Life Stages Of Parasite Research Articles

Staining-Independent Malaria Parasite Detection and Life Stage Classification in Blood Smear Images

Malaria is a leading cause of morbidity and mortality in tropical and sub-tropical regions. This research proposed a malaria diagnosis system based on the you only look once algorithm for malaria parasite detection and the convolutional neural network algorithm for malaria parasite life stage classification. Two public datasets are utilized: MBB and MP-IDB. The MBB dataset includes human blood smears infected with Plasmodium vivax (P. vivax). While the MP-IDB dataset comprises 4 species of malaria parasites: P. vivax, P. ovale, P. malariae, and P. falciparum. Four distinct stages of life exist in every species, including ring, trophozoite, schizont, and gametocyte. For the MBB dataset, detection and classification accuracies of 0.92 and 0.93, respectively, were achieved. For the MP-IDB dataset, the proposed algorithms yielded the accuracies for detection and classification as follows: 0.84 and 0.94 for P. vivax; 0.82 and 0.93 for P. ovale; 0.79 and 0.93 for P. malariae; and 0.92 and 0.96 for P. falciparum. The detection results showed the models trained by P. vivax alone provide good detection capabilities also for other species of malaria parasites. The classification performance showed the proposed algorithms yielded good malaria parasite life stage classification performance. The future directions include collecting more data and exploring more sophisticated algorithms.

Ternary structure of Plasmodium vivaxN-myristoyltransferase with myristoyl-CoA and inhibitor IMP-0001173.

Plasmodium vivax is a major cause of malaria, which poses an increased health burden on approximately one third of the world's population due to climate change. Primaquine, the preferred treatment for P. vivax malaria, is contraindicated in individuals with glucose-6-phosphate dehydrogenase (G6PD) deficiency, a common genetic cause of hemolytic anemia, that affects ∼2.5% of the world's population and ∼8% of the population in areas of the world where P. vivax malaria is endemic. The Seattle Structural Genomics Center for Infectious Disease (SSGCID) conducted a structure-function analysis of P.vivax N-myristoyltransferase (PvNMT) as part of efforts to develop alternative malaria drugs. PvNMT catalyzes the attachment of myristate to the N-terminal glycine of many proteins, and this critical post-translational modification is required for the survival of P. vivax. The first step is the formation of aPvNMT-myristoyl-CoA binary complex that can bind to peptides. Understanding how inhibitors prevent protein binding will facilitate the development of PvNMT as a viable drug target. NMTs are secreted in all life stages of malarial parasites, making them attractive targets, unlike current antimalarials that are only effective during the plasmodial erythrocytic stages. The 2.3 Å resolution crystal structure of the ternary complex of PvNMT with myristoyl-CoA and a novel inhibitor is reported. One asymmetric unit contains two monomers. The structure reveals notable differences between the PvNMT and human enzymes and similarities to other plasmodial NMTs that can be exploited to develop new antimalarials.

Ontological representation, modeling, and analysis of parasite vaccines

BackgroundPathogenic parasites are responsible for multiple diseases, such as malaria and Chagas disease, in humans and livestock. Traditionally, pathogenic parasites have been largely an evasive topic for vaccine design, with most successful vaccines only emerging recently. To aid vaccine design, the VIOLIN vaccine knowledgebase has collected vaccines from all sources to serve as a comprehensive vaccine knowledgebase. VIOLIN utilizes the Vaccine Ontology (VO) to standardize the modeling of vaccine data. VO did not model complex life cycles as seen in parasites. With the inclusion of successful parasite vaccines, an update in parasite vaccine modeling was needed.ResultsVIOLIN was expanded to include 258 parasite vaccines against 23 protozoan species, and 607 new parasite vaccine-related terms were added to VO since 2022. The updated VO design for parasite vaccines accounts for parasite life stages and for transmission-blocking vaccines. A total of 356 terms from the Ontology of Parasite Lifecycle (OPL) were imported to VO to help represent the effect of different parasite life stages. A new VO class term, ‘transmission-blocking vaccine,’ was added to represent vaccines able to block infectious transmission, and one new VO object property, ‘blocks transmission of pathogen via vaccine,’ was added to link vaccine and pathogen in which the vaccine blocks the transmission of the pathogen. Additionally, our Gene Set Enrichment Analysis (GSEA) of 140 parasite antigens used in the parasitic vaccines identified enriched features. For example, significant patterns, such as signal, plasma membrane, and entry into host, were found in the antigens of the vaccines against two parasite species: Plasmodium falciparum and Toxoplasma gondii. The analysis found 18 out of the 140 parasite antigens involved with the malaria disease process. Moreover, a majority (15 out of 54) of P. falciparum parasite antigens are localized in the cell membrane. T. gondii antigens, in contrast, have a majority (19/24) of their proteins related to signaling pathways. The antigen-enriched patterns align with the life cycle stage patterns identified in our ontological parasite vaccine modeling.ConclusionsThe updated VO modeling and GSEA analysis capture the influence of the complex parasite life cycles and their associated antigens on vaccine development.

