Parasitic Nematodes Research Articles

Morphological and molecular identification of plant parasitic nematodes in wheat fields of Eastern Anatolian Region (Türkiye)

The aim of this study is to identify plant-parasitic nematodes molecularly and morphologically in wheat (Triticum spp.) fields in the Eastern Anatolia Region (Türkiye) between 2017-2019. For this purpose, a total of 258 soil samples were collected from 7 provinces (Erzincan, Elazığ, Erzurum, Iğdır, Kars, Malatya, and Sivas) in the Eastern Anatolia Region. Nematodes were morphologically identified using a light microscope at the genus (some of them species) level. DNA extraction was performed and PCR products were used to DNA sequencing and nucleotide analysis for 28S ribosomal DNA region by comparing the results with the database. According to the obtained data, a total of 20 genera and 7 species belonging to 2 orders and 9 families were identified: H. digonicus Perry, 1959; H. canadensis Waseem, 1961; H. vulgaris Yuen, 1964 (Nematoda: Hoplolaimidae); Ditylenchus myceliophagus Goodey, 1958 (Nematoda: Anguinidae); Amplimerlinius macrurus (Goodey, 1932) (Nematoda: Dolichodoridae); Scutylenchus quadrifer (Andrassy, 1954); (Nematoda: Dolichodoridae), and Pratylenchoides alkani Yüksel, 1977 (Nematoda: Pratylenchidae). H. vulgaris was identified at the species level using molecular techniques. The rates of presence of economically important plant parasitic nematodes were determined as 73%, 43%, 36%, 33% and 28% for Ditylenchus spp., Pratylenchus spp., Aphelenchus spp., Xiphinema spp. and Helicotylenchus spp. respectively. It is thought that the results obtained will help to plan nematode control methods in the region.

First report of the nematode‐trapping fungus Arthrobotrys thaumasia in Türkiye and its biocontrol potential against Meloidogyne incognita

AbstractRoot‐knot nematodes, particularly, Meloidogyne incognita, are among the most destructive endoparasitic nematodes, infecting a diverse range of plant hosts. Nematode‐trapping fungi are known for their potential application as biological control agents against plant parasitic nematodes. In the present study, the nematode‐trapping fungus Arthrobotrys thaumasia was isolated from the rhizosphere soil of tomato plants in the West Mediterranean Region of Türkiye. Two hundred and twenty‐three tomato plant rhizosphere soil samples yielded six nematode‐trapping fungal isolates, giving an occurrence frequency of 2.69%. Using morphology and molecular marker sequences (ITS and β‐tubulin loci), the species of the fungi was confirmed to be A. thaumasia. In vitro, A. thaumasia reduced second‐stage juveniles of M. incognita by 77.5% (isolate I‐Y4‐2) and 72.5% (isolate B‐G5‐1). This is the first report on the isolation and characterization of the nematode‐trapping fungus A. thaumasia from Türkiye. Two isolates of A. thaumasia (I‐Y4‐2 and B‐G5‐1) appear to be promising biological control agents that may be utilized for controlling M. incognita‐caused root‐knot diseases.

Inconsistency of in vitro exsheathment triggers for gastrointestinal nematode parasites of sheep, cattle and deer

