Search

Blastocystis is the most prevalent intestinal eukaryotic microorganism with significant impacts on both human and animal health. Despite extensive research, its pathogenicity remains controversial. The COST Action CA21105, " Blastocystis under One Health" (OneHealthBlastocystis), aims to bridge gaps in our understanding by fostering a multidisciplinary network. This initiative focuses on developing standardised diagnostic methodologies, establishing a comprehensive subtype and microbiome databank, and promoting capacity building through education and collaboration. The Action is structured into five working groups, each targeting specific aspects of Blastocystis research, including epidemiology, diagnostics, 'omics technologies, in vivo and in vitro investigations, and data dissemination. By integrating advances across medical, veterinary, public, and environmental health, this initiative seeks to harmonise diagnostics, improve public health policies, and foster innovative research, ultimately enhancing our understanding of Blastocystis and its role in health and disease. This collaborative effort is expected to lead to significant advancements and practical applications, benefiting the scientific community and public health., Competing Interests: No competing interests were disclosed., (Copyright: © 2024 Tsaousis AD et al.)

Background: Blastocystis is a unicellular eukaryote commonly found in the intestinal tract of humans and other animals. The prevalence of Blastocystis has been investigated in both developed and developing countries, yet its occurrence and distribution in rural locations has been less studied. Herein, we aimed to examine the distribution of Blastocystis colonization in Thai adults representing background populations along a rural/peri-urban gradient, as well as associations between colonization and personal characteristics., Methodology: A total of 238 participants were recruited from rural and peri-urban areas situated in three provinces. The presence of Blastocystis in feces was evaluated using PCR and qPCR. Information on gender, age, region (province), rural/peri-urban location, and body mass index (BMI) was collected., Principal Findings: The overall rate of Blastocystis carriage was 67.2%. Univariate analysis revealed significant associations between Blastocystis carriage and region (p<0.05), location (p<0.001) and age group (p<0.05). Logistic regression analysis revealed that rural/peri-urban location and BMI were significantly associated with Blastocystis carriage. Nine subtypes (ST1-ST7, ST10 and ST23) were identified with ST3, ST7 and ST1 as the most abundant ones, in this order. The greatest diversity of subtypes, in terms of numbers, was found in the middle aged group (nine subtypes), while the least diversity was found in the young adult and obese (three subtypes each) groups., Conclusions: This study increases the understanding of the epidemiology of Blastocystis colonization and its association with population parameters and characteristics in middle-income countries., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Jinatham et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Gregarines are symbiotic protists that are found in a broad spectrum of invertebrates, including insects, crustaceans, and annelids. Among these the globally distributed amphipod Gammarus pulex is one of the earliest recognized hosts for aquatic gregarines and is prevalent among macroinvertebrates in freshwater environments. In this study, samples of G. pulex were collected in the Water of Leith river, Scotland, UK. Gregarines were identified using light and scanning electron microscopy as well as standard molecular techniques. We identified three septate eugregarine symbionts-Heliospora longissima, Cephaloidophora gammari, and the here newly characterized Cephaloidophora conus n. sp. (formerly Cephaloidophora sp.) associated with Gammarus pulex in the Water of Leith. Prevalences for identified gregarine species were calculated and seasonal dynamics of gregarine infections/colonization were analyzed. Prevalences were highest in autumn and spring reaching almost 50 %. While the two Cephaloidophora species showed similar colonization patterns, the prevalence of Heliospora showed an opposite trend. Identifying gregarine infection/colonization patterns is one step towards better understanding the gregarine-host relationship, as well as possible impacts of the gregarines on their hosts., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier GmbH.. All rights reserved.)

Cryptosporidium is an important water-borne and food-borne parasite with a high burden of disease. This organism has been shown to contaminate various leafy vegetables; however, studies assessing the presence of Cryptosporidium spp in pre-washed and ready-to-eat vegetables are limited. Routine surveillance in the UK revealed a nationwide exceedance of human cases of Cryptosporidium. Therefore, this study aims to assess the presence of this parasite in pre-washed vegetables from supermarkets in the UK. A total of 36 samples were purchased from four different supermarkets. A nested PCR targeting the SSU rRNA was carried out on 24 samples, 58% were PCR-positive for Cryptosporidium. Sanger sequencing confirmed that, of these sequences, 4/24 (17%) produced significant similarities to Cryptosporidium parvum. This study provides evidence for the presence of C. parvum in pre-washed and ready-to-eat vegetables. Future work to identify the point of contamination is required., (© 2024. The Author(s).)

Blastocystis is the most common gastrointestinal protist found in humans and animals. Although the clinical significance of Blastocystis remains unclear, the organism is increasingly being viewed as a commensal member of the gut microbiome. However, its impact on the microbiome is still being debated. It is unclear whether Blastocystis promotes a healthy gut and microbiome directly or whether it is more likely to colonize and persist in a healthy gut environment. In healthy people, Blastocystis is frequently associated with increased bacterial diversity and significant differences in the gut microbiome. Based on current knowledge, it is not possible to determine whether differences in the gut microbiome are the cause or result of Blastocystis colonization. Although it is possible that some aspects of this eukaryote's role in the intestinal microbiome remain unknown and that its effects vary, possibly due to subtype and intra-subtype variations and immune modulation, more research is needed to characterize these mechanisms in greater detail. This review covers recent findings on the effects of Blastocystis in the gut microbiome and immune modulation, its impact on the microbiome in autoimmune diseases, whether Blastocystis has a role like bacteria in the gut-brain axis, and its relationship with probiotics.

