Haem transporter HRG-1 is essential in the barber's pole worm and an intervention target candidate.

Parasitic roundworms (nematodes) have lost genes involved in the de novo biosynthesis of haem, but have evolved the capacity to acquire and utilise exogenous haem from host animals. However, very little is known about the processes or mechanisms underlying haem acquisition and utilisation in parasites. Here, we reveal that HRG-1 is a conserved and unique haem transporter in a broad range of parasitic nematodes of socioeconomic importance, which enables haem uptake via intestinal cells, facilitates cellular haem utilisation through the endo-lysosomal system, and exhibits a conspicuous distribution at the basal laminae covering the alimentary tract, muscles and gonads. The broader tissue expression pattern of HRG-1 in Haemonchus contortus (barber's pole worm) compared with its orthologues in the free-living nematode Caenorhabditis elegans indicates critical involvement of this unique haem transporter in haem homeostasis in tissues and organs of the parasitic nematode. RNAi-mediated gene knockdown of hrg-1 resulted in sick and lethal phenotypes of infective larvae of H. contortus, which could only be rescued by supplementation of exogenous haem in the early developmental stage. Notably, the RNAi-treated infective larvae could not establish infection or survive in the mammalian host, suggesting an indispensable role of this haem transporter in the survival of this parasite. This study provides new insights into the haem biology of a parasitic nematode, demonstrates that haem acquisition by HRG-1 is essential for H. contortus survival and infection, and suggests that HRG-1 could be an intervention target candidate in a range of parasitic nematodes.

An apicoplast-localized deubiquitinase contributes to the cell growth and apicoplast homeostasis of Toxoplasma gondii.

Toxoplasma gondii is among the most important parasites worldwide. The apicoplast is a unique organelle shared by all Apicomplexan protozoa. Increasing lines of evidence suggest that the apicoplast possesses its own ubiquitination system. Deubiquitination is a crucial step executed by deubiquitinase (DUB) during protein ubiquitination. While multiple components of ubiquitination have been identified in T. gondii, the deubiquitinases involved remain unknown. The aim of the current study was to delineate the localization of TgOTU7 and elucidate its functions. TgOTU7 was specifically localized at the apicoplast, and its expression was largely regulated during the cell cycle. Additionally, TgOTU7 efficiently breaks down ubiquitin chains, exhibits linkage-nonspecific deubiquitinating activity and is critical for the lytic cycle and apicoplast biogenesis, similar to the transcription of the apicoplast genome and the nuclear genes encoding apicoplast-targeted proteins. Taken together, the results indicate that the newly described deubiquitinase TgOTU7 specifically localizes to the apicoplast and affects the cell growth and apicoplast homeostasis of T. gondii.

Acyl-CoA oxidase ACOX-1 interacts with a peroxin PEX-5 to play roles in larval development of Haemonchus contortus.

Hypobiosis (facultative developmental arrest) is the most important life-cycle adaptation ensuring survival of parasitic nematodes under adverse conditions. Little is known about such survival mechanisms, although ascarosides (ascarylose with fatty acid-derived side chains) have been reported to mediate the formation of dauer larvae in the free-living nematode Caenorhabditis elegans. Here, we investigated the role of a key gene acox-1, in the larval development of Haemonchus contortus, one of the most important parasitic nematodes that employ hypobiosis as a routine survival mechanism. In this parasite, acox-1 encodes three proteins (ACOXs) that all show a fatty acid oxidation activity in vitro and in vivo, and interact with a peroxin PEX-5 in peroxisomes. In particular, a peroxisomal targeting signal type1 (PTS1) sequence is required for ACOX-1 to be recognised by PEX-5. Analyses on developmental transcription and tissue expression show that acox-1 is predominantly expressed in the intestine and hypodermis of H. contortus, particularly in the early larval stages in the environment and the arrested fourth larval stage within host animals. Knockdown of acox-1 and pex-5 in parasitic H. contortus shows that these genes play essential roles in the post-embryonic larval development and likely in the facultative arrest of this species. A comprehensive understanding of these genes and the associated ß-oxidation cycle of fatty acids should provide novel insights into the developmental regulation of parasitic nematodes, and into the discovery of novel interventions for species of socioeconomic importance.

