Silver and gold nanoparticles as a novel approach to fight Sarcoptic mange in rabbits.

Various kinds of pets have been known to contract the ectoparasite Sarcoptes scabiei. Current acaricides are becoming less effective because of the resistance developed by the mite besides their adverse effects on the general activity and reproductive performance of domestic pets. For this reason, the present study aims to discover a novel and safe approach using silver and gold nanoparticles to fight Sarcoptic mange in rabbits as well as to explain their mechanism of action. 15 pet rabbits with clinical signs of Sarcoptic mange that were confirmed by the microscopic examination were used in our study. All rabbits used in this study were assessed positive for the presence of different developing stages of S. scabiei. Three groups of rabbits (n = 5) were used as follows: group (1) didn't receive any treatment, and group (2 and 3) was treated with either AgNPs or GNPs, respectively. Both nanoparticles were applied daily on the affected skin areas via a dressing and injected subcutaneously once a week for 2 weeks at a dose of 0.5 mg/kg bwt. Our results revealed that all rabbits were severely infested and took a mean score = 3. The skin lesions in rabbits that didn't receive any treatments progressed extensively and took a mean score = of 4. On the other hand, all nanoparticle-treated groups displayed marked improvement in the skin lesion and took an average score of 0-1. All NPs treated groups showed remarkable improvement in the microscopic pictures along with mild iNOS, TNF-α, and Cox-2 expression. Both nanoparticles could downregulate the m-RNA levels of IL-6 and IFγ and upregulate IL-10 and TGF-1ß genes to promote skin healing. Dressing rabbits with both NPs didn't affect either liver and kidney biomarkers or serum Ig levels indicating their safety. Our residual analysis detected AgNPs in the liver of rabbits but did not detect any residues of GNPs in such organs. We recommend using GNPs as an alternative acaricide to fight rabbit mange.

In vitro acaricidal activity of several natural products against ibex-derived Sarcoptes scabiei.

In this study we analysed the effect of the temperature, diverse strains of Bacillus thuringiensis, Lysinibacillus sphaericus and nanoformulations with essential plant oils (EONP) on the survival of Sarcoptes scabiei mites derived from naturally-infested Iberian ibex (Capra pyrenaica). In general, mites maintained at 12ºC survived more than those maintained at 35ºC (40.7 hr and 31.2 hr, respectively). Mites with no treatment survived 27.6 h on average. Mites treated with B. thuringiensis serovar. konkukian and geranium EONP showed significant reduction in their survival. Despite the fact that these agents seem to be promising candidates for controlling sarcoptic mange in the field, further research is still needed to get stable, efficient and eco-friendly acaricides.

The effects of spatially-constrained treatment regions upon a model of wombat mange.

The use of therapeutic agents is a critical option to manage wildlife disease, but their implementation is usually spatially constrained. We seek to expand knowledge around the effectiveness of management of environmentally-transmitted Sarcoptes scabiei on a host population, by studying the effect of a spatially constrained treatment regime on disease dynamics in the bare-nosed wombat Vombatus ursinus. A host population of wombats is modelled using a system of non-linear partial differential equations, a spatially-varying treatment regime is applied to this population and the dynamics are studied over a period of several years. Treatment could result in mite decrease within the treatment region, extending to a lesser degree outside, with significant increases in wombat population. However, the benefits of targeted treatment regions within an environment are shown to be dependent on conditions at the start (endemic vs. disease free), as well as on the locations of these special regions (centre of the wombat population or against a geographical boundary). This research demonstrates the importance of understanding the state of the environment and populations before treatment commences, the effects of re-treatment schedules within the treatment region, and the transient large-scale changes in mite numbers that can be brought about by sudden changes to the environment. It also demonstrates that, with good knowledge of the host-pathogen dynamics and the spatial terrain, it is possible to achieve substantial reduction in mite numbers within the target region, with increases in wombat numbers throughout the environment.

Assessment of the in vitro acaricidal activity of Bravecto® (fluralaner) and a proposed orange oil-based formulation vehicle for the treatment of Sarcoptes scabiei.