Purpose: The present study aimed to estimate the prevalence and molecular characterization of Cryptosporidium spp. in six different fish species both from marine and freshwater environments., Methods: During a period of 2 years (2018-2020), a total of 415 fecal samples and 565 intestinal scrapings were collected in seven provinces from the central and eastern Algeria. From those, 860 fish belonged to six different species, two of which are cultured marine and four are wild freshwater fish. All samples were screened for Cryptosporidium spp. presence using molecular techniques. Nested PCR approach was performed to amplify partial sequences of the small subunit ribosomal RNA (SSU rRNA) and 60-kDa glycoprotein (GP60) genes for Cryptosporidium genotyping and subtyping. Detailed statistical analysis was performed to assess the prevalence variation of Cryptosporidium infection according to different risk factors., Results: Nested PCR analysis of SSU gene revealed 173 Cryptosporidium positive fish, giving an overall prevalence of 20.11% (17.5-23.0). Cryptosporidium spp. was detected in 8.93% (42/470) of cultured marine fish and 33.58% (131/390) of wild freshwater fish. Overall, the prevalence was affected by all studied risk factors, except the gender. Molecular characterization and subtyping of Cryptosporidium isolates showed occurrence of IIaA16G2R1 and IIaA17G2R1 subtypes of C. parvum in the fish species Sparus aurata., Conclusion: The present study provides the first epidemiological data on the prevalence and associated risk factors of Cryptosporidium spp. in farmed marine and wild freshwater fish and the first molecular data on the occurrence of zoonotic C. parvum in fish from North Africa (Algeria)., (© 2022. The Author(s) under exclusive licence to Witold Stefański Institute of Parasitology, Polish Academy of Sciences.)

Naegleria fowleri is both a pathogenic and a free-living microbial eukaryote, responsible for the development of primary amoebic meningoencephalitis (PAM) in humans. PAM is a rapid, severe and fatal underestimated infectious disease, which has been reported in countries with warmer climates. The major drawbacks with PAM are the lack of effective therapies and delay in diagnosis. The current frontline treatment presents a low rate of recovery (5%) and severe adverse effects. For example, many drug candidates lack efficacy, since they do not effectively cross the blood-brain-barrier. Consequently, more effective drugs are urgently needed. Herein, we report a new in vitro method suitable for medium- and high-throughput drug discovery assays, using the closely related Naegleria gruberi as a model. We have subsequently used this method to screen a library of 1175 Food and Drug Administration-approved drugs. As a result, we present three drugs (camptothecin, pyrimethamine, and terbinafine) that can be repurposed, and are anticipated to readily cross the blood-brain-barrier with activity against Naegleria species in therapeutically achievable concentrations. Successively, we integrated several in vitro assays that resulted in identifying fast-acting and high amoebicidal drugs. In conclusion, we present a new approach for the identification of anti-Naegleria drugs along with three potential drug candidates for further development for the treatment of PAM., (Copyright © 2021. Published by Elsevier Ltd.)

The type 2 secretion system (T2SS) is present in some Gram-negative eubacteria and used to secrete proteins across the outer membrane. Here we report that certain representative heteroloboseans, jakobids, malawimonads and hemimastigotes unexpectedly possess homologues of core T2SS components. We show that at least some of them are present in mitochondria, and their behaviour in biochemical assays is consistent with the presence of a mitochondrial T2SS-derived system (miT2SS). We additionally identified 23 protein families co-occurring with miT2SS in eukaryotes. Seven of these proteins could be directly linked to the core miT2SS by functional data and/or sequence features, whereas others may represent different parts of a broader functional pathway, possibly also involving the peroxisome. Its distribution in eukaryotes and phylogenetic evidence together indicate that the miT2SS-centred pathway is an ancestral eukaryotic trait. Our findings thus have direct implications for the functional properties of the early mitochondrion.

