysis focused on analyzing the ESPs from adult schistosomes would likely be informative, beyond current understanding of the complement of ESP proteins. Here, taking advantage of highly accurate and sensitive mass spectrometers, the excretory/secretory proteome from culturedSchistosoma mansonimale or female adult worms was identified, quantified, compared and contrasted using a label-free proteomic approach. Approximately 1,000 proteins were identified, from which almost 800 could be quantified. Considering the proteins uniquely identified and proteins with a significantly regulated expression pattern in male or female flukes, a total of 370 and 140 proteins were uniquely or more abundantly secreted by males and females, respectively. Using functional analysis networks showing the gene ontology terms and KEGG pathways with the highest significance, we observed that male schistosomes secrete proteins related to carbohydrate metabolism and cytoskeletal organization more abundantly than females, while female worms secreted more hydrolases and proteins involved in cellular homeostasis than males. This analysis doubles the number of reported excreted/secreted proteins fromS. mansoni, contributing to deeper understanding of the host-parasite interaction and parasitism. Furthermore, these findings expand potential vaccine and diagnostic candidates for this neglected tropical disease pathogen, and thereby also provide leads for novel intervention to control this disease and its transmission.

Probing the Peculiarity of EhRabX10, a pseudoRab GTPase, from the Enteric Parasite Entamoeba histolytica through In Silico Modeling and Docking Studies.

Entamoeba histolytica (Eh) is a pathogenic eukaryote that often resides silently in humans under asymptomatic stages. Upon indeterminate stimulus, it develops into fulminant amoebiasis that causes severe hepatic abscesses with 50% mortality. This neglected tropical pathogen relies massively on membrane modulation to flourish and cause disease; these modulations range from the phagocytic mode for food acquisition to a complex trogocytosis mechanism for tissue invasion. Rab GTPases form the largest branch of the Ras-like small GTPases, with a diverse set of roles across the eukaryotic kingdom. Rab GTPases are vital for the orchestration of membrane transport and the secretory pathway responsible for transporting the pathogenic effectors, such as cysteine proteases (EhCPs) which help in tissue invasion. Rab GTPases thus play a crucial role in executing the cytolytic effect of E. histolytica. First, they interact with Gal/Nac lectins required for adhering to the host cells, and then, they assist in the secretion of EhCPs. Additionally, amoebic Rab GTPases are vital for encystation because substantial vesicular trafficking is required to create dormant amoebic cysts. These cysts are the infective agent and help to spread the disease. The absence of a “bonafide” vesicular transport machinery in Eh and the existence of a diverse repertoire of amoebic Rab GTPases (EhRab) hint at their contribution in supporting this atypical machinery. Here, we provide insights into a pseudoRab GTPase, EhRabX10, by performing physicochemical analysis, predictive 3D structure modeling, protein-protein interaction studies, and in silico molecular docking. Our group is the first one to classify EhRabX10 as a pseudoRab GTPase with four nonconserved G-motifs. It possesses the basic fold of the P-loop containing nucleotide hydrolases. Through this in silico study, we provide an introduction to the characterization of the atypical EhRabX10 and set the stage for future explorations into the mechanisms of nucleotide recognition, binding, and hydrolysis employed by the pseudoEhRab GTPase family.

Exposure to dexamethasone modifies transcriptomic responses of free-living stages of Strongyloides stercoralis.

Hyperinfection and disseminated infection by the parasitic nematode Strongyloides stercoralis can be induced by iatrogenic administration of steroids and immunosuppression and lead to an elevated risk of mortality. Responses of free-living stages of S. stercoralis to the therapeutic corticosteroid dexamethasone (DXM) were investigated using RNA-seq transcriptomes of DXM-treated female and male worms. A total of 17,950 genes representing the transcriptome of these free-living adult stages were obtained, among which 199 and 263 were differentially expressed between DXM-treated females and DXM-treated males, respectively, compared with controls. According to Gene Ontology analysis, differentially expressed genes from DXM-treated females participate in developmental process, multicellular organismal process, cell differentiation, carbohydrate metabolic process and embryonic morphogenesis. Others are involved in signaling and signal transduction, including cAMP, cGMP-dependent protein kinase pathway, endocrine system, and thyroid hormone pathway, as based on Kyoto Encyclopedia of Genes and Genomes analysis. The novel findings warrant deeper investigation of the influence of DXM on growth and other pathways in this neglected tropical disease pathogen, particularly in a setting of autoimmune and/or allergic disease, which may require the clinical use of steroid-like hormones during latent or covert strongyloidiasis.