BACKGROUND: Sarcoptic mange is a serious animal welfare concern in bare-nosed wombats (Vombatus ursinus). Fluralaner (Bravecto®) is a novel acaricide that has recently been utilised for treating mange in wombats. The topical 'spot-on' formulation of fluralaner can limit treatment delivery options in situ, but dilution to a volume for 'pour-on' delivery is one practicable solution. This study investigated the in vitro acaricidal activity of Bravecto, a proposed essential oil-based diluent (Orange Power®), and two of its active constituents, limonene and citral, against Sarcoptes scabiei. METHODS: Sarcoptes scabiei were sourced from experimentally infested pigs. In vitro assays were performed to determine the lethal concentration (LC50) and survival time of the mites when exposed to varying concentrations of the test solutions. RESULTS: All compounds were highly effective at killing mites in vitro. The LC50 values of Bravecto, Orange Power, limonene and citral at 1 h were 14.61 mg/ml, 4.50%, 26.53% and 0.76%, respectively. The median survival times of mites exposed to undiluted Bravecto, Orange Power and their combination were 15, 5 and 10 min, respectively. A pilot survival assay of mites collected from a mange-affected wombat showed survival times of < 10 min when exposed to Bravecto and Orange Power and 20 min when exposed to moxidectin. CONCLUSIONS: These results confirm the acaricidal properties of Bravecto, demonstrate acaricidal properties of Orange Power and support the potential suitability of Orange Power and its active constituents as a diluent for Bravecto. As well as killing mites via direct exposure, Orange Power could potentially enhance the topical delivery of Bravecto to wombats by increasing drug penetration in hyperkeratotic crusts. Further research evaluating the physiochemical properties and modes of action of Orange Power and its constituents as a formulation vehicle would be of value.

A systematic review of immunosuppressive risk factors and comorbidities associated with the development of crusted scabies.

OBJECTIVES: Crusted scabies (CS, Norwegian scabies) is a severe form of scabies, characterized by hyper-infestation of Sarcoptes scabiei mites. CS is commonly associated with immunosuppression but is also reported in overtly immunocompetent individuals. We reviewed immunosuppressive risk factors and comorbidities associated with CS. METHODS: The National Library of Medicine (PubMed) database was reviewed for patient case reports of CS from January 1998 to July 2023. Two authors screened records for eligibility, extracted data, and one critically appraised the quality of the studies. SYSTEMATIC REVIEW REGISTRATION: PROSPERO CRD42023466126. RESULTS: A total of 436 records were identified, of which 204 were included for systematic review. From these, 683 CS patients were included. CS impacted both genders equally. Adults (21-59 years) were more commonly affected (45.5%) compared to children (0-20 years, 21%). Corticosteroid use was the most prevalent immunosuppressive risk factor identified (27.7% of all cases). About 10.2% of reports were associated with HIV/AIDS, and 8.5% with HTLV-1 infection. 10.5% of patients were overtly immunocompetent with no known risk factors. Overall, 41 (6.0%) died, many subsequent to secondary bacteremia. CONCLUSION: This study represents the first systematic review undertaken on immunosuppressive risk factors associated with CS. This provides insights into trends of immunosuppression and mechanisms of CS development.

Distribution of Scabies Infestations in Stray Dogs in Bursa Province.

Objective: Scabies infestation is a major parasitic disease affecting both human and animal health worldwide. This study aimed to determine the distribution of scabies infestation in stray dogs in Bursa province. Methods: The study material was obtained from stray dogs suspected of mange kept in a municipal shelter in Bursa between June 2020 and February 2022. Samples from stray dogs were examined in a laboratory, and dogs with scabies were determined. Results: During the study, samples were collected from 205 (115 males, 90 females) scabies-suspected stray dogs, 58 (28.29%) of which had scabies. Demodex spp. were detected in 35 of the positive dogs (60.34%) (D. canis, D. injae), 19 (32.76%) Sarcoptes scabiei canis, 2 (3.44%) mixed infestation (Sarcoptes and Demodex), and 2 (3.44%) Otodectes cynotis agents. It was determined that 32 (55.17%) of 58 scabies-positive dogs were male and 26 (44.83%) were female. The association of infestations with age, season, and sex has not been found to be statistically meaningful. Conclusion: This study identified scabies agents and their prevalence rate in stray dogs of Bursa province, Türkiye.

Scabies: Immunopathogenesis and pathological changes.