Exsheathment is crucial in the transition from free-living to parasitic phase for most strongyle nematode species. A greater understanding of this process could help in developing new parasitic control methods. This study aimed to identify commonalities in response to exsheathment triggers (heat acclimation, CO2 and pH) in a wide range of species (Haemonchus contortus, Trichostrongylus spp., Cooperia spp., Oesophagostomum spp., Chabertia ovina, and members of the subfamily Ostertagiinae) from sheep, cattle and farmed deer. The initial expectation of similarity in pH requirements amongst species residing within the same organ was not supported, with unexpected pH preferences for exsheathment of Trichostrongylus axei, Trichostrongylus vitrinus, Trichostrongylus colubriformis and Cooperia oncophora. We also found differences between species in their response to temperature acclimation, with higher exsheathment in response to heat shock observed for H. contortus, Ostertagia ostertagi, T. axei, T. vitrinus and Oesophagostomum sikae. Furthermore, some species showed poor exsheathment under all experimental conditions, such as Cooperia curticei and the large intestinal nematodes C. ovina and Oesophagostomum venulosum. Interestingly, there were some significant differences in response depending on the host from which the parasites were derived. The host species significantly impacted on the exsheathment response for H. contortus, Teladorsagia circumcincta, T. vitrinus and T. colubriformis. Overall, the data showed variability between nematode species in their response to these in vitro exsheathment triggers, highlighting the complexity of finding a common set of conditions for all species in order to develop a control method based on triggering the exsheathment process prematurely.

Enteric tuft cells coordinate timely expulsion of the tapeworm Hymenolepis diminuta from the murine host by coordinating local but not systemic immunity.

Recognizing that enteric tuft cells can signal the presence of nematode parasites, we investigated whether tuft cells are required for the expulsion of the cestode, Hymenolepis diminuta, from the non-permissive mouse host, and in concomitant anti-helminthic responses. BALB/c and C57BL/6 mice infected with H. diminuta expelled the worms by 11 days post-infection (dpi) and displayed DCLK1+ (doublecortin-like kinase 1) tuft cell hyperplasia in the small intestine (not the colon) at 11 dpi. This tuft cell hyperplasia was dependent on IL-4Rα signalling and adaptive immunity, but not the microbiota. Expulsion of H. diminuta was slowed until at least 14 dpi, but not negated, in tuft cell-deficient Pou2f3-/- mice and was accompanied by delayed goblet cell hyperplasia and slowed small bowel transit. Worm antigen and mitogen evoked production of IL-4 and IL-10 by splenocytes from wild-type and Pou2f3-/- mice was not appreciably different, suggesting similar systemic immune reactivity to infection with H. diminuta. Wild-type and Pou2f3-/- mice infected with H. diminuta displayed partial protection against subsequent infection with the nematode Heligmosomoides bakeri. We speculate that, with respect to H. diminuta, enteric tuft cells are important for local immune events driving the rapidity of H. diminuta expulsion but are not critical in initiating or sustaining systemic Th2 responses that provide concomitant immunity against secondary infection with H. bakeri.

Plant Parasitic Nematodes Associated with Three African Indigenous Vegetables in Southwest Nigeria

Plant Parasitic Nematodes Associated with Three African Indigenous Vegetables in Southwest Nigeria

Inference of Essential Genes of the Parasite Haemonchus contortus via Machine Learning.

Over the years, comprehensive explorations of the model organisms Caenorhabditis elegans (elegant worm) and Drosophila melanogaster (vinegar fly) have contributed substantially to our understanding of complex biological processes and pathways in multicellular organisms generally. Extensive functional genomic-phenomic, genomic, transcriptomic, and proteomic data sets have enabled the discovery and characterisation of genes that are crucial for life, called 'essential genes'. Recently, we investigated the feasibility of inferring essential genes from such data sets using advanced bioinformatics and showed that a machine learning (ML)-based workflow could be used to extract or engineer features from DNA, RNA, protein, and/or cellular data/information to underpin the reliable prediction of essential genes both within and between C. elegans and D. melanogaster. As these are two distantly related species within the Ecdysozoa, we proposed that this ML approach would be particularly well suited for species that are within the same phylum or evolutionary clade. In the present study, we cross-predicted essential genes within the phylum Nematoda (evolutionary clade V)-between C. elegans and the pathogenic parasitic nematode H. contortus-and then ranked and prioritised H. contortus proteins encoded by these genes as intervention (e.g., drug) target candidates. Using strong, validated predictors, we inferred essential genes of H. contortus that are involved predominantly in crucial biological processes/pathways including ribosome biogenesis, translation, RNA binding/processing, and signalling and which are highly transcribed in the germline, somatic gonad precursors, sex myoblasts, vulva cell precursors, various nerve cells, glia, or hypodermis. The findings indicate that this in silico workflow provides a promising avenue to identify and prioritise panels/groups of drug target candidates in parasitic nematodes for experimental validation in vitro and/or in vivo.