Efficient flavinylation of glycosomal fumarate reductase by its own ApbE domain in Trypanosoma brucei.

A subset of flavoproteins has a covalently attached flavin prosthetic group enzymatically attached via phosphoester bonding. In prokaryotes, this is catalysed by alternative pyrimidine biosynthesis E (ApbE) flavin transferases. ApbE-like domains are present in few eukaryotic taxa, for example the N-terminal domain of fumarate reductase (FRD) of Trypanosoma, a parasitic protist known as a tropical pathogen causing African sleeping sickness. We use the versatile reverse genetic tools available for Trypanosoma to investigate the flavinylation of glycosomal FRD (FRDg) invivo in the physiological and organellar context. Using direct in-gel fluorescence detection of covalently attached flavin as proxy for activity, we show that the ApbE-like domain of FRDg has flavin transferase activity invivo. The ApbE domain is preceded by a consensus flavinylation target motif at the extreme N terminus of FRDg, and serine 9 in this motif is essential as flavin acceptor. The preferred mode of flavinylation in the glycosome was addressed by stoichiometric expression and comparison of native and catalytically inactive ApbE domains. In addition to the trans-flavinylation activity, the ApbE domain catalyses the intramolecular cis-flavinylation with at least fivefold higher efficiency. We discuss how the higher efficiency due to unusual fusion of the ApbE domain to its substrate protein FRD may provide a selective advantage by faster FRD biogenesis during rapid metabolic adaptation of trypanosomes. The first 37 amino acids of FRDg, including the consensus motif, are sufficient as flavinylation target upon fusion to other proteins. We propose FRDg(1-37) as 4-kDa heat-stable, detergent-resistant fluorescent protein tag and suggest its use as a new tool to study glycosomal protein import.

The Fusarium solani species complex infecting cannabis (Cannabis sativa L., marijuana) plants and a first report of Fusarium (Cylindrocarpon) lichenicola causing root and crown rot

Greenhouse-grown cannabis (Cannabis sativa L., marijuana) plants with yellowing, crown rot and root-browning symptoms were sampled from six production facilities during 2019–2020. Among 34 fungal isolates recovered, 28 were identified as Fusarium solani and six isolates were provisionally identified as Cylindrocarpon sp. based on morphology. These latter isolates produced slow-growing colonies with grey-white aerial mycelium and a chestnut-brown colour below. Cylindrical 1–3 septate conidia without a distinctive foot cell were produced. Microconidia were absent and chlamydospores were produced in culture. Phylogenetic analysis of three isolates based on the elongation factor (TEF-1 α) and ITS1-5.8S-ITS2 regions identified them as Fusarium lichenicola (formerly Cylindrocarpon lichenicola), a member of F. solani species complex (FSSC) subclade 16. Pathogenicity tests using a mycelial and spore suspension were performed on cannabis cuttings and rooted plants. Isolates of F. solani originating from diseased crowns caused symptoms in 10–14 days, while those of F. lichenicola caused yellowing and wilting after 3 weeks, suggesting that F. lichenicola is less virulent. Inoculum of F. lichenicola was detected in coco coir samples used for plant propagation. Previous reports of F. lichenicola are from tropical climates, where the fungus has been associated with dermal and ocular infections of human tissues, with a few reports of it causing diseases on pomelo fruits, taro corms and tea plants. This study demonstrates the first occurrence worldwide of F. lichenicola on cannabis plants, on which it is considered a weak introduced tropical pathogen, likely to have originated from coco coir imported into Canada.

