Unveiling Dirofilaria asiatica sp. nov.: First Clinical Insights and Treatment Challenges for this Feline Zoonotic Filarioid Infection—A Cautionary Tale

Unveiling Dirofilaria asiatica sp. nov.: First Clinical Insights and Treatment Challenges for this Feline Zoonotic Filarioid Infection—A Cautionary Tale | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Case Report Unveiling Dirofilaria asiatica sp. nov.: First Clinical Insights and Treatment Challenges for this Feline Zoonotic Filarioid Infection—A Cautionary Tale Angel Almendros, Stefan Hobi, Zhou You, Tiffany Wong, Rebecca Traub This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6921015/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Background: Dirofilaria asiatica sp. nov. (syn. Dirofilaria sp. Hong Kong genotype) is an emerging zoonotic filarioid nematode, initially described in human subcutaneous nodules in Hong Kong and later demonstrated in dogs and cats. This report includes the first description of clinical signs, diagnostic findings, including comparative clinicopathology, treatment and associated clinical complications in a feline infection. Methods: An 18-month-old, spayed female Domestic Shorthair cat presented with subcutaneous nodules and microfilariae in blood smears. Diagnostic work-up included PCR, a quantitative modified Knott’s test, haematology, echocardiography, and abdominal ultrasonography. Results: PCR confirmed the presence of Dirofilaria asiatica sp. nov. Haematology revealed neutrophilia (11.35 × 10³/µL) with left shift, and hyperproteinaemia (80 g/L). A quantitative modified Knott’s test revealed a microfilaremia of 36,907 per ml. Treatment with oral doxycycline and topical moxidectin triggered an acute onset of respiratory distress, managed with oxygen and dexamethasone. Nodules regressed, and microfilariae cleared by day 70. Conclusions: This case supports that indoor feline pets are susceptible to infection with Dirofilaria asiatica sp. nov. In addition, we describe for the first time, the clinical and clinicopathological findings associated with infection and highlight the risks of immunologic reactions to microfilaricidal therapy. Moxidectin and doxycycline were critical for resolution, but pre-treatment with corticosteroids is recommended. Exorbitant microfilaremia might be a critical feature in cats. The zoonotic potential of this parasite warrants heightened surveillance in endemic regions. Dirofilaria hongkongensis asiatica feline dirofilariasis zoonosis moxidectin subcutaneous nodules acute pulmonary inflammation respiratory distress Figures Figure 1 Figure 2 Figure 3 Figure 4 Background Dirofilaria sp. Hong Kong genotype, recently classified as Dirofilaria asiatica sp. nov. [ 1 ], is a putative species that was first identified in 2012 from human subcutaneous nodules and subsequently detected in dogs by PCR, suggesting a zoonotic cycle involving Culicinae vectors [ 2 ]. Phylogenetically, it clusters within the Nochtiella subgenus, closely related to D. repens , but exhibits distinct genetic markers such as cox1 and 12S rRNA sequences [ 3 , 4 ]. While D. repens primarily causes subcutaneous and ocular infections in dogs and humans across Europe and Asia, Dirofilaria asiatica sp. nov. appears restricted to Asia, with reports from India, Sri-Lanka, Bhutan [ 5 – 7 ], Thailand, and more recently Hong Kong [ 8 , 9 ], primarily affecting dogs and to a lesser degree cats. Zoonotic cases of Dirofilaria asiatica sp. nov. in humans have been reported in India and Thailand, and in returned travellers from Europe and Australia [ 10 – 14 ]. Recent studies confirmed Dirofilaria asiatica sp. nov. within subcutaneous nodules of dogs and cats in Hong Kong [ 9 ]. Nodules were posteriorly located and filaroids were identified by histopathological assessment and species confirmed by PCR. The nematodes had longitudinal cuticular ridges typical of the subgenus Notchiella , and the lesions resembled D. repens , consisting of granulomatous to pyogranulomatous dermatitis with eosinophilic infiltrates [ 9 ]. Nevertheless, how Dirofilaria asiatica sp. nov. affects dogs or cats remains undocumented. Diagnosis relies on PCR targeting cox1 and 12S rRNA due to morphological overlap of microfilaria with D. repens [ 3 ]. Treatment has never been described in dogs and cats but might present challenges mirroring those of D. immitis in dogs, where potent microfilaricidal drugs such as