Strongyloides stercoralis, commonly known as the human threadworm, is a skin-penetrating gastrointestinal parasitic nematode that infects hundreds of millions of people worldwide. Like other Strongyloides species, S. stercoralis is capable of cycling through a single free-living generation. Although S. stercoralis and the free-living nematode Caenorhabditis elegans are evolutionarily distant, the free-living adults of S. stercoralis are similar enough in size and morphology to C. elegans adults that techniques for generating transgenics and knockouts in C. elegans have been successfully adapted for use in S. stercoralis. High-quality genomic and transcriptomic data are also available for S. stercoralis. Thus, one can use a burgeoning array of functional genomic tools in S. stercoralis to probe questions about parasitic nematode development, physiology, and behavior. Knowledge gained from S. stercoralis will inform studies of other parasitic nematodes such as hookworms that are not yet amenable to genetic manipulation. This review describes the basic anatomy of S. stercoralis.

The skin-penetrating gastrointestinal parasitic nematode Strongyloides stercoralis causes strongyloidiasis, which is a neglected tropical disease that is associated with severe chronic illness and fatalities. Unlike other human-infective nematodes, S. stercoralis cycles through a single free-living generation and thus serves as a genetically tractable model organism for understanding the mechanisms that enable parasitism. Techniques such as CRISPR/Cas9-mediated mutagenesis and transgenesis are now routinely performed in S. stercoralis by introducing exogenous DNA into free-living adults and then screening their F1 progeny for transgenic or mutant larvae. However, transgenesis in S. stercoralis has been severely hindered by the inability to establish stable transgenic lines that can be propagated for multiple generations through a host; to date, studies of transgenic S. stercoralis have been limited to heterogeneous populations of transgenic F1 larvae. Here, we develop an efficient pipeline for the generation of stable transgenic lines in S. stercoralis. We also show that this approach can be used to efficiently generate stable transgenic lines in the rat-infective nematode Strongyloides ratti. The ability to generate stable transgenic lines circumvents the limitations of working with heterogeneous F1 populations, such as variable transgene expression and the inability to generate transgenics of all life stages. Our transgenesis approach will enable novel lines of inquiry into parasite biology, such as transgene-based comparisons between free-living and parasitic generations.

The Caenorhabditis elegans egg hatching methodology is a valuable tool for assessing the anthelmintic activity of drugs and compounds and evaluating anthelmintic drug efficacy. Isolated eggs from gravid adults are exposed to different concentrations of selected drugs and the percentage of egg hatching is determined with respect to the control condition. The assay allows the construction of concentration-response curves and determination of EC50 or EC90 values for egg hatching inhibition. Also, it allows measurements of inhibition as a function of time of exposure. This approach addresses the urgent need for new anthelmintics, as resistance to current treatments poses a significant challenge in parasitic nematode infection. This resistance not only affects humans but also animals and plants, causing significant economic losses in livestock farming and agriculture. By using the free-living nematode C. elegans as a parasitic model organism, researchers can efficiently screen for potential treatments and assess drug combinations for synergistic effects. Importantly, this assay offers a cost-effective and accessible alternative to traditional methods, eliminating the need for specialized infrastructure, hosts, and trained animal maintenance personnel. Additionally, the methodology closely mimics natural conditions, providing insights into egg development and potential therapeutic targets. This method allows for evaluating the direct negative impact of drugs on egg hatching, which correlates with long-term anthelmintic effects, offering advantages in preventing or reducing the transmission and spread of worm infections by eggs. Overall, this approach represents a significant advancement for anthelmintic discovery, offering both practical applications and avenues for further scientific research. •The C. elegans egg hatching assay is a robust and effective method for assessing the anthelmintic potential of various drugs and compounds, allowing the generation of concentration-response curves.•By leveraging the free-living nematode C. elegans as a parasitic model organism, this method facilitates efficient screening of potential treatments and evaluation of drug combinations.•The method addresses the urgent need for new anthelmintics, offering a cost-effective and accessible alternative to traditional approaches.

Skin-penetrating nematodes infect nearly one billion people worldwide. The developmentally arrested infective larvae (iL3s) seek out hosts, invade hosts via skin penetration, and resume development inside the host in a process called activation. Activated infective larvae (iL3as) traverse the host body, ending up as parasitic adults in the small intestine. Skin-penetrating nematodes respond to many chemosensory cues, but how chemosensation contributes to host seeking, intra-host development, and intra-host navigation - three crucial steps of the parasite-host interaction - remains poorly understood. Here, we investigate the role of carbon dioxide (CO2) in promoting parasite-host interactions in the human-infective threadworm Strongyloides stercoralis. We show that S. stercoralis exhibits life-stage-specific preferences for CO2: iL3s are repelled, non-infective larvae and adults are neutral, and iL3as are attracted. CO2 repulsion in iL3s may prime them for host seeking by stimulating dispersal from host feces, while CO2 attraction in iL3as may direct worms toward high-CO2 areas of the body such as the lungs and intestine. We also identify sensory neurons that detect CO2; these neurons are depolarized by CO2 in iL3s and iL3as. In addition, we demonstrate that the receptor guanylate cyclase Ss-GCY-9 is expressed specifically in CO2-sensing neurons and is required for CO2-evoked behavior. Ss-GCY-9 also promotes activation, indicating that a single receptor can mediate both behavioral and physiological responses to CO2. Our results illuminate chemosensory mechanisms that shape the interaction between parasitic nematodes and their human hosts and may aid in the design of novel anthelmintics that target the CO2-sensing pathway.

