Spirocerca lupi, the dog oesophageal nematode, causes a potentially fatal disease in domestic dogs, and is currently clinically diagnosed by coproscopy and oesophagoscopy. To date, a single molecular ...method, a semi-nested PCR, targeting the cox1 gene, has been developed to aid in the diagnosis of spirocercosis. The present study describes three novel high-resolution melt (HRM) quantitative PCR (qPCR) assays targeting fragments of the ITS1, 18S and cytb loci of S. lupi. The performance of these molecular assays in feces was compared to fecal flotation and to the previously described cox1 gene semi-nested PCR in 18 fecal samples from dogs with clinical oesophageal spirocercosis diagnosed by oesophagoscopy.
The HRM qPCR for ITS1 and 18S were both able to detect 0.2 S. lupi eggs per gram (epg), while the HRM qPCR for the cytb and the semi-nested PCR for the cox1 detected 6 epg and 526 epg, respectively. Spirocerca lupi was detected in 61.1%, 44.4%, 27.8%, 11.1% and 5.6% of the fecal samples of dogs diagnosed with spirocercosis by using the ITS1 and 18S HRM qPCR assays, fecal flotation, cytb HRM qPCR and cox1 semi-nested PCR, respectively. All dogs positive by fecal flotation were also positive by ITS1 and 18S HRM qPCRs. Quantification of S. lupi eggs was successfully achieved in the HRM qPCRs and compared to the fecal flotation with no significant difference in the calculated concentrations between the HRM qPCRs that detected the 18S and ITS1 loci and the fecal flotation. The HRM qPCR for the 18S cross-amplified DNA from Toxocara canis and Toxascaris leonina. In contrast, the HRM qPCR for ITS1 did not cross-amplify DNA from other canine gastrointestinal parasites.
This study presents two new molecular assays with significantly increased sensitivity for confirming and quantifying fecal S. lupi eggs. Of these, the HRM qPCR for ITS1 showed the best performance in terms of the limit of detection and absence of cross-amplification with other parasites. These assays will be useful in detecting infection and for follow-up during therapy.
Spirocerca lupi is a parasitic nematode of canids that can lead to a severe and potentially fatal disease. Recently, a new species, Spirocerca vulpis, was described from red foxes in Europe, ...suggesting a high genetic diversity of the Spirocerca spp. infecting canids. The genetic variation and phylogenetic relationships of S. lupi collected from naturally-infected domestic dogs from Australia, Hungary, Israel, Italy, India and South Africa, and S. vulpis from red foxes from Bosnia and Herzegovina, Italy and Spain, was studied using mitochondrial and rDNA markers.
A high intra-individual variation was found in the first internal transcribed spacer (ITS1) locus in all Spirocerca spp., ranging between 0.37-2.84%, with up to six haplotypes per specimen. In addition, a combination of phylogenetic and haplotype analyses revealed a large variability between S. lupi specimens collected from different geographical locations using the ITS1 (0.37-9.33%) and the cytochrome c oxidase subunit 1 (cox1) gene (1.42-6.74%). This genetic diversity led to the identification of two S. lupi genotypes circulating among dogs (PTP support > 0.829), including genotype 1 found in S. lupi from Australia, India, Israel and South Africa, and genotype 2 represented by specimens from Hungary and Italy. These genotypes presented pairwise nucleotide distances of 0.14%, 8.06% and 6.48 ± 0.28% in the small rDNA subunit (18S), ITS1 and cox1 loci, respectively. Additionally, Nei's genetic distance in the ITS1 showed a further subdivision of genotype 1 worms into 1A (Israel and South Africa) and 1B (Australia and India). A morphological analysis of the anterior and posterior extremities of genotype 1 and genotype 2 worms using scanning electron microscopy did not show any differences between the specimens, contrary to the morphological differences between S. lupi and S. vulpis.
These findings demonstrate the high genetic variability among Spirocerca spp. from different geographical locations, thereby expanding our understanding of the epidemiology, evolution and phylogenetic variability within the genus.