diethylcarbamazine (DEC) can provoke fatal anaphylactic reactions owing to a sudden and significant destruction of circulating microfilariae [ 15 ] and pulmonary inflammation [ 16 ]. This case report offers novel insights into feline Dirofilaria asiatica sp. nov. infection, highlighting the challenges associated with its therapeutic management. Case Presentation An 18-month-old spayed female Domestic Shorthair cat was referred to the Veterinary Medical Centre of City University of Hong Kong (VMC CityUHK) for further investigation of a 2-week history of subcutaneous nodules previously explored at the referring veterinarian, revealing the presence of a long, thin worm. A blood smear had identified microfilaria but no further investigation had been performed. Other than intermittent fur ball vomiting, the cat was bright and alert, had a good appetite and no other gastrointestinal signs were reported. On day 1 (D1) the body condition score (BCS) was 5/9 with a weight of 4.14 kg. Initial physical and dermatological examination revealed two palpable subcutaneous, intact and non-moveable nodules, including a small, raised nodule (2 mm) on the left elbow region, and a larger nodule (5 mm) on the left pinnae. The nodules were reported not to be pruritic or painful. Cardiothoracic auscultation was unremarkable, with a heart rate (HR) of 186 bpm, and a respiratory rate (RR) of 32 bpm. Her mucous membranes were pink and moist, and no obvious pulse deficits were detected. The rest of the physical exam was unremarkable. An in-house blood smear exam revealed numerous microfilariae morphologically suggestive of filarial species (Fig. 1 ). Blood samples were submitted to the Veterinary Diagnostic Laboratory of City University of Hong Kong (VDL CityUHK) for a complete cell count and to the Department of Infectious Diseases and Public Health of CityUHK for a Knott’s test and PCR assay for identification of microfilariae. On day 2, the results of the haematological assessment revealed neutrophilia (11.35 × 10³/µL; reference interval - RI, 2.30–10.29) with a left shift (bands: 0.15 × 10³/µL; RI, 0.00–0.10), hyperproteinaemia (80 g/L; RI, 59–75), and a normal eosinophil count (0.92 × 10³/µL; RI, 0.10–1.80). For quantitation and species identification, 1 ml of whole blood was subjected to a Modified Knott's Test (MKT) stained with 0.1% methylene blue [ 17 ]. The average count across three 10 µl aliquots of the modified Knott’s test sediment revealed a microfilarial count of 36,907/ml of blood. Microfilariae were unsheathed, with a blunt head and curved tail (Fig. 2 ) as per previous descriptions of Dirofilaria sp. Hong Kong genotype / asiatica sp. nov. [ 6 ]. For molecular characterization, DNA was extracted from whole blood using the DNeasy Blood and Tissue Kit (Qiagen, Germany) according to the manufacturer’s protocols. Conventional PCR amplification was performed targeting a 763 bp region of the mitochondrial (mt) cytochrome oxidase – 1 (cox-1) gene [ 4 ]. PCR reactions were carried out in a final volume of 20 µl, containing 10 µl 2X Phanta Flash Master Mix (Vazyme, China), 1 µl of each primer (10 µM), 6 µl distilled water and 2 µl of template DNA. The mixtures were amplified using 35 cycles under the following conditions: denaturation at 98°C for 40 s, annealing at 58°C for 30 s, and extension at 72°C for 1 min, followed by a final extension at 72°C for 1 min using the SimpliAmp thermocycler (Thermofisher, USA). The PCR products were visualized after electrophoresis using 2% agarose gel and submitted for bidirectional Sanger sequencing at BGI Genomics (Guangdong, China). Clear and readable sequences of the 763 bp region of the cox gene were directly compared with reference sequences using BLASTn, which revealed between 99.86–100% sequence identity with Dirofilaria sp. ‘ hongkongensis ’ over 88–96% sequence coverage, isolated from cats, dogs and humans originating in India and in Hong Kong (GenBank accession numbers PQ327004-5, NC031365, OL744441). The PCR result was available on day 15 and the patient returned on day 17 for further evaluation. The cat was bright and alert, eating well, with a BCS of 5/9, and a body weight of 4.04 kg. A new nodule had developed on her right forelimb of approximately 5 mm, while the previously reported nodules on the left forelimb and the left pinnae remained unchanged. An echocardiograph was unremarkable with no visible adult parasites in the