NRPS-like ATRR in Plant-Parasitic Nematodes Involved in Glycine Betaine Metabolism to Promote Parasitism.

Plant-parasitic nematodes (PPNs) are among the most serious phytopathogens and cause widespread and serious damage in major crops. In this study, using a genome mining method, we identified nonribosomal peptide synthetase (NRPS)-like enzymes in genomes of plant-parasitic nematodes, which are conserved with two consecutive reducing domains at the N-terminus (A-T-R1-R2) and homologous to fungal NRPS-like ATRR. We experimentally investigated the roles of the NRPS-like enzyme (MiATRR) in nematode (Meloidogyne incognita) parasitism. Heterologous expression of Miatrr in Saccharomyces cerevisiae can overcome the growth inhibition caused by high concentrations of glycine betaine. RT-qPCR detection shows that Miatrr is significantly upregulated at the early parasitic life stage (J2s in plants) of M. incognita. Host-derived Miatrr RNA interference (RNAi) in Arabidopsis thaliana can significantly decrease the number of galls and egg masses of M. incognita, as well as retard development and reduce the body size of the nematode. Although exogenous glycine betaine and choline have no obvious impact on the survival of free-living M. incognita J2s (pre-parasitic J2s), they impact the performance of the nematode in planta, especially in Miatrr-RNAi plants. Following application of exogenous glycine betaine and choline in the rhizosphere soil of A. thaliana, the numbers of galls and egg masses were obviously reduced by glycine betaine but increased by choline. Based on the knowledge about the function of fungal NRPS-like ATRR and the roles of glycine betaine in host plants and nematodes, we suggest that MiATRR is involved in nematode-plant interaction by acting as a glycine betaine reductase, converting glycine betaine to choline. This may be a universal strategy in plant-parasitic nematodes utilizing NRPS-like ATRR to promote their parasitism on host plants.

Order and timing of infection with different parasite life stages impacts host and parasite life histories.

Co-exposure to multiple parasites can alter parasite success and host life history when compared to single infections. These infection outcomes can be affected by the order of parasite arrival, the host immune response, and the interspecific interactions among co-infecting parasites. In this study, we examined how the arrival order of two trematode parasites, Schistosoma mansoni and Echinostoma caproni, influenced parasite ecology and the life history of their snail host, Biomphalaria glabrata. Snail hosts were exposed to E. caproni cercariae before, with, and after their exposure to S. mansoni miracidia. We then measured the effects of this timing on infection prevalence, infection intensity of E. caproni metacercariae, and cercarial output of S. mansoni, as well as on snail reproduction and survival. Snails infected only with S. mansoni and snails exposed to E. caproni after S. mansoni both shed more cercariae than simultaneously exposed snails. Additionally, S. mansoni prevalence was lower in snails that were first exposed to E. caproni compared to snails that were exposed to E. caproni after S. mansoni. Moreover, snails exposed to E. caproni before S. mansoni did not differ in their survival compared to control snails, whereas simultaneously exposed snails and snails exposed to E. caproni after S. mansoni had lower survival than control snails. Combined, this prevalence and survival data suggest a potential protective role of early E. caproni exposure. The timing of E. caproni exposure impacts S. mansoni establishment and reproduction, but host survival patterns are likely driven by S. mansoni prevalence alone.