Hymenolepis diminuta, phylogenetic analyses of nuclear rRNA + ITS and mitochondrial cox1 and its infections in non-human primates.

Hymenolepis diminuta is a tapeworm commonly found worldwide in small rodents such as rats with occasional reports in other definitive hosts such as primates including chimpanzees and humans. It has not been reported in African green monkey (AGM, Chlorocebus sabaeus), and the parasite's molecular phenotype and phylogeny remain primitively sketchy. The aims of the current study were to determine if H. diminuta infected AGMs, to molecularly characterize H. diminuta and to review its infection in non-human primates. Feces of AGMs were examined visually for adult helminths and microscopically for eggs using centrifugation flotation. Total DNA extracted from eggs was amplified by PCR followed by DNA sequencing of targeted sequences of nuclear rRNA + internal transcribed spacers (ITS) and mitochondrial cox1. Phylogenetic analyses were performed. The DNA sequences of both nuclear rRNA + ITS and mitochondrial cox1 showed more than 98% and 99% identity to the known sequences respectively. Hymenolepis diminuta has been reported in various non-human primates with the highest prevalence of 38.5% in the white-headed capuchin monkey. The study presented here confirms that this tapeworm is capable of infecting various species of non-human primates with the first report of infections in AGM. Phylogenetic analyses of rRNA + ITS and mitochondrial cox1 demonstrated three separated clades I, II and III with the newly described AGM1 isolate belonging to the clade I. Whether these differences are at species level remains to be confirmed.

Lateral flow immunoassay with peptide-functionalized gold nanoparticles for rapid detection of protein tyrosine phosphatase 1B.

In this work, a lateral flow immunoassay (LFIA) with peptide functionalized gold nanoparticles (termed as biotin-ppeptide-AuNPs) has been developed for rapid, semi-quantitative detection of PTP1B activity without using any sophisticated equipment. In this method, the anti-phosphotyrosine (anti-pY) monoclonal antibody and streptavidin were used as test line and control line, respectively. The biotin-ppeptide-AuNPs contain 10% biotinylated peptide ligand carry a motif SDGHEpYIYVDP with pY (phosphotyrosine) and 90% pentapeptide (CALNN) ligand, which are used as PTP1B substrates and LFIA labelling probes. The experimental results demonstrate that the as-proposed LFIA with biotin-ppeptide-AuNPs exhibits a wide linear range (from 50 ng/mL to 10 µg/mL), a relatively low limit of detection (LOD, 44 ng/mL), and good specificity. In addition, the LFIA with biotin-ppeptide-AuNPs has been successfully used to evaluate activity levels of PTP1B in four cell lysates and the detection results exhibit a consistent trend with that of commercial kit.

A Novel Mitochondrial Genome Fragmentation Pattern in the Buffalo Louse Haematopinus tuberculatus (Psocodea: Haematopinidae).

Sucking lice are obligate ectoparasites of mammalian hosts, causing serious public health problems and economic losses worldwide. It is well known that sucking lice have fragmented mitochondrial (mt) genomes, but many remain undetermined. To better understand patterns of mt genome fragmentation in the sucking lice, we sequenced the mt genome of the buffalo louse Haematopinus tuberculatus using next-generation sequencing (NGS). The mt genome of H. tuberculatus has ten circular minichromosomes containing a total of 37 genes. Each minichromosome is 2.9-5.0 kb long and carries one to eight genes plus one large non-coding region. The number of mt minichromosomes of H. tuberculatus (ten) is different from those of congeneric species (horse louse H. asini, domestic pig louse H. suis and wild pig louse H. apri) and other sucking lice. Two events (gene translocation and merger of mt minichromosome) are observed in Haematopinus. Compared to other studies, our phylogeny generated from mt genome datasets showed a different topology, suggesting that inclusion of data other than mt genomes would be required to resolve phylogeny of sucking lice. To our knowledge, this is the first report of a ten mt minichromosomes genome in sucking lice, which opens a new outlook into unexplored mt genome fragmentation patterns in sucking lice.

