Dog Hookworm Uncinaria stenocephala Still Vulnerable to Key Dewormers, Italian Field Study Finds

In the rolling rural landscapes of Basilicata in southern Italy, a quiet parasitological drama has been unfolding in the dogs that roam villages and farmyards. A team of veterinary parasitologists led by Renata Fagundes-Moreira and Domenico Otranto of the University of Bari Aldo Moro, together with Jan Šlapeta of the University of Sydney, has now delivered one of the most detailed field assessments to date of Uncinaria stenocephala, a canine hookworm that is increasingly reported across Europe yet remains poorly understood under natural conditions. Their study, published in Parasites & Vectors, combines clinical pathology, parasitological monitoring, molecular diagnostics, and drug-susceptibility testing in naturally infected dogs, offering a rare integrated picture of how this parasite behaves outside the laboratory.

The research focused on a population of domestic dogs from an endemic rural area, sampling between July and October 2025. Twenty naturally Uncinaria-infected dogs and two puppies were enrolled, alongside three uninfected dogs that served as comparators. Fecal and blood samples were collected repeatedly, with egg shedding quantified on days minus 14, minus 7, and day 0 using the Mini-FLOTAC technique, a flotation-based counting method prized for its accuracy in quantifying parasite eggs per gram of feces. At day 0, the dogs were treated with one of three common anthelmintics: fenbendazole, moxidectin, or milbemycin oxime. Efficacy was then measured with the fecal egg count reduction test, or FECRT, the gold-standard field method that compares egg counts before and after treatment, in this case at day 14 post-treatment.

One of the most striking findings concerns the sheer volatility of egg shedding before any drug was administered. Egg counts ranged from zero to 3,105 eggs per gram across the pre-treatment sampling window, with sharp swings between consecutive sampling days. The mean egg count rose from 122 eggs per gram at day minus 14 to 647.2 at day minus 7, only to fall back to 162.2 at day 0. Such dramatic fluctuation within a matter of days carries a sober practical message: a single fecal sample can badly misrepresent the true infection status of a dog. A negative or low count on one day might mask a substantial infection that would be revealed a week later, which is precisely why the authors argue for multi-sample fecal diagnostics in both clinical practice and surveillance programs.

When it came to treatment, the results split cleanly. Fenbendazole, a member of the benzimidazole class that disrupts the parasite’s microtubule machinery by binding to β-tubulin, achieved a 100 percent fecal egg count reduction. Moxidectin, a milbemycin-class macrocyclic lactone that interferes with glutamate-gated chloride channels in the worm’s nervous system, came close behind at 99.3 percent efficacy. Milbemycin oxime, however, managed only 76.8 percent reduction, a figure that falls short of the thresholds typically used to declare acceptable efficacy and that raises questions about how this drug performs against U. stenocephala under field conditions. Importantly, the researchers did not stop at counting eggs; they probed the molecular and phenotypic basis of drug susceptibility.

To assess benzimidazole resistance, the team performed an in vitro egg hatch assay, exposing hookworm eggs to increasing concentrations of a benzimidazole and measuring the concentration that inhibits hatching in half of the eggs, the IC50. Two IC50 values were recorded, 0.03 with a 95 percent confidence interval of 0.03 to 0.04, and 0.09 with a confidence interval of 0.08 to 0.10, both consistent with phenotypic susceptibility to the drug class. In other words, despite the modest performance of milbemycin oxime in the live animals, the benzimidazole pathway itself showed no functional signs of resistance in this population.

Molecular screening reinforced that conclusion. Using amplicon deep sequencing of the internal transcribed spacer 2 region of the ribosomal RNA gene, the researchers confirmed that U. stenocephala was the sole hookworm species present in the samples, ruling out hidden co-infections with other hookworms such as Ancylostoma species that could confound efficacy calculations. They then sequenced the β-tubulin isotype-1 gene, the canonical molecular target whose mutations are the best-validated markers of benzimidazole resistance in parasitic nematodes. Across codons 134, 167, 198, and 200, the positions most frequently implicated in resistance, no canonical resistance-associated single nucleotide polymorphisms were detected, either before or after treatment. The genetic evidence and the phenotypic egg hatch data thus tell a consistent story: this southern Italian population of U. stenocephala remains fully susceptible to benzimidazoles.