Morphological data are used to describe a new nematode species, Heterocheilus floridensis sp. n. (Heterocheilidae) from the digestive tract of the Florida manatee Trichechus manatus latirostris (Harlan) (Trichechidae, Sirenia) from Florida, USA. Examination by light and scanning electron microscopy revealed that the new species differs from the related Heterocheilus tunicatus Diesing, 1839 mainly by having dentigerous ridges on the inner surface of the lips, a median unpaired papilla located anterior to the cloaca, and a considerably larger body size. Sequence data for subunits I and II of mitochondrial cytochrome oxidase gene, 18S small subunit and 28S ribosomal RNA genes were provided for molecular characterisation of the new species. However, the current unavailability of homologous sequence data for congeneric specimens precluded a molecular assessment of the morphological species hypothesis, and ascaridoid phylogenetic hypotheses could not be advanced. Specimens of Heterocheilus sp. collected from the Antillean manatee Trichechus manatus manatus Linnaeus in Puerto Rico, on loan from the US National Museum of Natural History, were morphologically consistent with the new species, so apparently all congeneric nematodes reported from both subspecies of the West Indian manatee Trichechus manatus Linnaeus and previously identified as H. tunicatus belong rather to H. floridensis sp. n. Heterocheilus hagenbecki (Khalil et Vogelsang, 1932) Sprent 1980 is here considered to be a species inquirenda. A key to valid species of Heterocheilus Diesing, 1839 is provided.

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The raccoon roundworm (Baylisascaris procyonis), a gastrointestinal nematode of the raccoon (Procyon lotor), may cause a severe form of larva migrans in humans, which can lead to death or permanent neurological damage. Although roundworms were inadvertently introduced to Europe alongside their raccoon hosts, the parasite is not present in every raccoon population. It is important to understand the geographic distribution of B. procyonis, as early and rapid treatment can prevent severe pathologies in humans. We present evidence for the roundworm spreading into a naive raccoon population through natural dispersal of infected raccoons. We sampled 181 raccoons from Saxony-Anhalt, a German federal state containing contact zones of different raccoon populations, two of which were previously free of the parasite. We screened the raccoons for roundworms and used microsatellite-based assignment tests to determine the genetic origin of the raccoons and their parasites. We detected roundworms in 16 of 45 raccoons sampled in a previously roundworm-free area in the northern part of the state. The largest proportion of the genetic ancestry (≥ 0.5) of the 16 raccoon hosts was assigned to the previously naive raccoon population. Conversely, the genetic ancestry of almost all the roundworms was assigned to the nearest roundworm population in the southern part of the state. Infected raccoons have, therefore, spread to the north of the state, where they interbred with and infected local raccoons. It seems likely that the roundworms will continue to spread. Health authorities should consider continuous surveillance programmes of naive populations and raise public awareness.

BACKGROUND: Ribosome biogenesis is the process of assembling ribosome complexes that regulate cell proliferation and differentiation with potential regulatory effects on development. Many factors regulate ribosome biological processes. Nin one binding protein (Nob1) has received widespread attention as key genes regulating ribosome biogenesis-the 3\' end of the 20S rRNA is cleaved by Nob1 at cleavage site D to form 18S rRNA, generating translationally capable 40S subunit. As a ribosome biogenesis factor, Nob1 may regulate the development of organisms, but almost nothing is known about the function of Nob1 for any parasitic nematode. We explored the functional role of NOBP-1 (the homologous gene of Nob1) encoding gene from a parasitic nematode-Strongyloides stercoralis.
METHODS: The full-length cDNA, gDNA and promoter region of Ss-nobp-1 was identified using protein BLAST in WormBase ParaSite according to the Caenorhabditis elegans NOBP-1 sequence to analyze the gene structure. RNA sequencing (RNA-seq) data in wormbase were retrieved and analyzed to assess the transcript abundance of Ss-nobp-1 in seven developmental stages of S. stercoralis. The standard method for gonadal microinjection of constructs was carried out to determine the anatomic expression patterns of Ss-nobp-1. The interaction between Ss-NOBP-1 and partner of NOBP-1 (Ss-PNO-1) was assessed by yeast two-hybridization and bimolecular fluorescence complementarity (BiFC) experiments.
RESULTS: The NOBP-1 encoding gene Ss-nopb-1 from the zoonotic parasite S. stercoralis has been isolated and characterized. The genomic DNA representing Ss-nobp-1 includes a 1599-bp coding region and encodes a protein comprising 403 amino acids (aa), which contains conserved PIN domain and zinc ribbon domain. RNA-seq analysis revealed that Ss-nobp-1 transcripts are present throughout the seven developmental stages in S. stercoralis and have higher transcription levels in iL3, L3 and P Female. Ss-nobp-1 is expressed mainly in the intestine of transgenic S. stercoralis larvae, and there is a direct interaction between Ss-NOBP-1 and Ss-PNO-1.
CONCLUSIONS: Collectively, Ss-NOBP-1 has a potential role in embryo formation and the infective process, and findings from this study provide a sound foundation for investigating its function during the development of parasitic nematode.