Control and eradication of bovine trichomonosis in Wyoming, USA by testing and culling positive bulls.

Bovine trichomonosis is caused by Tritrichomonas foetus. Thirty-three US states have state rules on this disease and render it reportable due to potential huge economic losses to cattle industry. The various rules of different states generally mandate testing and culling T. foetus-positive bulls as well as prohibiting import of T. foetus-positive animals. Wyoming has enforced these rules for over 20 year beginning in 2000. From 2017 to 2019, 3 years in a row, not even one T. foetus-positive bull has been detected throughout the entire state among over ten thousand bulls tested annually. Wyoming is the first US state to achieve total control and eradication of bovine trichomonosis by testing and culling T. foetus-positive bulls.

Clinical isolates of Tritrichomonas foetus in bulls in Wyoming, South Dakota and Montana, USA.

BACKGROUND: Several Tritrichomonas species have been found in mammalian hosts. Among these trichomonads T. foetus is often found in the urogenital tract of cattle and the gastrointestinal tract of the domestic cat, resulting in sexually transmitted bovine trichomonosis and fecal-orally transmitted feline trichomonosis, respectively. The aims of the current study were to molecularly characterize clinical isolates of T. foetus in cattle populations in Wyoming, South Dakota, and Montana of the United States of America and to phylogenetically analyze Tritrichomonas species of mammalian hosts. RESULTS: DNA sequencing of rRNA genes showed over 99% identity of the newly described isolates to other bovine isolates. Further, T. foetus isolates of various mammalian hosts originated in different geographic regions worldwide were clustered into two well-defined clades by phylogenetic analysis of rRNA and cysteine protease 2 genes. Clade I consisted of isolates originated from cattle, pig, and human whereas clade II contained isolates of cat and dog. CONCLUSION: It is concluded that all mammalian Tritrichomonas spp. apparently belong to T. foetus. Analysis of more sequences is warranted to support this conclusion.

Two Tales of Cytauxzoon felis Infections in Domestic Cats.

Cytauxzoonosis is an emerging infectious disease that affects wild felids as well as the domestic cat; it is caused by the apicomplexan protozoan parasites belonging to the genus Cytauxzoon. Cytauxzoonfelis is the species of major concern, whose transmission occurs via the bite of an infected tick. Cytauxzoonosis of the domestic cat has historically been considered uniformly fatal, with a short course of illness, and most domestic cats die within 9 to 15 days postinfection. However, increasing evidence of domestic cats surviving C. felis infection suggests the existence of different strains with various levels of pathogenicity. Although wild felids are considered natural reservoirs for this parasite, a number of studies suggest that domestic cats that have survived nonlethal infections may serve as an additional reservoir. The current article comprehensively reviews the parasite and its life cycle, geographic distribution, genetic variability, and pathogenesis, as well as host immunology and the diagnosis, treatment, and prevention of infection in the domestic cat. This information should provide a basis for better understanding the parasite as well as the pathogenesis of the disease.

Parasites of small Indian mongoose, Herpestes auropunctatus, on St. Kitts, West Indies.

Herpestes auropunctatus, the small Indian mongoose, is an invasive omnivore introduced to the Caribbean, including the island of St. Kitts over 150 years ago. It has played a role in changing native fauna and can carry zoonotic pathogens of public health importance. The aim of the current study was to estimate the prevalence of parasites harbored by mongooses. In total, 87 mongooses trapped from April to July 2015 were examined for parasites using (1) hair plucks (N = 79), ear swabs (N = 79), and general coat and skin examination (N = 87) for mites, ticks, lice, and fleas; (2) dissection of the trachea, bronchi, and lungs for lungworms and flukes (N = 76); (3) a double centrifugation fecal flotation method for parasites of the gastrointestinal tract (N = 75); and (4) PCR of heart homogenates for Toxoplasma gondii (N = 60). The only ectoparasite seen was Ctenocephalides felis (79.3%; 69/87), with most mongooses having > 10 fleas (based on a subjective assessment) but insufficient numbers to result in signs of pruritus or anemia. On fecal flotation, coccidial oocysts were found with a prevalence of 69.3% (52/75). Neither T. gondii, lungworm, nor fluke infections were detected with the methods used. The high number of C. felis-infested mongooses and the infestation level of the individual mongooses suggest that they could serve as a reservoir for these potential vectors of pathogens. No evidence was found to support that mongooses are a component of T. gondii cycles on St. Kitts, although this finding needs to be confirmed with a larger sample size from other geographic locations.