Scabies is an itchy skin disease caused by the burrowing mite Sarcoptes scabiei. During their lifespan, the female mites invade the stratum corneum and create tunnels, in which they reside, move, feed, deposit fecal pellets, and lay eggs. Recently, scabies was included in the World Health Organization roadmap for neglected tropical diseases 2021-2030. This review attempts to summarize our knowledge about the mite's biology and the disease pathogenesis, pathological changes, and complications. Generally, the host-parasite interaction in scabies is highly complex and involves different mechanisms, some of which are yet largely unknown. Elucidation of the nature of such interaction as well as the underlying mechanisms could allow a better understanding of the mite's biology and the development of novel diagnostic and therapeutic options for scabies control programs. Moreover, identification of the molecular basis of such interaction could unveil novel targets for acaricidal agents and vaccines.

Escabiose: aspectos gerais e a importância no reconhecimento e tratamento precoces / Scabies: general aspects and the importance of early recognition and treatment

A escabiose é uma dermatose infecto parasitária mais prevalente no mundo, sobretudo nas regiões tropicais e em países de baixa renda. Surtos são comuns em locais de aglomeração como presídios, escolas e campos de refugiados, e tanto o atraso no diagnóstico como o tratamento inadequado são responsáveis pela propagação da doença. Este trabalho tem por objetivos destacar os principais aspectos da escabiose bem como as apresentações dermatológicas, a fim de auxiliar no diagnóstico e tratamentos precoces, tendo como foco o médico generalista. O estudo foi realizado no Departamento de Clínica de Dermatologia do Hospital das Clínicas da Faculdade de Medicina de São Paulo (HCFMUSP), através de uma revisão de literatura com acesso aos bancos de dados eletrônicos PubMed. A escabiose é causada pelo Sarcoptes scabiei, caracterizada pelo prurido intenso e por lesões cutâneas sugestivas e localizações típicas, mas que podem variar de acordo com a faixa etária ou estados de imunossupressão. É considerada pela Organização Mundial da Saúde uma doença tropical negligenciada, podendo causar grande impacto socioeconômico e, ainda que com menor frequência, levar a complicações, muitas vezes decorrentes de infecções bacterianas secundárias, sobretudo nas formas mais graves da doença (vistas principalmente em pacientes imunossuprimidos). Por esses motivos, o reconhecimento das principais formas de apresentação clínica e sintomas associados são importantes para que o diagnóstico seja estabelecido de forma breve, possibilitando a instituição correta do tratamento e, com isso, cessando o ciclo de transmissão do ácaro.

Resolution of Clinical Signs of Sarcoptic Mange in American Black Bears (Ursus americanus), in Ivermectin-Treated and Nontreated Individuals.

The parasitic mite Sarcoptes scabiei causes mange in nearly 150 species of mammals by burrowing under the skin, triggering hypersensitivity responses that can alter animals' behavior and result in extreme weight loss, secondary infections, and even death. Since the 1990s, sarcoptic mange has increased in incidence and geographic distribution in Pennsylvania black bear (Ursus americanus) populations, including expansion into other states. Recovery from mange in free-ranging wildlife has rarely been evaluated. Following the Pennsylvania Game Commission's standard operating procedures at the time of the study, treatment consisted of one subcutaneous injection of ivermectin. To evaluate black bear survival and recovery from mange, from 2018 to 2020 we fitted 61 bears, including 43 with mange, with GPS collars to track their movements and recovery. Bears were collared in triplicates according to sex and habitat, consisting of one bear without mange (healthy control), one scabietic bear treated with ivermectin when collared, and one untreated scabietic bear. Bears were reevaluated for signs of mange during annual den visits, if recaptured during the study period, and after mortality events. Disease status and recovery from mange was determined based on outward gross appearance and presence of S. scabiei mites from skin scrapes. Of the 36 scabietic bears with known recovery status, 81% fully recovered regardless of treatment, with 88% recovered with treatment and 74% recovered without treatment. All bears with no, low, or moderate mite burdens (<16 mites on skin scrapes) fully recovered from mange (n=20), and nearly half of bears with severe mite burden (≥16 mites) fully recovered (n=5, 42%). However, nonrecovered status did not indicate mortality, and mange-related mortality was infrequent. Most bears were able to recover from mange irrespective of treatment, potentially indicating a need for reevaluation of the mange wildlife management paradigm.

Molecular characterization of Sarcoptes scabiei causing severe mange in two Andean foxes (Lycalopex culpaeus) from Peru.