A Persisting Nontropical Focus of Burkholderia pseudomallei with Limited Genome Evolution over Five Decades.

Burkholderia pseudomallei is the causative agent of the high-mortality disease melioidosis. Although melioidosis is classified as a tropical disease, rare autochthonous cases have been reported from temperate climatic regions, with uncertainty as to whether B. pseudomallei is persistent in the local environment and whether specific genetic mechanisms facilitate the survival of B. pseudomallei outside the tropics. Sporadic cases of melioidosis occurred in a valley region (latitude 31.6°S) in southwest Western Australia, Australia, between 1966 and 1992. We report a new melioidosis cluster in the same region following high rainfall in January 2017. More than 20 animals died, and B. pseudomallei was isolated from four alpacas, a parrot, and three environmental samples taken from the farm where the alpacas resided. Epidemiological data and genomics revealed that two locations on the farm were the probable sources of the alpaca infections. We determined that B. pseudomallei isolates from the 2017 cluster belonged to sequence type 284 (ST-284), as did all isolates recovered from 1966 to 1992. Genomic analysis confirmed that the ST-284 isolates were clonal and contained conserved genomic islands and limited evidence of recombination. We identified protein-coding regions unique to these isolates that might influence the persistence of B. pseudomallei in this temperate region. We demonstrate the environmental persistence of B. pseudomallei in a temperate region for over 50 years, with limited genetic changes suggesting a latent state and with activation, potential aerosolization, and local dispersal following unusually high rainfall.IMPORTANCE Burkholderia pseudomallei is predominantly a tropical pathogen uncommonly found in the environment of temperate climatic regions. It is unclear if introduction into temperate regions is sporadic and temporary or if B. pseudomallei can persist in such environments. B. pseudomallei was identified in the environment of southwest Western Australia with melioidosis cases between 1966 and 1991. We report a new cluster with 23 animal fatalities in the same region from 2017, with B. pseudomallei again being recovered from the environment. Comparison of the isolates from the first and second clusters using genomics revealed a single sequence type, high clonality, and limited recombination, even though the time of recovery of the isolates spanned 51 years. This is a major contrast to the extensive genomic diversity seen in the tropics. Our data support the suggestion that B. pseudomallei has the ability to persist in nontropical environments, potentially in a latent state, and has the ability to activate following favorable conditions (rainfall) and then infect animals and humans.

Ribavirin inhibits Chandipura virus replication in Vero cells.

Chandipura virus (CHPV) is an emerging tropical pathogen in India. The virus has been reported to be associated with an acute encephalitis syndrome in young children with a case fatality rate of 55% to 75%. Clinical management with symptomatic treatment is the only option available to treat infected patients. No vaccines are available for prophylaxis. In light of the prophylactic limitations, antiviral therapy would play an important role in control of CHPV infection. In the present study, ribavirin (RBV), an antiviral drug widely accepted for human use and having an antiviral effect on many RNA and DNA viruses, was tested against the CHPV. A screening assay that scores for the virus-mediated plaque formation in the cultured Vero cells was used. RBV exhibited 50% inhibitory concentration (IC50 ) at 89.84 ± 1.8 µM. The drug was very effective when the cells were treated either within an hour postinfection or 4 to 6 hours before infection. The plaque morphology was different in RBV treated cells; the plaques were smaller in size as compared with the plaques in the virus infected cells. The study reports the antiviral activity of RBV against CHPV, and hence, suggests the possible utility of RBV in CHPV infected patients to mitigate the disease. A further clinical trial is needed before introducing the drug for human use against CHPV infection.

Transcriptome of the parasitic flatworm Schistosoma mansoni during intra-mammalian development.