Freedom from Tritrichomonas foetus infection in cattle in St. Kitts.

Trichomonosis is an endemic disease in cattle that are reared under extensive conditions and bred by natural mating. It causes profound economic losses to the producers by increasing calving interval, increasing embryo losses, and decreasing pregnancy rates. The aim of this study was to determine whether Tritrichomonas foetus infections were absent from cattle in St. Kitts. Using the modified hypergeometric method, preputial samples from bulls (n = 78) were tested using the InPouch™ culture for presence of T. foetus. Results highlighted an absence of trichomoniasis in bulls on St. Kitts with a 95% confidence.

Tritrichomonas foetus infection, a cause of chronic diarrhea in the domestic cat.

Tritrichomonas foetus is a very intriguing trichomonad protozoan with respect to its varied choice of residence in the different host species. It is an obligate parasite of the reproductive and the gastrointestinal tract of bovine and feline host respectively, leading to trichomonosis. Bovine trichomonosis is a sexually transmitted disease whereas feline trichomonosis is a disease with a purported fecal-oral route of spread. Further, the trichomonad is a commensal in the nasal passages, stomach, cecum and colon of swine host. Advances have been exponential in understanding the trichomonad biology and specifically feline trichomonosis since late 1990s and early 2000s when T. foetus was soundly determined to be a causative agent of chronic diarrhea in the domestic cat. It is a challenging task, even for a skilled investigator not to mention the busy clinical veterinarian, to keep up with the vast volume of information. Here we comprehensively reviewed the trichomonad biology, clinical manifestations, pathogenesis, host immunity, world map of distribution, risk factors, diagnosis and treatment. Risk factors associated with T. foetus-positive status in the domestic cat include young age, purebred, history of diarrhea, co-infections with other enteral pathogens. In addition, molecular similarity of bovine and feline isolates of T. foetus in DNA sequence was concisely discussed. The data presented serve as an information source for veterinarians, and investigators who are interested in biology of T. foetus and feline trichomonosis.

The plasma viral communities associate with clinical profiles in a large-scale haematological patients cohort.

BACKGROUND: Haematological patients exhibit immune system abnormalities that make them susceptible to viral infections. Understanding the relationship between the virome in the blood plasma of haematological patients and their clinical characteristic is crucial for disease management. We aimed to explore the presence of viral pathogens and identify close associations between viral infections and various clinical features. RESULTS: A total of 21 DNA viruses and 6 RNA viruses from 12 virus families were identified from 1383 patients. Patients with haematological diseases exhibited significantly higher diversity, prevalence, and co-detection rates of viral pathogens. During fever episodes, pathogen detection was notably higher, with Epstein-Barr virus (EBV) and Mucorales infections being the most probable culprits for fever symptoms in non-haematological patients. The detection rate of torque teno virus (TTV) significantly increases in haematological patients after transplantation and during primary lung infections. Additionally, TTV-positive patients demonstrate significantly higher absolute neutrophil counts, while C-reactive protein and procalcitonin levels are notably lower. Furthermore, TTV, cytomegalovirus, and parvovirus B19 (B19V) were found to be more prevalent in non-neutropenic patients, while non-viral pathogenic infections, such as Gram-negative bacteria and Mucorales, were more common in neutropenic patients. Pegivirus C (HPgV-C) infection often occurred post-transplantation, regardless of neutropenia. Additionally, some viruses such as TTV, B19V, EBV, and HPgV-C showed preferences for age and seasonal infections. CONCLUSIONS: Analysis of the plasma virome revealed the susceptibility of haematological patients to plasma viral infections at specific disease stages, along with the occurrence of mixed infections with non-viral pathogens. Close associations were observed between the plasma virome and various clinical characteristics, as well as clinical detection parameters. Understanding plasma virome aids in auxiliary clinical diagnosis and treatment, enabling early prevention to reduce infection rates in patients and improve their quality of life. Video Abstract.