Optimization of General Threshold Value for Preprocessing in Plasmodium Parasites Detection

The high mortality rate of malaria makes it a severe disease that spreads throughout all-region by infected female Anopheles mosquitoes, especially in tropical countries. Accurate early malaria detection is one of the ways to reduce the mortality rate. Microscopy-based malaria examinations are still considered the gold standard. Due to numerous large malaria patients with limited parasitologists, an automated detection system is needed as a second opinion to assist parasitologists. This study proposed an optimization method for finding an optimal global threshold value for pre-processing parasite detection. There were three stages of the proposed method. The first is to pre-process digital microscopic images using color channel selection, contrast stretching, and morphological operation. The second is to find the global threshold value using multiple modified Otsu’s. The third is to determine the optimum global threshold value. In the last stage, predicted threshold values are generated using a pattern recognition approach to determine the optimum global threshold value. The proposed method evaluated 468 microscopic images captured from hundreds of thin smear blood slides. The slides are provided by the Department of Parasitology-UGM and the Eijkman Institute for Molecular Biology. The set image contains 691 malaria parasites in all types and life stages of malaria parasites. The proposed method obtained a sensitivity of 99.6 % and the smallest FPs number compared to without the optimization. It indicates that the proposed method has the potential to be implemented in the initial stages of the malaria detection system.

Optimization of General Threshold Value for Preprocessing in Plasmodium Parasites Detection

The high mortality rate of malaria makes it a severe disease that spreads throughout all-region by infected female Anopheles mosquitoes, especially in tropical countries. Accurate early malaria detection is one of the ways to reduce the mortality rate. Microscopy-based malaria examinations are still considered the gold standard. Due to numerous large malaria patients with limited parasitologists, an automated detection system is needed as a second opinion to assist parasitologists. This study proposed an optimization method for finding an optimal global threshold value for pre-processing parasite detection. There were three stages of the proposed method. The first is to pre-process digital microscopic images using color channel selection, contrast stretching, and morphological operation. The second is to find the global threshold value using multiple modified Otsu’s. The third is to determine the optimum global threshold value. In the last stage, predicted threshold values are generated using a pattern recognition approach to determine the optimum global threshold value. The proposed method evaluated 468 microscopic images captured from hundreds of thin smear blood slides. The slides are provided by the Department of Parasitology-UGM and the Eijkman Institute for Molecular Biology. The set image contains 691 malaria parasites in all types and life stages of malaria parasites. The proposed method obtained a sensitivity of 99.6 % and the smallest FPs number compared to without the optimization. It indicates that the proposed method has the potential to be implemented in the initial stages of the malaria detection system.

Plasmodium vivax transcriptomics: What is new?

Malaria is the leading human parasitosis and is transmitted through the bite of anopheline mosquitoes infected with parasites of the genus Plasmodium spp. Among the seven species that cause malaria in humans, Plasmodium vivax is the most prevalent species in Latin America. In recent years, there have been an increasing number of reports of clinical complications caused by P. vivax infections, which were previously neglected and underestimated. P. vivax biology remains with large gaps. The emergence of next-generation sequencing technology has ensured a breakthrough in species knowledge. Coupled with this, the deposition of the P. vivax Sal-1 reference genome allowed an increase in transcriptomics projects by accessing messenger RNA. Thus, the regulation of differential gene expression according to the parasite life stage was verified, and several expressed genes were linked to different biological functions. Today, with the progress associated with RNA sequencing technologies, it is possible to detect nuances and obtain robust results. Discoveries provided by transcriptomic studies allow us to understand topics such as RNA expression and regulation and proteins and metabolic pathways involved during different stages of the parasite life cycle. The information obtained enables a better comprehension of immune system evasion mechanisms; invasion and adhesion strategies used by the parasite; as well as new vaccine targets, potential molecular markers, and others therapeutic targets. In this review, we provide new insights into P. vivax biology by summarizing recent findings in transcriptomic studies.