Schistosomes are parasitic blood flukes that survive for many years within the mammalian host vasculature. How the parasites establish a chronic infection in the hostile bloodstream environment, whilst evading the host immune response is poorly understood. The parasite develops morphologically and grows as it migrates to its preferred vascular niche, avoiding or repairing damage from the host immune system. In this study, we investigated temporal changes in gene expression during the intra-mammalian development of Schistosoma mansoni. RNA-seq data were analysed from parasites developing in the lung through to egg-laying mature adult worms, providing a comprehensive picture of in vivo intra-mammalian development. Remarkably, genes involved in signalling pathways, developmental control, and adaptation to oxidative stress were up-regulated in the lung stage. The data also suggested a potential role in immune evasion for a previously uncharacterised gene. This study not only provides a large and comprehensive data resource for the research community, but also reveals new directions for further characterising host-parasite interactions that could ultimately lead to new control strategies for this neglected tropical disease pathogen.

Genome sequencing of Rigidoporus microporus provides insights on genes important for wood decay, latex tolerance and interspecific fungal interactions

Fungal plant pathogens remain a serious threat to the sustainable agriculture and forestry, despite the extensive efforts undertaken to control their spread. White root rot disease is threatening rubber tree (Hevea brasiliensis) plantations throughout South and Southeast Asia and Western Africa, causing tree mortality and severe yield losses. Here, we report the complete genome sequence of the basidiomycete fungus Rigidoporus microporus, a causative agent of the disease. Our phylogenetic analysis confirmed the position of R. microporus among the members of Hymenochaetales, an understudied group of basidiomycetes. Our analysis further identified pathogen’s genes with a predicted role in the decay of plant cell wall polymers, in the utilization of latex components and in interspecific interactions between the pathogen and other fungi. We also detected putative horizontal gene transfer events in the genome of R. microporus. The reported first genome sequence of a tropical rubber tree pathogen R. microporus should contribute to the better understanding of how the fungus is able to facilitate wood decay and nutrient cycling as well as tolerate latex and utilize resinous extractives.

Epidemiology of Bladder Cancer.

Based on the latest GLOBOCAN data, bladder cancer accounts for 3% of global cancer diagnoses and is especially prevalent in the developed world. In the United States, bladder cancer is the sixth most incident neoplasm. A total of 90% of bladder cancer diagnoses are made in those 55 years of age and older, and the disease is four times more common in men than women. While the average 5-year survival in the US is 77%, the 5-year survival for those with metastatic disease is a measly 5%. The strongest risk factor for bladder cancer is tobacco smoking, which accounts for 50–65% of all cases. Occupational or environmental toxins likewise greatly contribute to disease burden (accounting for an estimated 20% of all cases), though the precise proportion can be obscured by the fact bladder cancer develops decades after exposure, even if the exposure only lasted several years. Schistosomiasis infection is the common cause of bladder cancer in regions of Africa and the Middle East and is considered the second most onerous tropical pathogen after malaria. With 81% of cases attributable to known risk factors (and only 7% to heritable mutations), bladder cancer is a prime candidate for prevention strategies. Smoking cessation, workplace safety practices, weight loss, exercise and schistosomiasis prevention (via water disinfection and mass drug administration) have all been shown to significantly decrease the risk of bladder cancer, which poses a growing burden around the world.

Functional analysis of vasa/PL10-like genes in the ovary of Schistosoma mansoni

The RNA helicase Vasa plays a pivotal role in the development of the germ line. To decipher the functional roles of vasa/PL10-like genes in the human blood fluke Schistosoma mansoni, we performed RNA interference followed by the analysis of the ovary in the adult female. Double-stranded RNA targeting the schistosome vasa-like gene Smvlg1 reduced the volume of the ovary. Changes in morphology of the ovary were analysed using carmine red-staining of the parasites followed by a novel confocal laser scanning microscopy (CLSM)-based approach to control for natural autofluorescence in female schistosome tissues. The reduction in the ovary volume may have been promoted by the loss of germ cells. By contrast, significant differences were not apparent in the number of eggs produced or hatching rate of eggs laid by the female schistosomes transfected with Smvlg1-specific dsRNA. The findings suggested a role for S. mansoni vasa/PL10-like gene -1 in germ cell development within the schistosome ovary that might impact in the pathogenesis and disease transmission by this neglected tropical disease pathogen.