Attenuation of Leishmania infantum chagasi metacyclic promastigotes by sterol depletion.

The infectious metacyclic promastigotes of Leishmania protozoa establish infection in a mammalian host after they are deposited into the dermis by a sand fly vector. Several Leishmania virulence factors promote infection, including the glycosylphosphatidylinositol membrane-anchored major surface protease (MSP). Metacyclic Leishmania infantum chagasi promastigotes were treated with methyl-beta-cyclodextrin (MßCD), a sterol-chelating reagent, causing a 3-fold reduction in total cellular sterols as well as enhancing MSP release without affecting parasite viability in vitro. MßCD-treated promastigotes were more susceptible to complement-mediated lysis than untreated controls and reduced the parasite load 3-fold when inoculated into BALB/c mice. Paradoxically, MßCD-treated promastigotes caused a higher initial in vitro infection rate in human or murine macrophages than untreated controls, although their intracellular multiplication was hindered upon infection establishment. There was a corresponding larger amount of covalently bound C3b than iC3b on the parasite surfaces of MßCD-treated promastigotes exposed to healthy human serum in vitro, as well as loss of MSP, a protease that enhances C3b cleavage to iC3b. Mass spectrometry showed that MßCD promotes the release of proteins into the extracellular medium, including both MSP and MSP-like protein (MLP), from virulent metacyclic promastigotes. These data support the hypothesis that plasma membrane sterols are important for the virulence of Leishmania protozoa at least in part through retention of membrane virulence proteins.

Human infections by the rat tapeworm Hymenolepis diminuta in China.

The rat tapeworm Hymenolepis diminuta is a parasite that usually uses rats as a definitive host. It also infects humans and non-human primates. Human infections have been reported in 80 countries worldwide, including China. Nevertheless, nearly all the literature on human infections in China by the rat tapeworm is in Chinese journals, which are very difficult to access by readers outside China. The main aim of the current manuscript was to systematically review human infections by the rat tapeworm in China for readers inside and outside the country. Chinese characters for H. diminuta were used to search several databases, including Google Scholar. In total, 511 infections were reported in 24 Chinese provinces/autonomous regions, which surpassed 320 in Costa Rica as the country with the highest number of infections. Furthermore, three nationwide surveys on parasitic infections in the past 3 decades revealed detailed prevalence of this parasite along with that of roundworm, whipworm, hookworm and pinworm in Chinese populations. These data contribute to better understanding of this greatly neglected zoonosis in the world's most populated country.

Correction: Matthew et al. A Loop-Mediated Isothermal Amplification (LAMP) Assay Specific to Trichomonas tenax Is Suitable for Use at Point-of-Care. Microorganisms 2022, 10, 594.

In the original publication [...].

Roles of Cysteine Proteases in Biology and Pathogenesis of Parasites.

Cysteine proteases, also known as thiol proteases, are a class of nucleophilic proteolytic enzymes containing cysteine residues in the enzymatic domain. These proteases generally play a pivotal role in many biological reactions, such as catabolic functions and protein processing, in all living organisms. They specifically take part in many important biological processes, especially in the absorption of nutrients, invasion, virulence, and immune evasion of parasitic organisms from unicellular protozoa to multicellular helminths. They can also be used as parasite diagnostic antigens and targets for gene modification and chemotherapy, as well as vaccine candidates, due to their species and even life-cycle stage specificity. This article highlights current knowledge on parasitic cysteine protease types, biological functions, and their applications in immunodiagnosis and chemotherapy.

One-pot synthesis of AuPt@FexOy nanoparticles with excellent peroxidase-like activity for development of ultrasensitive colorimetric lateral flow immunoassay of cardiac troponin I.