Cases of sarcoptic mange in the wildlife population have been increasing around the world in recent years. In this study, we report the first case of severe sarcoptic mange in two Andean foxes (Lycalopex culpaeus) and molecularly analyze the collected mites. The National Forestry and Wildlife Service of Lima, Peru, found an adult male Andean fox in the province of Callao in August 2022. The veterinarians decided to euthanize the fox due to the severity of the mange. In August 2023, an adult male Andean fox was found dead in the province of Huaral by veterinarians. Both foxes were sent to the veterinary school in Lima for necropsy. Skin samples from different body zones were digested in buffer lysis, and mites were detected in the tissue samples. A morphological diagnosis identified the mites as Sarcoptes scabiei. The mites from both foxes had the same nuclear (ITS2) and mitochondrial (cox1) genetic marker sequences as the S. scabiei sequences from dogs, vicunas, Andean foxes, and water buffalos recorded in GenBank. Unlike ITS2, phylogenetic analysis of S. scabiei cox1 showed host-related nucleotide sequence polymorphisms. Future molecular studies of S. scabiei from different hosts and localities will be necessary to better understand the transmission of this disease in Andean foxes.

Epizootiology and Pathology of Sarcoptic Mange in Two Species of Fox (Lycalopex spp.) in Human-Dominated Landscapes of Central Chile.

Sarcoptic mange caused by Sarcoptes scabiei can have catastrophic consequences for wildlife. We inspected 122 Andean foxes (Lycalopex culpaeus), collected by active (n=66) or passive (n=56) surveillance, and 28 South American gray foxes (Lycalopex griseus; all from passive surveillance) for mange in Chile (2015-19). In Andean foxes, gross lesions of mange were diagnosed in 24% of passively and 9% of actively collected foxes, although observed prevalences might be underestimated. Seroprevalence was 37 and 18%, respectively, indicating that some individuals recovered from infection or were developing the disease. No differences were found between age and sex groups. Comparing data from passive surveillance, occurrence of gross lesions was lower in gray foxes (5%). Body condition was significantly better in Andean foxes without lesions than in diseased foxes, which had significantly lower albumin concentrations than healthy individuals. Among the 12 foxes with gross lesions, four, six and two individuals were categorized as having type I, type II, and type III lesions, respectively, based on clinical severity. Histologic severity correlated with gross lesions and included irregular epidermal hyperplasia with hyperkeratosis, which was marked in type II and III infections. Conventional PCR targeting of the cox1 gene fragment revealed four nucleotide sequence types, showing 99-100% identity among them and between 99% and 100% identity with previously published sequences of S. scabiei. A significant association between the occurrence of mange in foxes and distance to the nearest house was found. We speculate that diseased foxes tended to approach human settlements, perhaps in search of food. Visual inspection of 211 rural dogs from the study area did not reveal gross mange lesions in any animal. Sarcoptic mange is enzootic in the Andean fox in the study area and should be considered in the management of the species.

Plantar keratoderma-like crusted scabies in an immunocompetent infant after topical steroids.

A healthy 6-month-old girl presented with plantar keratoderma-like lesions unresponsive to topical corticosteroids. Nocturnal pruritus in 13 relatives, presence of burrows on clinical exam, and the positive scabies preparation led to the diagnosis of crusted scabies. She was successfully treated with topical and oral scabicides. Crusted scabies is a severe form of Sarcoptes scabiei infection uncommon in immunocompetent subjects, in whom previous corticosteroid use may favor its occurrence.

Scabies in Spain: Trends in Public Interest and Ectoparasiticide Use. / Escabiosis en España: tendencias del interés público y consumo de ectoparasiticidas.

Scabies is an ectoparasitic dermatosis caused by the Sarcoptes scabiei var. hominis mite, which lives and reproduces in humans. Its incidence in Spain has increased in recent years. The aim of this study was to complement existing evidence on the increasing number of scabies cases in our country by analyzing changes in ectoparasiticide prescriptions and Internet searches for scabies infestations measured by Google Trends. We also examined correlations between these two variables. Our results show that public interest in scabies has increased in recent years and is positively and significantly correlated with an increasing use of ectoparasiticides. We believe that Google Trends should be considered as a complementary tool for monitoring real-time trends in scabies infestations in Spain.

Camera traps reveal a high prevalence of sarcoptic mange in red foxes from northern Spain.