Dating the Cryptococcus gattii Dispersal to the North American Pacific Northwest.

The emergence of Cryptococcus gattii, previously regarded as a predominantly tropical pathogen, in the temperate climate of the North American Pacific Northwest (PNW) in 1999 prompted several questions. The most prevalent among these was the timing of the introduction of this pathogen to this novel environment. Here, we infer tip-dated timing estimates for the three clonal C.gattii populations observed in the PNW, VGIIa, VGIIb, and VGIIc, based on whole-genome sequencing of 134 C.gattii isolates and using Bayesian evolutionary analysis by sampling trees (BEAST). We estimated the nucleotide substitution rate for each lineage (1.59 × 10-8, 1.59 × 10-8, and 2.70 × 10-8, respectively) to be an order of magnitude higher than common neutral fungal mutation rates (2.0 × 10-9), indicating a microevolutionary rate (e.g., successive clonal generations in a laboratory) in comparison to a species' slower, macroevolutionary rate (e.g., when using fossil records). The clonal nature of the PNW C.gattii emergence over a narrow number of years would therefore possibly explain our higher mutation rates. Our results suggest that the mean time to most recent common ancestor for all three sublineages occurred within the last 60 to 100years. While the cause of C.gattii dispersal to the PNW is still unclear, our research estimates that the arrival is neither ancient nor very recent (i.e., <25 years ago), making a strong case for an anthropogenic introduction. IMPORTANCE The recent emergence of the pathogenic fungus Cryptococcus gattii in the Pacific Northwest (PNW) resulted in numerous investigations into the epidemiological and enzootic impacts, as well as multiple genomic explorations of the three primary molecular subtypes of the fungus that were discovered. These studies lead to the general conclusion that the subtypes identified likely emerged out of Brazil. Here, we conducted genomic dating analyses to determine the ages of the various lineages seen in the PNW and propose hypothetical causes for the dispersal events. Bayesian evolutionary analysis strongly suggests that these independent fungal populations in the PNW are all 60 to 100 years old, providing a timing that is subsequent to the opening of the Panama Canal, which allowed for more direct shipping between Brazil and the western North American coastline, a possible driving event for these fungal translocation events.

A novel Zika virus mouse model reveals strain specific differences in virus pathogenesis and host inflammatory immune responses.

Zika virus (ZIKV) is a mosquito borne flavivirus, which was a neglected tropical pathogen until it emerged and spread across the Pacific Area and the Americas, causing large human outbreaks associated with fetal abnormalities and neurological disease in adults. The factors that contributed to the emergence, spread and change in pathogenesis of ZIKV are not understood. We previously reported that ZIKV evades cellular antiviral responses by targeting STAT2 for degradation in human cells. In this study, we demonstrate that Stat2-/- mice are highly susceptible to ZIKV infection, recapitulate virus spread to the central nervous system (CNS), gonads and other visceral organs, and display neurological symptoms. Further, we exploit this model to compare ZIKV pathogenesis caused by a panel of ZIKV strains of a range of spatiotemporal history of isolation and representing African and Asian lineages. We observed that African ZIKV strains induce short episodes of severe neurological symptoms followed by lethality. In comparison, Asian strains manifest prolonged signs of neuronal malfunctions, occasionally causing death of the Stat2-/- mice. African ZIKV strains induced higher levels of inflammatory cytokines and markers associated with cellular infiltration in the infected brain in mice, which may explain exacerbated pathogenesis in comparison to those of the Asian lineage. Interestingly, viral RNA levels in different organs did not correlate with the pathogenicity of the different strains. Taken together, we have established a new murine model that supports ZIKV infection and demonstrate its utility in highlighting intrinsic differences in the inflammatory response induced by different ZIKV strains leading to severity of disease. This study paves the way for the future interrogation of strain-specific changes in the ZIKV genome and their contribution to viral pathogenesis.