Detection of cardiac troponin I (cTnI) plays a critical role in diagnosing acute myocardial infarction (AMI). In this report, a new kind of spherical AuPt@FexOy core@shell nanoparticles (termed as AuPt@FexOy NPs) were one-pot synthesized by a redox interaction-engaged strategy (RIES) without the addition of any surfactants or reducing agents. The as-synthesized AuPt@FexOy NPs not only retain the plasmonic activity of gold nanoparticles (AuNPs), but also possess excellent catalytic activities of platinum nanoparticles (PtNPs) and FexOy nanoclusters. The features of AuPt@FexOy NPs enable greatly enhance the colorimetric detection sensitivity of lateral flow immunoassay (LFIA) through integrating AuPt@FexOy NPs labeling procedure and catalyzing oxidation of chromogenic substrate 3,3',5,5'-tetramethylbenzidine (TMB) signal amplification strategy. The as-developed colorimetric LFIA (termed as AuPt@FexOy-LFIA) exhibits the limit of detection (LOD) as 26.0 pg mL-1 cTnI under the TMB signal amplification mode. In particular, the detection results of cTnI in 40 clinical seral samples by AuPt@FexOy-LFIA are correlated well with those of cTnI in the same samples by commercial enzyme-linked immunosorbent assay (ELISA) detection kit (R2 = 0.97, slope = 1), demonstrating the highly reliable analytical performance and good application prospect of AuPt@FexOy-LFIA.

Trichomonas tenax: A Neglected Protozoan Infection in the Oral Cavities of Humans and Dogs-A Scoping Review.

Trichomonas tenax is a flagellated protozoan parasite found in the oral cavities of humans and animals and has been associated with periodontal disease, the most prevalent inflammatory disease affecting them all. Studies have shown that T. tenax can cause damage to mammalian cells and secretes virulent proteins, such as cysteine. It is presently considered zoonotic. Despite the few studies that have been done, the pathogenicity of this oral protozoan is still not fully understood. A database search was performed in July 2022 using PubMed and Google Scholar to retrieve data eligible for this study. PRISMA-ScR guidelines were followed to conduct this scoping review. A total of 321 articles were found with 87 included in this review after applying the exclusion criteria. Due to its increasing prevalence worldwide in both humans and dogs, detecting and elucidating the pathogenicity of this parasite is paramount for effective global control and prevention of periodontal disease. However, there is a paucity in the literature on this neglected zoonotic trichomonad, which is in large contrast to the closely related human pathogen T. vaginalis. Here, we comprehensively review the history, morphology and reproduction, host, prevalence, diagnosis, pathogenicity, control, and prevention of T. tenax. Hopefully, this article will call attention to both medical and veterinary professionals as well as epidemiologists on this most neglected and zoonotic protozoan. More epidemiological and clinical studies need to be conducted on T. tenax to gain a better understanding of its pathogenicity, to increase the chances of developing effective drugs to aid in the control of this oral parasite, and reduce the spread of periodontal disease worldwide.

Redundant targeting signals of the apicoplast-resident protein TgMnmA in Toxoplasma gondii involve trans-organellar function.

The apicoplast, which is the result of secondary endosymbiosis, is a distinctive subcellular organelle and a crucial therapeutic target for apicomplexan parasites. The majority of apicoplast-resident proteins are encoded by the nuclear genome and target the apicoplast via bipartite targeting signals consisting of a signal peptide and a transit peptide. The properties and functions of these peptides are poorly understood, which hinders the identification of apicoplast proteins and the study for plastid evolution. Here, the targeting signals of the recently discovered apicoplast tRNA thiouridylase TgMnmA of Toxoplasma gondii were analyzed. Our data using a reporter (the enhanced green fluorescent protein) fused with individual fragments containing various numbers of its N-terminal amino acids unequivocally revealed that the first 28 amino acids of TgMnmA functioned as a signal peptide for cellular secretion. The N-terminal 150 amino acids were sufficient to direct the fusion protein to the apicoplast, whereas its deletion caused the fusion protein to be localized to the mitochondrion. Our data further demonstrated that the apicoplast, rhoptry, and mitochondrion shared similar targeting signals, indicating that the apicoplast localization peptide was trans-organellar in function. In addition, the apicoplast localization peptide was important for the healthy proliferation of tachyzoites. In conclusion, the targeting signals of the nucleus-encoded apicoplast-targeted protein TgMnmA have been mapped out and the importance of this localization peptide has been elucidated in the current study.