The mite Sarcoptes scabiei affects numerous mammal species causing the sarcoptic mange, a widespread disease with relevance for wildlife conservation, welfare, and management. The red fox (Vulpes vulpes) could become infested by direct and indirect routes leading to external skin lesions potentially recognizable by devices such as camera traps (CTs). In the present study, 86 randomly placed CTs were used to investigate the apparent prevalence and severity of S. scabiei in a red fox population from northern Spain. Their potential environmental and population-related drivers were also assessed. A total of 341 independent encounters were examined to visually identify mange-compatible lesions. The apparent prevalence was 19.16% (confidence interval (CI) 95%: 15.08-23.80) of which 82.81% (CI95%: 71.33-91.10) were severe. Our results revealed that habitat attributes such as lower altitudes, higher coverage of water-linked habitats and woodland predominance, were significant predictors of the apparent risk of mange. The models also suggested that the apparent prevalence of mange was associated with poor body condition and elevated frequencies of spatial coincidence among fox encounters, which facilitates indirect transmission. Interestingly, we did not observe mange-compatible lesions in other sympatric wild species (>15,000 encounters examined). This could be explained by the mite's host specificity and the low probability that these other potential hosts use sites where transmission among foxes usually occurs, such as dens. This study illustrates how camera trapping can be used as an interesting tool for the surveillance of wildlife diseases, thus overcoming the logistic constraints derived from direct sampling and allowing the early detection and better management of pathogens in the riskiest areas.

[Translated article] Scabies in Spain: Trends in Public Interest and Ectoparasiticide Use.

Scabies is an ectoparasitic dermatosis caused by the Sarcoptes scabiei var. hominis mite, which lives and reproduces in humans. Its incidence in Spain has increased in recent years. The aim of this study was to complement existing evidence on the increasing number of scabies cases in our country by analyzing changes in ectoparasiticide prescriptions and Internet searches for scabies infestations measured by Google Trends. We also examined correlations between these two variables. Our results show that public interest in scabies has increased in recent years and is positively and significantly correlated with an increasing use of ectoparasiticides. We believe that Google Trends should be considered as a complementary tool for monitoring real-time trends in scabies infestations in Spain.

Comparison of topical permethrin 5% vs. benzyl benzoate 25% treatment in scabies: a double-blinded randomized controlled trial.

BACKGROUND: Scabies is a pruritic parasitic infestation of the skin. High-income countries have reported an increasing incidence over the last few years. Studies have indicated a reduction in the sensitivity of scabies mites to the standard treatment of choice, topical permethrin 5%. OBJECTIVES: To evaluate in a head-to-head manner the efficacy of two topical scabicides [permethrin 5% and benzyl benzoate 25% (BB)] in the treatment of scabies using the same administration modality; and to address potential confounding factors such as incorrectly performed treatment and hygiene measures. METHODS: In total, 110 patients with dermoscopy-verified scabies infestation were enrolled and randomized into two equally sized groups in a double-blinded manner. Fifty-five received topical permethrin 5% and 55 received topical BB 25%, both for daily use over a period of three consecutive days. Treatment outcome was evaluated by dermoscopy at a 3-week follow-up visit. RESULTS: Treatment resulted in a dermoscopy-verified cure rate of 27% in the permethrin group and 87% in the BB group. The tolerability and safety profile of permethrin 5% cream was excellent, while the BB emulsion produced a burning sensation in 43% of patients. CONCLUSIONS: Topical permethrin demonstrated a lack of efficacy in the majority of scabies cases, whereas BB demonstrated an excellent cure rate and reasonable tolerability. Considering the reduced sensitivity of scabies mites to permethrin 5%, our results suggest that BB is an appropriate first-line therapy in the treatment of scabies.

Crusted scabies in a rabbit model: a severe skin disease or more?

BACKGROUND: Around 200-300 million people are estimated to be affected by scabies annually worldwide. However, the mechanisms by which this disease may affect the general condition of the host are not entirely clear. The aim of the present study was to evaluate the systemic changes that may accompany crusted scabies in both treated and non-treated experimental animals. METHODS: Male New Zealand rabbits were infected with Sarcoptes scabiei var. cuniculi and divided into the following three groups: control, ivermectin-treated, and fluralaner-treated. Several methods were used to evaluate the systemic changes, including histopathological examination of the liver, kidney, heart, and spleen, as well as the measurement of serum biochemical parameters and immunological parameters. RESULTS: Several definite structural and functional changes at the systemic level were revealed, as evidenced by the observed histopathological changes in the tissue sections of internal organs and the highly significant increases in markers of systemic inflammation, serum procalcitonin, and oxidative stress markers. Abnormalities in the liver and renal function results, as well as in the serum lipid profile, were also noted. Additionally, a disorganized immune response was noted, evidenced by a mixed type 1 and type 2 helper T cell response. Although there was notable clinical and parasitological cure in the ivermectin-treated group, the histopathological, biochemical, and immunological markers indicated incomplete resolution. In contrast, the fluralaner-treated group exhibited a nearly complete resolution of changes in these parameters. CONCLUSIONS: We conclude that crusted scabies is a systemic syndrome that can affect several organs besides the skin. Inflammation, oxidative stress, and possibly bacterial infections, are all implicated as underlying mechanisms of tissue damage due to the disease. We recommend that fluralaner, a promising scabicidal agent, should be studied for possible human use, and especially for control programs.