Cockroach microbiome studies generally focus on pest cockroach species belonging to the Blattidae and Ectobiidae families. There are no reports characterizing the gut microbiome of non-pest cockroach species Blaberus discoidalis (family Blaberidae), which is commonly used as a food source for insectivorous animals. We discovered the parasitic nematode Leidynema appendiculata in the B. discoidalis hindgut during initial work characterizing the gut microbiome of this organism. To determine the proportion of the B. discoidalis colony that was colonized by L. appendiculata, 28 S rDNA was amplified using two Methods: endpoint polymerase chain reaction (PCR) and loop-mediated isothermal amplification (LAMP). B. discoidalis colonies were raised on three diet types (control, high fibre, and high fat and salt) for 21 days before dissection. Each individual was sexed during dissection to identify potential sex-based effects of colonization. Data collected were analysed to determine if diet and sex impacted parasite colonization patterns. LAMP detected a higher proportion of parasite positive samples when compared to endpoint PCR. No sex- or diet-based differences in L. appendiculata colonization were found. This study adds to the limited existing knowledge of the B. discoidalis gut microbiome.

Based on light microscopical and scanning electron microscopical (SEM) examinations, two North American species of Spinitectus Fourment, 1884, S. acipenseri Choudhury & Dick, 1992 and S. micracanthus Christian, 1972 (Nematoda, Cystidicolidae) are redescribed from museum voucher specimens (S. acipenseri) and those newly collected from centrarchid and some other fishes in the Upper San Marcos River in Texas and the Santee River in South Carolina, USA. The first use of SEM to study S. acipenseri, a parasite of lake sturgeon Acipenser fulvescens Rafinesque (Acipenseridae) in Canada, made it possible to describe dorsal and ventral lips, amphids and sublabia, and the presence of a dorsal barb on the right spicule, which was confirmed to be the most characteristic feature of this species. The SEM study of S. micracanthus, a parasite mainly of centrarchids, enabled us to correctly determine the location of the excretory pore in relation to rings of cuticular spines in the male, and to describe the exact structure of the tip of the male tail, sublabia, phasmids and the presence of a median ventral protuberance on the male tail. Some taxonomic problems of North American species of Spinitectus are discussed. Filaria serrata Linton, 1901 is considered a junior synonym of S. oviflagellis Fourment, 1884. To date, there are 13 valid species of Spinitectus parasitising fishes in North America. Keys to species of Spinitectus-like nematodes from fishes in North American waters are provided.
UNASSIGNED: Redescriptions de Spinitectus acipenseri et S. micracanthus (Nematoda, Cystidicolidae), et notes sur la taxonomie des nématodes de type Spinitectus parasitant les poissons nord-américains.
UNASSIGNED: Deux espèces nord-américaines de Spinitectus Fourment, 1884, S. acipenseri Choudhury & Dick, 1992 et S. micracanthus Christian, 1972 (Nematoda, Cystidicolidae) sont redécrites à partir de spécimens de musée et d’autres nouvellement collectés aux USA dans les rivières Upper San Marcos (Texas) and Santee (Caroline du Sud), avec l’aide de la microscopie optique et électronique à balayage (MEB). L’utilisation pour la première fois du MEB pour étudier S. acipenseri, un parasite de l’esturgeon Acipenser fulvescens Rafinesque (Acipenseridae) au Canada, a permis de décrire les lèvres dorsales and ventrales, les amphides et sublabia, ainsi que la barbe dorsale sur le spicule droit que nous confirmons être le critère le plus caractéristique de cette espèce. L’étude au MEB de S. micracanthus, un parasite principalement de centrarchides, nous a permis de déterminer correctement la place du pore excréteur en relation avec les couronnes d’épines cuticulaires chez le mâle, ainsi que de décrire la structure exacte de l’extrémité caudale du mâle, des sublabia, des phasmides et la présence d’une protubérance médiane et ventrale sur la queue du mâle. Des problèmes taxonomiques sur les espèces nord-américaines de Spinitectus sont discutés. Filaria serrata Linton, 1901 est considéré synonyme plus récent de S. oviflagellis Fourment, 1884. À ce jour, il y a 13 espèces valides de Spinitectus parasites de poissons en Amérique du Nord. Des clés des espèces des nématodes de type Spinitectus provenant des poissons des eaux nord-américaines sont fournies.