SlWRKY16 and SlWRKY31 of tomato, negative regulators of plant defense, involved in susceptibility activation following root-knot nematode Meloidogyne javanica infection

The involvement of WRKY transcription factors in plant-nematode interactions, and in particular, how these WRKYs participate in regulating the complex morphological and physiological changes occurring after nematode infection, are the topic of active research. We characterized the functional role of the unstudied tomato WRKY genes SlWRKY16 and SlWRKY31 in regulating tomato roots’ response to infection by the root-knot nematode Meloidogyne javanica. Using promoter–GUS reporter gene fusions and qRT-PCR, we show that both SlWRKYs are predominantly expressed during the first half of the parasitic life stages, when feeding-site induction and construction occur. Expression of SlWRKY16 increased sharply 15 days after inoculation, whereas SlWRKY31 was already induced earlier, but reached its maximum expression at this time. Both genes were downregulated at the mature female stage. To determine biological function, we produced transgenic lines overexpressing SlWRKY16 and SlWRKY31 in tomato hairy roots. Overexpression of both genes resulted in enhanced M. javanica infection, reflected by increased galling occurrence and reproduction. Expression profiling of marker genes responsive to defense-associated phytohormones indicated reductions in salicylic acid defense-related PR-1 and jasmonic acid defense-related PI in inoculated roots overexpressing SlWRK16 and SlWRKY31, respectively. Our results suggest that SlWRKY16 and SlWRKY31 function as negative regulators of plant immunity induced upon nematode infection.

Population-level variation in insecticide tolerance across three life stages of the trematode Echinostoma trivolvis

Ecotoxicological studies using single test populations may miss the inherent variation of natural systems and limit our understanding of how contaminants affect focal species. Though population-level variation in pesticide tolerance is commonly observed in host taxa, few studies have assessed population-level differences in the tolerance of parasites to different contaminants. We investigated population-level variation in insecticide tolerance of three Echinostoma trivolvis life stages (egg, miracidium, and cercaria) to three insecticides (carbaryl, chlorpyrifos, and diazinon). We tested two relevant metrics of insecticide tolerance—baseline and induced—across up to eight different parasite populations per life stage. Across all life stages, the insecticide treatments tended to reduce survival, but the magnitude of their effects often varied significantly among populations. Surprisingly, we found that exposure to chlorpyrifos increased the hatching success of echinostome eggs relative to the control treatment in three of six tested populations. We also found that cercariae shed from snails previously exposed to a sublethal concentration of chlorpyrifos had a significantly lower mortality rate when subsequently exposed to a lethal concentration of chlorpyrifos relative to individuals from snails that were not previously exposed; this suggests inducible tolerance in cercariae. We found no evidence that insecticide tolerance is correlated across parasite life stages within a population. Together the findings of our study demonstrate that single-population toxicity assays may greatly over- or underestimate the effects of pesticides on the survival of free-living parasite stages, insecticide tolerance levels may not be predictable from one parasite life stage to the next, and insecticides can have both expected and counterintuitive effects on non-target taxa.

Differential Effects of Antimalarial Drugs on Parasite Clearance Rates Are Reflected by Plasmodium falciparum Ring Ratio.