Interaction between bacterial microbiota and nematode parasite communities in sheep’s gastrointestinal tract

Interaction between bacterial microbiota and nematode parasite communities in sheep’s gastrointestinal tract

Counting nematodes made easy: leveraging AI-powered automation for enhanced efficiency and precision.

Counting nematodes is a labor-intensive and time-consuming task, yet it is a pivotal step in various quantitative nematological studies; preparation of initial population densities and final population densities in pot, micro-plot and field trials for different objectives related to management including sampling and location of nematode infestation foci. Nematologists have long battled with the complexities of nematode counting, leading to several research initiatives aimed at automating this process. However, these research endeavors have primarily focused on identifying single-class objects within individual images. To enhance the practicality of this technology, there's a pressing need for an algorithm that cannot only detect but also classify multiple classes of objects concurrently. This study endeavors to tackle this challenge by developing a user-friendly Graphical User Interface (GUI) that comprises multiple deep learning algorithms, allowing simultaneous recognition and categorization of nematode eggs and second stage juveniles of Meloidogyne spp. In total of 650 images for eggs and 1339 images for juveniles were generated using two distinct imaging systems, resulting in 8655 eggs and 4742 Meloidogyne juveniles annotated using bounding box and segmentation, respectively. The deep-learning models were developed by leveraging the Convolutional Neural Networks (CNNs) machine learning architecture known as YOLOv8x. Our results showed that the models correctly identified eggs as eggs and Meloidogyne juveniles as Meloidogyne juveniles in 94% and 93% of instances, respectively. The model demonstrated higher than 0.70 coefficient correlation between model predictions and observations on unseen images. Our study has showcased the potential utility of these models in practical applications for the future. The GUI is made freely available to the public through the author's GitHub repository (https://github.com/bresilla/nematode_counting). While this study currently focuses on one genus, there are plans to expand the GUI's capabilities to include other economically significant genera of plant parasitic nematodes. Achieving these objectives, including enhancing the models' accuracy on different imaging systems, may necessitate collaboration among multiple nematology teams and laboratories, rather than being the work of a single entity. With the increasing interest among nematologists in harnessing machine learning, the authors are confident in the potential development of a universal automated nematode counting system accessible to all. This paper aims to serve as a framework and catalyst for initiating global collaboration toward this important goal.

Lure and starve: Host root exudates to suppress field populations of cyst nematodes

Plant parasitic nematodes are a burden to global food security, accounting for substantial yield losses in agricultural production worldwide. Increasing concerns over the environment and health issues have led to diminishing control options at the disposal of agricultural producers. Lure and starve, a strategy that is based on stimulating hatching of second stage juveniles (J2) of some cyst forming plant parasitic nematodes in the absence of their host plant seems a promising approach to maintain sustainability in crop production. Here we examined in-situ hatching of encysted eggs of the potato cyst nematode Globodera pallida and the carrot cyst nematode Heterodera carotae following exogenous applications of host root exudates in repeated field experiments during autumn and spring seasons. Population densities (viable J2 g−1 of soil) were assessed prior and post-application of root exudates. Results showed hatching induction of up to 83 % for G. pallida and 54 % for H. carotae in field plots drenched with their respective host root exudates. Spring season experiments were characterised by limited soil moisture and did not reduce nematode population densities for G. pallida. The potential for host root exudates in suppressing field population of cyst nematode in lure and starve strategies looks promising for both cyst nematode species investigated.