heart chambers or in the pulmonary artery as it might occur in cases of D. immitis infection. An abdominal ultrasound showed mild reactive mid abdominal lymphadenopathy but there was no evidence of visceral parasites. Haematology and biochemistry were unremarkable and a urine analysis revealed urine dilution (SG 1.016), protein traces and a pH of 6 (RI, 6–7). For further exploration of the cutaneous nodules, the patient was sedated intramuscularly with butorphanol (Torbugesic, Zoetis, Australia) 0.3 mg/kg and dexmedetomidine (Dexdomitor, Zoetis, Australia) 6 µg/kg. One large live filaroid was extracted from the ear nodule and was submitted for pathological examination (Fig. 3 ). After confirming the absence of worms within the cardiovascular system, the patient was prescribed a topical heartworm preventative (Advocate, Elanco, USA), containing imidacloprid 10% and moxidectin 1% to reduce the microfilaria burden. Additionally, oral doxycycline (Vibramycin, Pfizer, USA) was prescribed for 30 days at 10 mg/kg q 24h alongside the topical treatment. The following day (D18), prednisolone (Macrolone, Mavlab, Australia) 2 mg/kg was additionally prescribed due to concerns regarding the effects of microfilarial death causing the increased respiratory rate reported by the owner, but the drugs were never collected. The cat presented to the emergency department on the evening of day 18, approximately 30 hours post-treatment, after developing an onset of acute respiratory distress with a RR of 92 bpm, oxygen saturation (SpO₂) of 83%, and diffuse interstitial lung patterns on radiography (Fig. 3 ). The cat was admitted to the hospital and was initially managed with oxygen therapy (15 L/min), dexamethasone (Dexafort, Merck, USA) 0.1 mg/kg IV q12h, and ampicillin-sulbactam (Unasyn, Pfizer, USA) 30 mg/kg IV q8h. Initial blood work upon admission revealed mild hyperglycaemia (11.34 mmol/L;RI, 4.11–8.84), likely due to stress, along with a mild decrease in urea (4.7 mmol/L; RI, 5.7–12.9). The white blood cell count was normal, but the patient was mildly neutropenic (2.11 × 10³/µL; RI, 2.30–10.29) and exhibited mild eosinopenia (0.08 × 10³/µL; RI, 0.17–1.57). Blood gas (venous) values included pH 7.31 (RI, 7.22–7.38), HCO3 19.5 mmol/L (RI, 15.4–23.4), pCO2 41.1 mmHg (RI, 32.8–50.8), pO2 33.3 mmHg (27.6–49.6). Oximetry parameters in venous blood included cHb 10.9 g/dL (9.8–15.4), sO2 43.4% (62.4 +/- 13.5), and the metabolite Lactate measurement was 2 mmol/L [ 18 , 19 ]. During hospitalization, a nasogastric tube (NGT) was inserted for nutritional support. Supportive treatment included 8 ml/hr IV Ringer’s lactate infusion solution (LRS), 0.1 mg/kg dexamethasone (Dexafort, Merck, USA) q12h IV, 30 mg/kg ampicillin-sulbactam (Unasyn, Pfizer, USA) q8h IV, 10 mg/kg doxycycline (Vibramycin, Pfizer, USA) q24h via NGT, 0.02 mg/kg buprenorphine (Zorbium, Elanco, USA) q6h IV, and 1 mg/kg maropitant (Cerenia, Zoetis, USA) q24h IV. The oxygen level was adjusted hourly, ranging from 10 to 15 L/min. The cat was discharged 7 days after admission when the RR returned to normal. On day 41, the patient returned for a blood test. Samples were collected in EDTA for a blood smear examination and a microfilaria test. A microscopical assessment in-house revealed absence of microfilaria under stained and fresh blood smear exams. The sample was submitted for further analysis to a referral laboratory, and resolution of infection was confirmed by a clear Knott’s test and a negative PCR on day 65. Nodules regressed by day 70. A final check-up on day 72 revealed no abnormalities on echocardiogram or abdominal ultrasound. The patient was discharged from the clinic with instructions to continue long-term application of imidacloprid 10% and moxidectin 1% spot-on formulation (Advocate, Elanco, USA). No recurrence of clinical signs was observed over 3 months after the last check-up. Discussion This case unveils for the first time the effect of Dirofilaria asiatica sp. nov.on a feline patient ( Felis catus ) describing detailed clinical signs, diagnostic confirmation and treatment outcome. Other than the description of histopathological lesions and molecular confirmation, little has been reported about this parasite and its effect on dogs and cats [ 9 , 20 ]. Weight loss and vomiting were mentioned in the history notes in one cat in a previous study, but no diagnostic tests to investigate the potential association were