Efficacy of a chewable tablet containing sarolaner, moxidectin, and pyrantel (Simparica Trio®) in the treatment of sarcoptic mange caused by Sarcoptes scabiei mite infestations in dogs.

BACKGROUND: Infestation with Sarcoptes scabiei in dogs is a debilitating disease if left untreated and is transmissible to humans. Two field studies were conducted to confirm the efficacy of orally administered sarolaner in combination with moxidectin and pyrantel (Simparica Trio®) in the treatment of sarcoptic mange in dogs. METHODS: Client-owned dogs with S. scabiei infestation were enrolled and received 2 monthly treatments. In the first, small-scale study, 12 dogs each were allocated randomly to treatment with either placebo or Simparica Trio®. Skin scrapings to detect live mites and assessment of clinical signs of sarcoptic mange were conducted on Days 0, 14, 30, 44, and 60. Efficacy was calculated based on the percent reduction in arithmetic mean live mite counts relative to placebo. In the second, large-scale study, 75 dogs were allocated randomly to treatment with Simparica Trio® and 37 to treatment with afoxolaner + milbemycin oxime (NexGard Spectra®). Skin scrapings to detect live mites and assessment of clinical signs of sarcoptic mange were conducted on Days 0, 14, 30, and 60. The parasitological cure rate (percentage of dogs without live mites) was determined and non-inferiority of Simparica Trio® to the control product was assessed. RESULTS: In the small-scale study, 2 monthly doses of Simparica Trio® resulted in a significant reduction (P ≤ 0.0050) in live S. scabiei mite numbers and provided a 99.2% reduction relative to placebo by Day 60. Clinical signs of sarcoptic mange improved throughout the study in Simparica Trio®-treated dogs. In the large-scale study, the parasitological cure rate on Days 30 and 60 was 97.3% and 100% in the Simparica Trio® group and 91.9% and 100% in the afoxolaner + milbemycin oxime group, respectively. The parasitological cure rate for Simparica Trio® was non-inferior to afoxolaner + milbemycin oxime at both time points. Clinical signs of sarcoptic mange improved throughout the study in both groups. CONCLUSIONS: Two-monthly doses of Simparica Trio® reduced S. scabiei mite counts by 99.2% relative to placebo in one study and eliminated S. scabiei mites in 100% of dogs in the second study, thus confirming that Simparica Trio® is highly effective in the treatment of sarcoptic mange in dogs caused by S. scabiei var. canis.

Optical microscopic study on a novel morphological classification method of multiple diagnostic features of Sarcoptes scabiei var. hominis.

Optical microscopy is the gold standard technique used to confirm the diagnosis of scabies. Multiple diagnostic features of the pathogen Sarcoptes scabiei var. hominis (S. scabiei) can be identified under a microscope and classified into 3 categories: mites, eggs and fecal pellets. However, mite and eggshell fragments can also be observed, which have been ignored in the 2020 International Alliance for the Control of Scabies (IACS) Criteria and by most researchers. In this study, we propose a novel morphological classification method that classifies multiple diagnostic features into 5 categories and 7 subcategories. Our results revealed that 65.2% (1893 of 2896) of the positive cases were confirmed through the identification of mites, eggs or fecal pellets, whereas up to 34.6% (1003 of 2896) of the positive cases were confirmed through the identification of mite or eggshell fragments. Therefore, the important diagnostic values of mite and eggshell fragments should be emphasized. Importantly, for the first time, mite and eggshell fragments were classified into 7 subcategories, some of which are easily ignored or confused with contaminating artefacts. We believe that this novel morphological classification method will be beneficial for operator training in interpreting slides and in improving the 2020 IACS Criteria.