Molecular Characterization of Sweet Potato Plant Parasitic Nematodes in Different Agroecological Zones of Kirinyaga County, Kenya

Plant parasitic nematodes (PPNs) are widely distributed in all agroecological zones (AEZs) within Kenya. Plant parasitic nematodes are important pests of many cultivated crops and they exert a detrimental influence on a wide range of vascular plants, leading to significant crop losses by reducing both quantity and quality of the yield. However, due to inadequate taxonomic descriptions and a low number of diagnostic features, the morphological diagnosis of many species remains a challenge. The objective of this study was to characterize PPNs associated with sweet potatoes in different agroecological zones of Kirinyaga County, Kenya, using 18S rRNA gene sequencing. Seventy-seven soil samples from sweet potato tubers rhizosphere were collected from different agroecological zones and nematodes were extracted using Baermann’s technique. Thirteen nematode isolates were obtained and only seven could be identified morphologically to their genus level based on their distinct phenotypes. Seven isolates that could not be identified through the microscope were advanced to molecular sequencing. The nematode DNA were extracted and the PCR amplification and sequencing of 18S rRNA gene carried out. The study identified six PPN species, including Mylonchulus hawaiiensis, Aporcelaimellus nigeriensis, Rotylenchulus reniformis, Rotylenchulus borealis, Aporcella femina, Heterodera dunensis, and a predatory nematode (Dorylaimus aff. stagnalis) as part of the soil biota. This study showed significant distribution of plant parasitic nematodes across the agroecological zones. Thus, the occurrence of a nematode species complex in sweet potato farms requires the development of specific and appropriate sustainable control strategies.

A Phase 1, Randomized, Double-Blind, Placebo-Controlled, Single Ascending Dose Trial of AWZ1066S, an Anti-Wolbachia Candidate Macrofilaricide.

AWZ1066S has been developed as a potential treatment for the neglected tropical diseases lymphatic filariasis and onchocerciasis. AWZ1066S targets the Wolbachia bacterial endosymbiont present in the causative nematode parasites. This phase 1, first-in-human study aimed to assess the safety and pharmacokinetics of AWZ1066S in healthy human participants. In a randomized double-blind, placebo-controlled, single ascending dose study, healthy adults received a single oral dose of AWZ1066S (or placebo) and were followed up for 10 days. The planned single doses of AWZ1066S ranged from 100 to 1600mg, and each dose was administered to a cohort of 8 participants (6 AWZ1066S and 2 placebo). In total 30 people participated, 18 (60%) female, median age 30.0 years (minimum 20, maximum 61). The cohorts administered 100, 200, 300, and 400mg of AWZ1066S progressed unremarkably. After single 700-mg doses all 4 participants developed symptoms of acute gastritis and transient increases in liver enzymes. The severity of these adverse events ranged from mild to severe, with 1 participant needing hospital admission. Pharmacokinetic analysis indicated that AWZ1066S is rapidly absorbed with predictable pharmacokinetics. In conclusion, safety concerns prevented this study from reaching the human exposures needed for AWZ1066S to be clinically effective against lymphatic filariasis and onchocerciasis.

A soybean cyst nematode suppresses microbial plant symbionts using a lipochitooligosaccharide-hydrolysing enzyme.

Cyst nematodes are the most damaging species of plant-parasitic nematodes. They antagonize the colonization of beneficial microbial symbionts that are important for nutrient acquisition of plants. The molecular mechanism of the antagonism, however, remains elusive. Here, through biochemical combined with structural analysis, we reveal that Heterodera glycines, the most notorious soybean cyst nematode, suppresses symbiosis by secreting an enzyme named HgCht2 to hydrolyse the key symbiotic signalling molecules, lipochitooligosaccharides (LCOs). We solved the three-dimensional structures of apo HgCht2, as well as its chitooligosaccharide-bound and LCO-bound forms. These structures elucidated the substrate binding and hydrolysing mechanism of the enzyme. We designed an HgCht2 inhibitor, 1516b, which successfully suppresses the antagonism of cyst nematodes towards nitrogen-fixing rhizobia and phosphorus-absorbing arbuscular mycorrhizal symbioses. As HgCht2 is phylogenetically conserved across all cyst nematodes, our study revealed a molecular mechanism by which parasitic cyst nematodes antagonize the establishment of microbial symbiosis and provided a small-molecule solution.