available [ 9 ]. The present case additionally highlights potential life threatening complications after the use of a macrocyclic lactone, a common treatment for gastrointestinal nematodes and prophylaxis for heartworm (Imidacloprid 10% and moxidectin 1% spot-on formulation), that was used for its known microfilaricidal and long-term adulticidal effects. This case supports previous findings where felines expanded the known host range beyond humans and dogs for Dirofilaria asiatica sp. nov. [ 9 , 20 ]. A clinicopathological overlap was demonstrated by the cat’s subcutaneous nodules and microfilariae, mirroring D. repens infections and further emphasizing the need for molecular differentiation in endemic regions [ 21 ]. In the present case, PCR unequivocally identified Dirofilaria sp. ‘hongkongensis’/ Dirofilaria asiatica sp. nov.. Unlike D. immitis , Dirofilaria asiatica sp. nov. exhibited tropism for subcutaneous tissues but absence of cardiac involvement, corroborating prior reports [ 9 , 20 ]. Serum chemistry was unremarkable in this case, with marginal hyperglycemia and hyperproteinemia likely due to stress and an ongoing inflammatory process. Neutrophilia and a left shift were present and were interpreted as an active inflammatory response. Other than occasional furball-associated vomiting, no clinical signs were associated with this cutaneous filarioid infection. The presence of mid abdominal reactive lymph nodes was non-specific, however, previous reports of D. repens and Dirofilaria asiatica sp. nov. have also identified lymphadenopathy [ 9 , 20 , 22 ]. Reactive lymphadenopathy is not uncommon and could be expected in young cats due to antigenic stimulation with or without gastrointestinal signs such as intermittent vomiting in this case [ 23 , 24 ]. Feline immunodeficiency virus (FIV) and feline leukaemia virus (FeLV) tests were negative in our cat, which did not show any obvious co-morbidities or co-infections as it might be seen in other cases of Dirofilaria infection [ 25 ]. The location of the skin lesions in this case were different to a previous study, where all 5 cases including 4 male cats and a female dog presented lesions posteriorly [ 9 ]. Different to previous reports the cat affected here was a female. In this case the skin lesions were anterior, at the left pinna and both elbow regions, which would align with the biting preferences of mosquitos. Interestingly, in the current case the nodules were neither pruritic nor painful, which is in contrast to other cases of cutaneous dirofilariasis where pruritus is commonly seen and may even mimic allergic dermatitis [ 26 ]. Whether this is associated with a reduced movement of Dirofilaria asiatica sp. nov., a modified expression of secretion products or individual factors, needs to be further elucidated. Treatment complications occurred due to an immunologic reaction demonstrated by the severe respiratory distress following moxidectin and doxycycline application. This has been observed with D. immitis treatment in dogs linked to a sudden and severe destruction of circulating microfilaria, most frequently seen with the administration of DEC [ 15 ]. Corticosteroids were critical to mitigating this reaction. Although macrocyclic lactones are significantly safer than DEC in attaining a more gradual reduction in microfilarial clearance, the extremely high microfilarial count in this cat coupled with the use of doxycycline in combination with moxidectin, could have potentiated this reaction. Macrocyclic lactones in combination with doxycycline are known to have significant higher microfilaricidal effects compared to macrocyclic lactones alone [ 27 ]. This warrants caution due to the risk of anaphylaxis. Treating with macrocytic lactones alone and pretreatment with prednisolone or dexamethasone is advisable in future cases. This case alerts clinicians for potential acute respiratory crises following the treatment with preventative care drugs such as moxidectin that can be further exacerbated with the administration of concurrent doxycycline. Since antigen tests are not available for this nematode species, microfilarial identification by microscopy might be a sensible consideration. There are several limitations of this study, including the design being retrospective in nature and including only one case. Avoidance of infection would be better achieved by having a confirmed vector. While Culicinae