The location of Plasmodium falciparum within the body is determined by the life cycle of the parasite; young rings are in the peripheral blood, whereas mature parasites are sequestered in deep tissues. We can calculate a "ring ratio," the proportion of parasites in the periphery to the total number of parasites in the body. Artesunate acts on all parasite life stages, whereas quinine is effective only on sequestered parasites. Children with cerebral malaria (CM) treated with artesunate clear parasites faster than those treated with quinine. In this study, we established the relationship between ring ratio and parasite clearance rate and used the ring ratio to determine if the benefit derived from artesunate treatment could be attributed to its broader effect on life cycle stages. Ring ratios were calculated for 400 hospitalized children with CM in Blantyre, Malawi between 2010 and 2019 (quinine: 2010-2013, artesunate: 2014-2019). In both treatment groups, parasite clearance rates were positively associated with the ring ratios, with a stronger association in the artesunate era than the quinine era. In the quinine era, an increase of 1-unit log10 difference between parasitemia and plasma P falciparum histidine-rich protein 2 (a proxy for ring ratio) resulted in a 0.27-unit increase in the parasite clearance rate, whereas in the artesunate era an equal increase resulted in a 0.41-unit increase (P = .04 for the difference). This analysis provides in vivo evidence supporting the hypothesis that more rapid parasite clearance rates in artesunate recipients are due to its superiority over quinine in killing ring-stage parasites.

Haemonchus contortus Adopt Isolate-Specific Life History Strategies to Optimize Fitness and Overcome Obstacles in Their Environment: Experimental Evidence.

Gastrointestinal nematodes (GIN) use flexible life history strategies to maintain their fitness under environmental challenges. Costs incurred by a challenge to one life trait can be recouped by increasing the expression of subsequent life traits throughout their life cycle. Anticipating how parasites respond to the challenge of control interventions is critical for the long-term sustainability of the practice and to further ensure that the parasites withstand favourable adaptive responses. There is currently limited information on whether distinct populations of a GIN species respond to the same environmental challenge in a consistent manner, with similar alterations to their life history strategies or comparable fitness outcomes. This study compared the life history traits and experimental fitness of three distinct Haemonchus contortus isolates exposed to environmental challenges at both the parasitic (i.e., passage through resistant or susceptible sheep) and free-living (i.e., exposure to diverse climatic conditions) life stages. The key findings show that H. contortus maintain their fitness under challenge with isolate-specific alterations to their life history strategies. Further, partial exploration of the H. contortus isolates transcriptomes using cDNA-AFLP methods confirmed disparate expression profiles between them. These results bring fresh insights into our understanding of the non-genetic adaptive processes of GIN that may hinder the efficacy of parasite control strategies.

Plasmodium falciparum gametocyte carriage in longitudinally monitored incident infections is associated with duration of infection and human host factors

Malaria transmission depends on the presence of Plasmodium gametocytes that are the only parasite life stage that can infect mosquitoes. Gametocyte production varies between infections and over the course of infections. Infection duration is highly important for gametocyte production but poorly quantified. Between 2017 and 2019 an all-age cohort of individuals from Tororo, eastern Uganda was followed by continuous passive and routine assessments. We longitudinally monitored 104 incident infections from 98 individuals who were sampled once every 28 days and on any day of symptoms. Among infections that lasted ≥ 3 months, gametocyte appearance was near-universal with 96% of infections having detectable gametocytes prior to clearance. However, most infections were of much shorter duration; 55.7% of asymptomatic infections were detected only once. When considering all asymptomatic infections, regardless of their duration, only 36.3% had detectable gametocytes on at least one time-point prior to parasite clearance. Infections in individuals with sickle-cell trait (HbAS) were more likely to have gametocytes detected (Hazard Rate (HR) = 2.68, 95% CI 1.12, 6.38; p = 0.0231) and had gametocytes detected at higher densities (Density Ratio (DR) = 9.19, 95% CI 2.79, 30.23; p = 0.0002) compared to infections in wildtype (HbAA) individuals. Our findings suggest that a large proportion of incident infections is too short in duration and of too low density to contribute to onward transmission.

Alterations to unionid transformation during agricultural and urban contaminants of emerging concern exposures