Chromosome-level genome assembly of the cereal cyst nematode Heterodera flipjevi

As an economically important plant parasitic nematode (PPN), Heterodera filipjevi causes great damage on wheat, and now it was widely recorded in many countries. While multiple genomes of PPNs have been published, high-quality genome assembly and annotation on H. filipjevi have yet to be performed. This study presents a chromosome-scale genome assembly and annotation for H. filipjevi, utilizing a combination of Illumina short-read, PacBio long-read, and Hi-C sequencing technologies. The genome consists of 9 pseudo-chromosomes that contain 134.19 Mb of sequence, with a scaffold N50 length of 11.88 Mb. In total, 10,036 genes were annotated, representing 75.20% of the total predicted protein-coding genes. Our study provides the first chromosome-scale genome for H. filipjevi, which is also the inaugural high-quality genome of cereal cyst nematodes (CCNs). It provides a valuable genomic resource for further biological research and pest management of cereal cyst nematodes disease.

Different metazoan parasites, different transcriptomic responses, with new insights on parasitic castration by digenetic trematodes in the schistosome vector snail Biomphalaria glabrata

BackgroundGastropods of the genus Biomphalaria (Family Planorbidae) are exploited as vectors by Schistosoma mansoni, the most common causative agent of human intestinal schistosomiasis. Using improved genomic resources, overviews of how Biomphalaria responds to S. mansoni and other metazoan parasites can provide unique insights into the reproductive, immune, and other systems of invertebrate hosts, and their responses to parasite challenges.ResultsUsing Illumina-based RNA-Seq, we compared the responses of iM line B. glabrata at 2, 8, and 40 days post-infection (dpi) to single infections with S. mansoni, Echinostoma paraensei (both digenetic trematodes) or Daubaylia potomaca (a nematode parasite of planorbid snails). Responses were compared to unexposed time-matched control snails. We observed: (1) each parasite provoked a distinctive response with a predominance of down-regulated snail genes at all time points following exposure to either trematode, and of up-regulated genes at 8 and especially 40dpi following nematode exposure; (2) At 2 and 8dpi with either trematode, several snail genes associated with gametogenesis (particularly spermatogenesis) were down-regulated. Regarding the phenomenon of trematode-mediated parasitic castration in molluscs, we define for the first time a complement of host genes that are targeted, as early as 2dpi when trematode larvae are still small; (3) Differential gene expression of snails with trematode infection at 40dpi, when snails were shedding cercariae, was unexpectedly modest and revealed down-regulation of genes involved in the production of egg mass proteins and peptide processing; and (4) surprisingly, D. potomaca provoked up-regulation at 40dpi of many of the reproduction-related snail genes noted to be down-regulated at 2 and 8dpi following trematode infection. Happening at a time when B. glabrata began to succumb to D. potomaca, we hypothesize this response represents an unexpected form of fecundity compensation. We also document expression patterns for other Biomphalaria gene families, including fibrinogen domain-containing proteins (FReDs), C-type lectins, G-protein coupled receptors, biomphalysins, and protease and protease inhibitors.ConclusionsOur study is relevant in identifying several genes involved in reproduction that are targeted by parasites in the vector snail B. glabrata and that might be amenable to manipulation to minimize their ability to serve as vectors of schistosomes.