State-of-the-art computer-vision algorithms rely on big and accurately annotated data, which are expensive, laborious and time-consuming to generate. This task is even more challenging when it comes to microbiological images, because they require specialized expertise for accurate annotation. Previous studies show that crowdsourcing and assistive-annotation tools are two potential solutions to address this challenge. In this work, we have developed a web-based platform to enable crowdsourcing annotation of image data; the platform is powered by a semi-automated assistive tool to support non-expert annotators to improve the annotation efficiency. The behavior of annotators with and without the assistive tool is analyzed, using biological images of different complexity. More specifically, non-experts have been asked to use the platform to annotate microbiological images of gut parasites, which are compared with annotations by experts. A quantitative evaluation is carried out on the results, confirming that the assistive tools can noticeably decrease the non-expert annotation's cost (time, click, interaction, etc.) while preserving or even improving the annotation's quality. The annotation quality of non-experts has been investigated using IoU (intersection over union), precision and recall; based on this analysis we propose some ideas on how to better design similar crowdsourcing and assistive platforms., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Diverse microbial ecosystems underpin life in the sea. Among these microbes are many unicellular eukaryotes that span the diversity of the eukaryotic tree of life. However, genetic tractability has been limited to a few species, which do not represent eukaryotic diversity or environmentally relevant taxa. Here, we report on the development of genetic tools in a range of protists primarily from marine environments. We present evidence for foreign DNA delivery and expression in 13 species never before transformed and for advancement of tools for eight other species, as well as potential reasons for why transformation of yet another 17 species tested was not achieved. Our resource in genetic manipulation will provide insights into the ancestral eukaryotic lifeforms, general eukaryote cell biology, protein diversification and the evolution of cellular pathways.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

The microbial parasite Blastocystis colonizes the large intestines of numerous animal species and increasing evidence has linked Blastocystis infection to enteric diseases with signs and symptoms including abdominal pain, constipation, diarrhea, nausea, vomiting, and flatulence. It has also recently been reported to be an important member of the host intestinal microbiota. Despite significant advances in our understanding of Blastocystis cell biology and host-parasite interactions, a genetic modification tool is absent. In this study, we successfully established a robust gene delivery protocol for Blastocystis subtype 7 (ST7) and ectopic protein expression was further tested using a high sensitivity nano-luciferase (Nluc) reporter system, with promoter regions from several genes. Among them, a strong promoter encompassing a region upstream of the legumain 5' UTR was identified. Using this promoter combined with the legumain 3' UTR, which contains a conserved, precise polyadenylation signal, a robust transient transfection technique was established for the first time in Blastocystis. This system was validated by ectopic expression of proteins harbouring specific localization signals. The establishment of a robust, reproducible gene modification system for Blastocystis is a significant advance for Blastocystis research both in vitro and in vivo. This technique will spearhead further research to understand the parasite's biology, its role in health and disease, along with novel ways to combat the parasite.

Emerging infectious diseases are frequently zoonotic, often originating in wildlife, but enteric protozoa are considered relatively minor contributors. Opinions regarding whether pathogenic enteric protozoa may be transmitted between wildlife and humans have been shaped by our investigation tools, and have led to oscillations regarding whether particular species are zoonotic or have host-adapted life cycles. When the only approach for identifying enteric protozoa was morphology, it was assumed that many enteric protozoa colonized multiple hosts and were probably zoonotic. When molecular tools revealed genetic differences in morphologically identical species colonizing humans and other animals, host specificity seemed more likely. Parasites from animals found to be genetically identical - at the few genes investigated - to morphologically indistinguishable parasites from human hosts, were described as having zoonotic potential. More discriminatory molecular tools have now sub-divided some protozoa again. Meanwhile, some infection events indicate that, circumstances permitting, some "host-specific" protozoa, can actually infect various hosts. These repeated changes in our understanding are linked intrinsically to the investigative tools available. Here we review how molecular tools have assisted, or sometimes confused, our understanding of the public health threat from nine enteric protozoa and example wildlife hosts ( Balantoides coli - wild boar; Blastocystis sp. - wild rodents; Cryptosporidium spp. - wild fish; Encephalitozoon spp. - wild birds; Entamoeba spp. - non-human primates; Enterocytozoon bieneusi - wild cervids; Giardia duodenalis - red foxes; Sarcocystis nesbitti - snakes; Toxoplasma gondii - bobcats). Molecular tools have provided evidence that some enteric protozoa in wildlife may infect humans, but due to limited discriminatory power, often only the zoonotic potential of the parasite is indicated. Molecular analyses, which should be as discriminatory as possible, are one, but not the only, component of the toolbox for investigating potential public health impacts from pathogenic enteric protozoa in wildlife.

In eukaryotes, determination of the subcellular location of a novel protein encoded in genomic or transcriptomic data provides useful clues as to its possible function. However, experimental localization studies are expensive and time-consuming. As a result, accurate in silico prediction of subcellular localization from sequence data alone is an extremely important field of study in bioinformatics. This is especially so as genomic studies expand beyond model system organisms to encompass the full diversity of eukaryotes. Here we review some of the more commonly used programs for prediction of proteins that function in mitochondria, or mitochondrion-related organelles in diverse eukaryotic lineages and provide recommendations on how to apply these methods. Furthermore, we compare the predictive performance of these programs on a mixed set of mitochondrial and non-mitochondrial proteins. Although N-terminal targeting peptide prediction programs tend to have the highest accuracy, they cannot be effectively used for partial coding sequences derived from high-throughput expressed sequence tag surveys where data for the N-terminus of the encoded protein is often missing. Therefore methods that do not rely on the presence of an N-terminal targeting sequence alone are extremely useful, especially for expressed sequence tag data. The best strategy for classification of unknown proteins is to use multiple programs, incorporating a variety of prediction strategies, and closely examine the predictions with an understanding of how each of those programs will likely handle the data.