Highly imperiled unionids have a complex life cycle including the metamorphosis of an obligate parasite life stage, larval glochidia, to the juvenile stage. Despite the known vulnerabilities of both glochidia and juveniles to pollutants, little is known on how metamorphosis success may be affected by chemical stress. Disruption of the transformation process in which glochidia encyst on the gills of a host fish, could lead to lowered recruitment and population declines. Transformation rates of Lampsilis cardium on host fish Micropterus salmoides were empirically derived from experimental exposures to low, medium, or high concentrations of an agricultural or urban mixture of contaminants of emerging concern (CECs) over two exposure durations. Transformation was characterized by: (1) a zero-inflated Poisson general linear mixed effects model to compare difference in transformation between exposure durations and (2) time response curves to describe the transformation curve using long-term exposure data. Lampsilis cardium transformation was similar between exposure durations. When compared to controls, CEC stress significantly reduced juvenile production (p « 0.05) except for the agricultural medium treatment and tended to increased encapsulation duration which while statistically insignificant (p = 0.16) may have ecological relevancy. Combining the empirically derived reduction of transformation rates with parameters values from the literature, a Lefkovich stage-based population model predicted strong declines in population size of L. cardium for all treatments if these results hold in nature. Management focus on urban CECs may lead to best conservation efforts though agricultural CECs may also have a concentration dependent impact on transformation and therefore overall recruitment and conservation success.

Mosquito bites and stage-specific antibody responses against Plasmodium falciparum in southern Ghana

BackgroundThe human host elicits specific immune responses after exposure to various life stages of the malaria parasite as well as components of mosquito saliva injected into the host during a mosquito bite. This study describes differences in IgG responses against antigens derived from the sporozoite (PfCSP), asexual stage parasite (PfEBA175) and the gametocyte (Pfs230), in addition to an Anopheles gambiae salivary gland antigen (gSG6-P1), in two communities in Ghana with similar blood stage malaria parasite prevalence.MethodsThis study used archived plasma samples collected from an earlier cross-sectional study that enrolled volunteers aged from 6 months to 70 years from Simiw, peri-urban community (N = 347) and Obom, rural community (N = 291). An archived thick and thin blood smear for microscopy was used for the estimation of Plasmodium parasite density and species and DNA extraction from blood spots and P. falciparum confirmation was performed using PCR. This study used the stored plasma samples to determine IgG antibody levels to P. falciparum and Anopheles salivary antigens using indirect ELISA.ResultsIndividuals from Simiw had significantly higher levels of IgG against mosquito gSG6-P1 [median (95%CI)] [2.590 (2.452–2.783) ng/mL] compared to those from Obom [2.119 (1.957–2.345) ng/mL], p < 0.0001. Both IgG responses against Pfs230proC (p = 0.0006), and PfCSP (p = 0.002) were significantly lower in volunteers from Simiw compared to the participants from Obom. The seroprevalence of PfEBA-175.5R (p = 0.8613), gSG6-P1 (p = 0.0704), PfCSP (p = 0.7798) IgG were all similar in Obom and Simiw. However, Pfs230 seroprevalence was significantly higher at Obom compared to Simiw (p = 0.0006). Spearman correlation analysis showed no significant association between IgG responses against gSG6-P1, PfCSP, Pfs230proC and PfEBA-175.5R and parasite density at both Obom and Simiw (p > 0.05).ConclusionIn conclusion, the study showed that participants from Simiw had higher concentrations of circulating gSG6-P1 IgG antibodies but lower concentrations of P. falciparum antibodies, PfCSP IgG and Pfs230proC IgG compared to participants from Obom.

Proximity-Dependent Biotinylation and Identification of Flagellar Proteins in Trypanosoma cruzi.