Biocontrol agent amendments shape the soybean rhizosphere in a cyst nematode (Heterodera glycines) conducive soil over a two-year field trial

Plant-associated beneficial microorganisms play a vital role in promoting plant health, fitness, and disease suppression, leading to improved plant growth and protection against specific plant parasites. Microbial amendments may reduce nematode parasite populations and ensure plant yield, yet their long-term impact on the native plant microbiome under field conditions is not well understood. In this study, we evaluated the effectiveness of biocontrol products against soybean cyst nematode (SCN) using two application methods over a two-year period under field conditions. Pochonia chlamydosporia PC10 (Rizotec), Bacillus methylotrophicus UFPEDA 20 (Onix), and Trichoderma koningiopsis GF 362 were applied either through seed inoculation or in-furrow treatment at planting. The treatments effectively reduced the nematode population, with T. koningiopsis showing significant deviations from the control and leading to a notable increase in yield. No difference in the Shannon diversity index was detected for the alpha-diversity of root-associated 16S, ITS2, and 18S communities. The dominant phyla were Proteobacteria, Acidobacteria, Actinobacteria for 16S; Ascomycota, Basidiomycota, Mortierellomycota for ITS2; and Ascomycota, and Cercozoa for 18S. In the 16S community, T. koningiopsis and B. methylotrophicus caused a 25 % increase in the relative abundance of Sphingomonas spp. compared to the control. Additionally, the relative abundance of Mortierella spp. significantly increased by 50 % in both the T. koningiopsis and B. methylotrophicus treatments compared to the control. Both treatments also led to a significant reduction in Fusarium spp. by 37.5 % and 31.5 %, respectively. Co-occurrence network analysis revealed that communities were disrupted after the first application, but grew more intricate and cohesive after the second year of biocontrol product amendment. T. koningiopsis and B. methylotrophicus demonstrated potential in reducing nematode and Fusarium populations, leading to increased yield production. However, under high nematode pressure, relying solely on biocontrol measures does not guarantee a reduction in SCN population or yield improvement. Manipulating the microbial community to reduce harmful organisms and promote biocontrol-related species may offer long-term benefits.

Grass-Endophyte Interactions and Their Associated Alkaloids as a Potential Management Strategy for Plant Parasitic Nematodes.

Claviceptaceous endophytic fungi in the genus Epichloë mostly form a symbiotic relationship with cool-season grasses. Epichloë spp. are capable of producing bioactive alkaloids such as peramines, lolines, ergot alkaloids, and indole-diterpenes, which protect the host plant from herbivory by animals, insects, and nematodes. The host also benefits from enhanced tolerance to abiotic stresses, such as salt, drought, waterlogging, cold, heavy metals, and low nitrogen stress. The bioactive alkaloids produced can have both direct and indirect effects towards plant parasitic nematodes. Direct interaction with nematodes' motile stages can cause paralysis (nematostatic effect) or death (nematicidal effect). Indirectly, the metabolites may induce host immunity which inhibits feeding and subsequent nematode development. This review highlights the different mechanisms through which this interaction and the metabolites produced have been explored in the suppression of plant parasitic nematodes and also how the specific interactions between different grass genotypes and endophyte strains result in variable suppression of different nematode species. An understanding of the different grass-endophyte interactions and their successes and failures in suppressing various nematode species is essential to enable the proper selection of grass-endophyte combinations to identify the alkaloids produced, concentrations required, and determine which nematodes are sensitive to which specific alkaloids.

Development of a novel rodent model for dog heartworm microfilaremia using the severe-combined immunodeficiency mouse

Dirofilaria immitis is a mosquito-borne parasitic nematode that causes fatal heartworm disease in canids. The microfilariae are essential for research, including drug screening and mosquito-parasite interactions. However, no reliable methods for maintaining microfilaria long-term are currently available. Therefore, we used severe combined immunodeficiency (SCID) mice to develop a reliable method for maintaining D. immitis microfilaria. SCID mice were injected intravenously with microfilariae isolated from a D. immitis-infected dog. Microfilariae were detected in blood collected from the tail vein 218 days post-inoculation (dpi) and via cardiac puncture 296 dpi. Microfilariae maintained in and extracted from SCID mice showed infectivity and matured into third-stage larvae (L3s) in the vector mosquito Aedes aegypti. L3s can develop into the fourth stage larvae in vitro. Microfilariae from SCID mice respond normally to ivermectin in vitro. The microfilariae in SCID mice displayed periodicity in the peripheral circulation. The SCID mouse model aided in the separation of microfilariae from cryopreserved specimens. The use of SCID mice enabled the isolation and sustained cultivation of microfilariae from clinical samples. These findings highlight the usefulness of the SCID mouse model for studying D. immitis microfilaremia in canine heartworm research.