Willow (Salix spp.) trees, found worldwide, contain secondary metabolites that are valuable as dietary supplements for animal feed and as antiparasitic compounds. We quantified secondary metabolites ...(phenolics, flavonoids, and salicylic acid) in ethanolic extracts from leaves and branches of three Salix acmophylla Boiss. genotypes and investigated their potential to inhibit Eimeria sp. sporulation, a major concern in ruminants. The total phenolic content of willow leaves and branches was similar in two of three different genotypes. The total flavonoid content of the branches was significantly higher than that of leaves of the same genotype; however, the salicylic acid content was significantly higher in leaves than in branches. Importantly, all extracts exhibited significant inhibition of Eimeria sporulation, where over 70% inhibition was obtained at concentrations as low as 750 mgL−1. The sporulation inhibition by branch or leaf extracts exceeded 80% for leaves and 90% for branches at concentrations above 1250 mgL−1. The study highlights the potential of using Salix extracts as bioactive compounds for biological control of coccidiosis in ruminants. We conclude that all parts and all investigated genotypes of S. acmophylla can provide secondary metabolites that act as a coccidiostat to treat Eimeria in goats.
Gastrointestinal parasites are one of the main restrictions to small ruminant production. Their pathological importance is primarily related to the major production losses, in quantity or quality, ...induced by the direct action of worms. Control of these parasites is based exclusively on the frequent use of anthelmintic drugs. However, the resistance to anthelmintics in worm populations after commercialisation of chemical drugs is now widespread. Therefore, there is a need to find new natural resources to ensure sustainable and effective treatment and control of these parasites. The aim of this study was to evaluate the anthelmintic activity, as minimum inhibitory concentration (IC
mg/mL), of different plant extracts using larval exsheathment inhibition assay using a two-species but steady population of parasitic nematodes (ca. 20% Teladorsagia circumcinta and 80% Trichostrongylus colubriformis).
The study showed that the ethanolic extracts of 22 out of the 48 plant extracts, obtained from 46 plant species, have an inhibitory effect >50% (at concentrations of 100 mg/mL) on the third stage larvae (L3) of the nematodes exhibited the strongest inhibition activity (94%) with IC
of 0.02 mg/mL, where other members of the Rhamnaceae family have shown to possess strong anthelmintic activity (70-89%).
Plant extracts are potential rich resources of anthelmintics to combat helminthic diseases. Our results suggest that extracts from Rhamnus elaternus, Epilobium hirsutum, Leucaena leucocephala and Rhamnus palaestinus have promising anthelmintic activity, with potential applications in animal therapeutics and feed.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Cryptosporidium parvum is an apicomplexan parasite that infects humans and ruminants. C. parvum isolated from cattle in northeastern Turkey and in Israel was genotyped using multiple polymorphic ...genetic markers, and the two populations were compared to assess the effect of cattle husbandry on the parasite's population structure. Dairy herds in Israel are permanently confined with essentially no opportunity for direct herd-to-herd transmission, whereas in Turkey there are more opportunities for transmission as animals range over wider areas and are frequently traded. A total of 76 C. parvum isolates from 16 locations in Israel and seven farms in the Kars region in northeastern Turkey were genotyped using 16 mini- and microsatellite markers. Significantly, in both countries distinct multilocus genotypes confined to individual farms were detected. The number of genotypes per farm was higher and mixed isolates were more frequent in Turkey than in Israel. As expected from the presence of distinct multilocus genotypes in individual herds, linkage disequilibrium among loci was detected in Israel. Together, these observations show that genetically distinct populations of C. parvum can emerge within a group of hosts in a relatively short time. This may explain the frequent detection of host-specific genotypes with unknown taxonomic status in surface water and the existence of geographically restricted C. hominis genotypes in humans.
The parasitic nematode Spirocerca lupi (Spirurida: Thelaziidae), the canine esophageal worm, is the causative agent of spirocercosis, a disease causing morbidity and mortality in dogs. Spirocerca ...lupi has a complex life cycle, involving an obligatory coleopteran intermediate host (vector), an optional paratenic host, and a definitive canid host. The diagnosis of spirocercosis is challenging, especially in the early disease stages, when adult worms and clinical signs are absent. Thus, alternative approaches are needed to promote early diagnosis. The interaction between nematodes and their bacterial symbionts has recently become a focus of novel treatment regimens for other helminthic diseases.
Using 16S rDNA-based molecular methods, here we found a novel bacterial symbiont in S. lupi that is closely related to Comamonas species (Brukholderiales: Comamonadaceae) of the beta-proteobacteria. Its DNA was detected in eggs, larvae and adult stages of S. lupi. Using fluorescent in situ hybridization technique, we localized Comamonas sp. to the gut epithelial cells of the nematode larvae. Specific PCR enabled the detection of this symbiont's DNA in blood obtained from dogs diagnosed with spirocercosis.