The flagellated kinetoplastid protozoan and causative agent of human Chagas disease, Trypanosoma cruzi, inhabits both invertebrate and mammalian hosts over the course of its complex life cycle. In these disparate environments, T. cruzi uses its single flagellum to propel motile life stages and, in some instances, to establish intimate contact with the host. Beyond its role in motility, the functional capabilities of the T. cruzi flagellum have not been defined. Moreover, the lack of proteomic information for this organelle, in any parasite life stage, has limited functional investigation. In this study, we employed a proximity-dependent biotinylation approach based on the differential targeting of the biotin ligase TurboID to the flagellum or cytosol in replicative stages of T. cruzi to identify proteins that are enriched in the flagellum by mass spectrometry. Proteomic analysis of the resulting biotinylated protein fractions yielded 218 candidate flagellar proteins in T. cruzi epimastigotes (insect stage) and 99 proteins in intracellular amastigotes (mammalian stage). Forty of these enriched flagellar proteins were common to both parasite life stages and included orthologs of known flagellar proteins in other trypanosomatid species, proteins specific to the T. cruzi lineage and hypothetical proteins. With the validation of flagellar localization for several of the identified candidates, our results demonstrate that TurboID-based proximity proteomics is an effective tool for probing subcellular compartments in T. cruzi. The proteomic data sets generated in this work offer a valuable resource to facilitate functional investigation of the understudied T. cruzi flagellum. IMPORTANCE Trypanosoma cruzi is a protozoan parasite that causes Chagas disease, which causes substantial morbidity and mortality in South and Central America. Throughout its life cycle, T. cruzi interacts with insect and mammalian hosts via its single flagellum, establishing intimate contact with host membranes. Currently, few flagellar proteins have been identified in T. cruzi that could provide insight into the mechanisms involved in mediating physical and biochemical interactions with the host. Here, we set out to identify flagellar proteins in the main replicative stages of T. cruzi using a proximity-labeling approach coupled with mass spectrometry. The >200 candidate flagellar proteins identified represent the first large-scale identification of candidate flagellar proteins in T. cruzi with preliminary validation. These data offer new avenues to investigate the biology of T. cruzi-host interactions, a promising area for development of new strategies aimed at the control of this pathogen.

Tracking life cycles of parasites across a broad taxonomic scale in a marine ecosystem

Parasitic helminths exhibit remarkable diversity in their life cycles, although few parasite species have their whole life cycles resolved. Owing to the fact that parasite life stages within hosts are often not comparable using morphological data, genetic data provides convincing evidence of transmission pathways between intermediate and definitive hosts. We took this approach to an ecosystem level, genetically matching parasite (acanthocephalan, cestode, nematode and trematode) life stages across a broad taxonomic range of intermediate and definitive hosts (invertebrates, seabirds, elasmobranchs and teleost fish) in Otago’s (New Zealand) coastal marine ecosystem. We identified which transmission routes are utilized by the most parasite species and assessed which intermediate hosts are most important in facilitating the transmission of parasites in this ecosystem. Our findings reveal 59 new records of larval parasites infecting their respective intermediate hosts and 289 transmission pathways utilized by 35 helminth species to complete their life cycles. Sprat, triplefin and arrow squid all hosted the highest number of larval parasite species, suggesting they play important roles as intermediate hosts. We then used the new life cycle data to provide a synthetic overview of the life cycles known for various parasite groups in New Zealand. This study highlights how studying parasite life cycles can enhance our understanding of the ecology and evolution of parasites and hosts in natural systems, beyond simply resolving life cycles.

The Intracellular Amastigote of Trypanosoma cruzi Maintains an Actively Beating Flagellum.

Throughout its complex life cycle, the uniflagellate parasitic protist, Trypanosoma cruzi, adapts to different host environments by transitioning between elongated motile extracellular stages and a nonmotile intracellular amastigote stage that replicates in the cytoplasm of mammalian host cells. Intracellular T. cruzi amastigotes retain a short flagellum that extends beyond the opening of the flagellar pocket with access to the extracellular milieu. Contrary to the long-held view that the T. cruzi amastigote flagellum is inert, we report that this organelle is motile and displays quasiperiodic beating inside mammalian host cells. Kymograph analysis determined an average flagellar beat frequency of ~0.7 Hz for intracellular amastigotes and similar beat frequencies for extracellular amastigotes following their isolation from host cells. Inhibitor studies reveal that flagellar motility in T. cruzi amastigotes is critically dependent on parasite mitochondrial oxidative phosphorylation. These novel observations reveal that flagellar motility is an intrinsic property of T. cruzi amastigotes and suggest that this organelle may play an active role in the parasite infection process. IMPORTANCE Understanding the interplay between intracellular pathogens and their hosts is vital to the development of new treatments and preventive strategies. The intracellular "amastigote" stage of the Chagas disease parasite, Trypanosoma cruzi, is a critical but understudied parasitic life stage. Previous work established that cytosolically localized T. cruzi amastigotes engage physically and selectively with host mitochondria using their short, single flagellum. The current study was initiated to examine the dynamics of the parasite flagellum-host mitochondrial interaction through live confocal imaging and led to the unexpected discovery that the T. cruzi amastigote flagellum is motile.