Seasonal population dynamics and impact of abiotic factors on plant parasitic nematodes under temperate conditions of Kashmir, India

Plant parasitic nematodes (PPNs) pose a significant threat to the successful cultivation of grapevines globally. Abiotic parameters such as temperature, moisture and pH critically influence nematode population growth. Ecological investigation was conducted on three grapevine cultivars (Sahibi, Hussaini and Thompson) in Kashmir, India, from January 2021 to December 2022. The population dynamics of the identified PPNs around the grapevine rhizosphere were studied in relation to various factors. Nematode abundance peaked during the warm months of the summer season and dipped during winter. Soil and air temperatures positively correlated with the abundance of nematodes; however, soil moisture and rainfall showed a negative correlation. This study underscores the critical role of environmental factors on the population dynamics of plant parasitic nematodes, with temperature being an important influential factor. Effective nematode management in grapevine cultivation requires a thorough understanding of these dynamics to mitigate the impact of these parasites.

Morphological and molecular data on acuariid nematodes in European great cormorants (Phalacrocorax carbo sinensis) and pygmy cormorants (Microcarbo pygmaeus)

The family Acuariidae is a speciose group of parasitic nematodes, infecting mostly birds as definitive hosts. This study focused on the characterization of two species of acuariids, collected in two different species of piscivorous birds, the European great cormorant Phalacrocorax carbo sinensis from Italy, and the pygmy cormorant Microcarbo pygmaeus from Israel. Parasites were analyzed using light and scanning electron microscopy and by amplification and sequencing of the 28S rDNA. The results of morphological and molecular analyses showed that Ph. carbo sinensis was infected by the acuariid Syncuaria squamata (12 females) and Cosmocephalus obvelatus (1 female), whereas M. pygmaeus was infected by C. obvelatus (2 males, 12 females). The present results provide new data on the distribution of acuariid parasites of piscivorous birds, the first report of Acuariidae in Israel, and the first molecular data on S. squamata and C. obvelatus, which will be useful in future epidemiological and phylogenetic studies of these widely distributed, but less molecularly studied parasites.

Evaluation of diagnostic techniques for early detection of heartworm in experimentally infected dogs: identification of Dirofilaria immitis-derived microRNA in the initial 28 weeks post-inoculation

BackgroundDirofilaria immitis, commonly known as heartworm (HW), is a parasitic nematode transmitted by various mosquito species, leading to heartworm disease (HWD) in dogs. Diagnosis of HW typically involves antigen or microfilariae detection, or visualization of adult worms through imaging or post mortem examination. Polymerase chain reaction (PCR) and micro RNA (miRNA) detection have been explored for HW diagnosis.MethodsThree dogs, previously experimentally infected with HW, underwent blood sampling every 4 weeks for 7 months. Samples were assessed for antigen presence after heat treatment, PCR amplification, and microfilaria examination using Giemsa-stained thick smears. Additionally, whole blood aliquots underwent miRNA deep sequencing and bioinformatic analysis.ResultsHeartworm antigen was detectable after heat treatment at 20 weeks post-inoculation and via PCR at 24 weeks, with microfilariae observed in peripheral blood smears at 28 weeks. However, deep miRNA sequencing revealed that the miRNA candidate sequences are not consistently expressed before 28 weeks of infection.ConclusionsWhile ancillary molecular methods such as PCR and miRNA sequencing may be less effective than antigen detection for detecting immature larval stages in an early stage of infection, our experimental findings demonstrate that circulating miRNAs can still be detected in 28 weeks post-infection.Graphical