The discovery of a new Comamonas sp. in S. lupi increase the complexity of the interactions among the organisms involved in this system, and may open innovative approaches for diagnosis and control of spirocercosis in dogs.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
•Spirocerca lupi is a nematode of dogs that utilizes dung beetles as its intermediate host.•Understanding the triggers of egg development may help in preventing infection.•Embryonation of S. lupi ...eggs was enhanced by potassium dichromate.•Hatching was enhanced by trypsin, pH levels, temperature of 26°C, 20% CO2 and mechanical pressure.•Eggs hatched in the buccal head parts and the midgut of Onthophagus sellatus beetles.
Spirocerca lupi is a parasitic nematode which causes spirocercosis, a severe disease of dogs. Its life cycle involves dung beetles as intermediate hosts and canids as definitive hosts. The effect of different physical conditions and chemical factors on the embryonation and hatching of S. lupi eggs were investigated in this study in order to understand the triggers for progression in the early development of this parasite.
Exposure to potassium dichromate significantly enhanced the embryonation of eggs compared to formaldehyde and controls (p<0.0001), reaching the maximum embryonation level of 83% within 2days of incubation. Hatching of eggs was significantly (p<0.05) enhanced in the presence of 2.5% trypsin, pH 6.0 and 8.0, a temperature of 26°C, 20% CO2 and mechanical force by stirring with 3-mm beads. Dissection of Onthophagus sellatus beetles 8h post-feeding with eggs showed that 13% of the ingested eggs hatched in the buccal cavity and the midgut. Finally, the pH range of the beetle’s gut was 6.0-6.2 compared to 7.2±0.4 in dog feces suggesting that this pH change may induce hatching in the beetle.
These findings contribute to the understanding of the early steps in the life cycle of S. lupi and may be used in the future to block the development of S. lupi and prevent dog infection and disease.
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•Trichinella britovi and Trichinella spiralis were genetically identified in Israel.•Examined isolates were analyzed using multiplex and extended PCR assays.•Out of total 58 samples, ...four mixed infections were confirmed using clone analysis.
Trichinellosis is a worldwide disease caused by nematode worms of the genus Trichinella, frequently diagnosed in Israel. However, the identity of the Israeli isolates have not been studied. Here we describe the molecular characterization of 58 isolates collected from jackals (Canis aureus), wild boar (Sus scrofa), foxes (Vulpes vulpes) and a wolf (Canis lupus) in central and northern Israel. Isolates were analyzed using the multiplex PCR analysis encompassing expansion segment V (ESV) and internal sequence 1 (ITS-1) markers, which identified 52 of the 58 samples. Out of the six unidentified samples, four were successfully identified using extended PCR assays for ESV and ITS-1, developed in this study. Our analysis identified 44 isolates as T. britovi, 8 as T. spiralis, four mixed infections, and two isolates were not identified. Clonal analysis of the ITS-1 sequences from six isolates confirmed the initial identification of four mixed infections. These results show that the prevalent species in Israel are T. britovi and T. spiralis, with nearly 7% (4 of 58) incidence of mixed infection.
Willow from Goats Haj-Zaroubi, Manal; Mattar, Nariman; Awabdeh, Sami ...
Agriculture (Basel),
05/2024, Letnik:
14, Številka:
5
Journal Article
Recenzirano
Willow (Salix spp.) trees, found worldwide, contain secondary metabolites that are valuable as dietary supplements for animal feed and as antiparasitic compounds. We quantified secondary metabolites ...(phenolics, flavonoids, and salicylic acid) in ethanolic extracts from leaves and branches of three Salix acmophylla Boiss. genotypes and investigated their potential to inhibit Eimeria sp. sporulation, a major concern in ruminants. The total phenolic content of willow leaves and branches was similar in two of three different genotypes. The total flavonoid content of the branches was significantly higher than that of leaves of the same genotype; however, the salicylic acid content was significantly higher in leaves than in branches. Importantly, all extracts exhibited significant inhibition of Eimeria sporulation, where over 70% inhibition was obtained at concentrations as low as 750 mgLsup.−1. The sporulation inhibition by branch or leaf extracts exceeded 80% for leaves and 90% for branches at concentrations above 1250 mgLsup.−1. The study highlights the potential of using Salix extracts as bioactive compounds for biological control of coccidiosis in ruminants. We conclude that all parts and all investigated genotypes of S. acmophylla can provide secondary metabolites that act as a coccidiostat to treat Eimeria in goats.
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