Two Candidate Meloidogyne javanica Effector Genes, MjShKT and MjPUT3: A Functional Investigation of Their Roles in Regulating Nematode Parasitism.

During parasitism, root-knot nematode Meloidogyne spp. inject molecules termed effectors that have multifunctional roles in construction and maintenance of nematode feeding sites. As an outcome of transcriptomic analysis of Meloidogyne javanica, we identified and characterized two differentially expressed genes encoding the predicted proteins MjShKT, carrying a Stichodactyla toxin (ShKT) domain, and MjPUT3, carrying a ground-like domain, both expressed during nematode parasitism of the tomato plant. Fluorescence in-situ hybridization revealed expression of MjShKT and MjPUT3 in the dorsal esophageal glands, suggesting their injection into host cells. MjShKT expression was upregulated during the parasitic life stages, to a maximum at the mature female stage, whereas MjPUT3 expression increased in third- to fourth-stage juveniles. Subcellular in-planta localization of MjShKT and MjPUT3 using a fused fluorescence marker indicated MjShKT co-occurrence with the endoplasmic reticulum, the perinuclear endoplasmatic reticulum, and the Golgi organelle markers, while MjPUT3 localized, to some extent, within the endoplasmatic reticulum and was clearly observed within the nucleoplasm. MjShKT inhibited programmed cell death induced by overexpression of MAPKKKα and Gpa2/RBP-1. Overexpression of MjShKT in tomato hairy roots allowed an increase in nematode reproduction, as indicated by the high number of eggs produced on roots overexpressing MjShKT. Roots overexpressing MjPUT3 were characterized by enhanced root growth, with no effect on nematode development on those roots. Investigation of the two candidate effectors suggested that MjShKT is mainly involved in manipulating the plant effector-triggered immune response toward establishment and maintenance of active feeding sites, whereas MjPUT3 might modulate roots morphology in favor of nematode fitness in the host roots. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Within-host and external environments differentially shape β-diversity across parasite life stages.

Uncovering drivers of community assembly is a key aspect of learning how biological communities function. Drivers of community similarity can be especially useful in this task as they affect assemblage-level changes that lead to differences in species diversity between habitats. Concepts of β-diversity originally developed for use in free-living communities have been widely applied to parasite communities to gain insight into how infection risk changes with local conditions by comparing parasite communities across abiotic and biotic gradients. Factors shaping β-diversity in communities of immature parasites, such as larvae, are largely unknown. This is a key knowledge gap as larvae are frequently the infective life-stage and understanding variation in these larval communities is thus key for disease prevention. Our goal was to uncover links between β-diversity of parasite communities at different life stages; therefore, we used gastrointestinal nematodes infecting African buffalo in Kruger National Park, South Africa, to investigate within-host and extra-host drivers of adult and larval parasite community similarity. We employed a cross-sectional approach using PERMANOVA that examined each worm community at a single time point to assess independent drivers of β-diversity in larvae and adults as well as a longitudinal approach with path analysis where adult and larval communities from the same host were compared to better link drivers of β-diversity between these two life stages. Using the cross-sectional approach, we generally found that intrinsic, within-host traits had significant effects on β-diversity of adult nematode communities, while extrinsic, extra-host variables had significant effects on β-diversity of larval nematode communities. However, the longitudinal approach provided evidence that intrinsic, within-host factors affected the larval community indirectly via the adult community. Our results provide key data for the comparison of community-level processes where adult and immature stages inhabit vastly different habitats (i.e. within-host vs. abiotic environment). In the context of parasitism, this helps elucidate host infection risk via larval stages and the drivers that shape persistence of adult parasite assemblages, both of which are useful for predicting and preventing infectious disease.