The clinical pathology component of the study adds a reassuring but nuanced dimension. Hematological and biochemical values in the infected dogs were mostly within reference limits, a profile consistent with low-intensity parasitism rather than clinically significant disease. The most frequent abnormalities were mild eosinophilia, the elevated eosinophil count typical of the immune response to helminths; slight hypoalbuminemia, a modest drop in the blood protein albumin that hookworms can induce through intestinal blood loss and protein leakage; and increased β-globulin fractions, reflecting the antibody-driven immune activation that chronic parasitic exposure produces. None of these findings amounted to the severe anemia or profound protein loss that can accompany heavy hookworm burdens, suggesting that in these naturally exposed dogs, U. stenocephala was more of a subclinical passenger than a dangerous pathogen.

That subclinical profile is itself part of the reason the parasite has flown under the radar. U. stenocephala, sometimes called the northern hookworm, is better adapted to cooler climates than its tropical and subtropical relatives, and its infections tend to produce lower egg outputs and milder clinical signs. Yet the parasite is being reported with increasing frequency in Europe, and without baseline data on its shedding dynamics, drug susceptibility, and clinical impact, veterinarians have had little ground to stand on when interpreting diagnostic results or choosing treatments. This study supplies exactly that baseline, and its message about the unreliability of single-sample diagnosis is likely to resonate well beyond the study region.

The work also carries broader significance for the global problem of anthelmintic resistance. In canine hookworms, particularly Ancylostoma caninum in North America, widespread resistance to multiple drug classes has become a serious clinical challenge, making the search for populations where susceptibility persists all the more valuable. By documenting, with molecular and phenotypic rigor, a European U. stenocephala population that still responds fully to fenbendazole and moxidectin, the researchers have established a reference point against which future changes can be measured. Continued surveillance of anthelmintic efficacy, they emphasize, is essential, because resistance mutations can spread rapidly once they arise, and the modest efficacy of milbemycin oxime observed here warrants closer attention in future studies.

For dog owners and veterinarians, the practical takeaways are concrete. Repeated fecal examinations over consecutive weeks give a far more accurate picture of hookworm infection than a single test, given the day-to-day variability in egg shedding documented here. Fenbendazole and moxidectin remain dependable choices against U. stenocephala in this region, while milbemycin oxime’s performance suggests it should not be assumed fully effective without verification. And as this underestimated hookworm continues its apparent expansion across Europe, the integrated approach demonstrated in Basilicata, pairing egg counts with clinical chemistry, deep amplicon sequencing, and egg hatch assays, offers a template for monitoring not just this species, but the wider and shifting landscape of canine parasitism in a changing world.

Subject of Research: Anthelmintic susceptibility and clinical impact of the canine hookworm Uncinaria stenocephala in naturally infected dogs

Article Title: Clinical pathology, parasitological evaluation, and anthelmintic resistance assessment in dogs naturally infected with Uncinaria stenocephala

Article References: Fagundes-Moreira, R., Bezerra-Santos, M. A., Šlapeta, J., & Otranto, D. (2026). Clinical pathology, parasitological evaluation, and anthelmintic resistance assessment in dogs naturally infected with Uncinaria stenocephala. Parasites & Vectors. https://doi.org/10.1186/s13071-026-07688-4

Image Credits: AI Generated

DOI: 10.1186/s13071-026-07688-4

Keywords: Uncinaria stenocephala, hookworms, dogs, anthelmintic resistance, benzimidazole, fenbendazole, moxidectin, milbemycin oxime, fecal egg count reduction test, egg hatch assay, β-tubulin, Parasites & Vectors