vectors such as Aedes spp. are suspected, vector competence studies are needed. Another limitation was the lack of skin biopsies due to owner refusal, although a nematode could be directly isolated from associated lesions. Nevertheless, a granulomatous inflammation seen in previous studies was suspected based on the appearance of lesions but could not be confirmed histopathologically. The blood gas analysis was from venous origin in this case, hence the ranges are adjusted. This might explain the oxygenation discrepancy in blood gas PO2 versus sPO2 where the latter is more reliable in venous samples. However, ventilation and acid-base can be estimated and evaluated respectively in venous samples when arterial samples are not available [ 18 ]. In this case we emphasize the zoonotic and epidemiological significance of felines as potential reservoirs for Dirofilaria asiatica sp. nov. Given the high microfilarial counts, surveillance and monthly prophylaxis for Hong Kong's feline population are crucial to mitigate risks to the public. The summer onset of symptoms in this case report coincides with the peak of the mosquito activity. Vector dynamics therefore are important and suggest summer months to be a higher risk for transmission [ 20 , 28 ]. Conclusion This case highlights Dirofilaria asiatica sp. nov. as an emerging feline pathogen with zoonotic potential. Affected individuals may only show subcutaneous nodules, mimicking neoplastic, inflammatory or other infectious pathologies, underlining the need for further diagnostics. Treatment requires multimodal therapy to address both parasitosis and immunologic complications. Public health messaging should emphasize on year-round parasite and vector prevention in pets, particularly in endemic regions like Hong Kong. Declarations I hereby confirm that the owner and legal guardian of the patient consented to participate in this clinical case and its publication. The authors confirm that the ethical policies of the journal, as noted on the journal’s author guidelines page, have been adhered to. No ethical approval was required as this is a retrospective study with no original research data but analysing only history records. Informed owner consent was obtained. Acknowledgements The authors acknowledge the help of clinicians at VMC CityUHK and pathologists at VDL CityUHK for their help with data collection and laboratory interpretation. Author contributions TW was the primary clinician of the case and collected clinical data, together with AA provided care and treatment for the cat. AA and TW collected nematode samples. AA drafted the initial manuscript and TW, SH and RT reviewed and edited the manuscript accordingly. YZ carried out the PCR assay and DNA sequencing analysis. RT performed and interpreted results of the Knott’s test and supervised the molecular diagnostics . All authors read and approved the final manuscript. Funding This research received no external funding. This study was funded with a grant from City University of Hong Kong to AA (Project No. 9610596) Availability of data and materials Data are available upon request, with patient identifiers redacted. Consent for publication All authors consent to this manuscript’s publication. Competing interests The authors disclose no conflicts of interest. References Colella V, Young ND, Manzanell R, Atapattu U, Sunita B, Sumanam LG, Huggins AV, Koehler, Robin B, Gasser (2025) Dirofilaria Asiatica Sp. Nov.(Spirurida: Onchocercidae)–Defined Using a Combined Morphological-Molecular Approach. 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Abdominal Lymph Nodes, Peritoneal Cavity and Aorta. J Feline Med Surg 23(9):835–849 Carbonara M, Perles L, Venco L, Gabrielli S, Vanessa R, Barrs G, Miró E, Papadopoulos (2025) Clara Lima, Emilie Bouhsira, and Gad Baneth. Dirofilaria Spp. Infection in Cats from the Mediterranean Basin: Diagnosis and Epidemiology. Int J Parasitol Tarello W (2011) Clinical Aspects of Dermatitis Associated with Dirofilaria Repens in Pets: A Review of 100 Canine and 31 Feline Cases (1990–2010) and a Report of a New Clinic Case Imported from Italy to Dubai. J Parasitol Res no. 1 (2011): 578385 Kramer L, Serena Crosara G, Gnudi M, Genchi C, Mangia A, Viglietti, Quintavalla C (2018) Wolbachia, Doxycycline and Macrocyclic Lactones: New Prospects in the Treatment of Canine Heartworm Disease. Vet Parasitol 254:95–97 Otranto D, Capelli G, Genchi C (2009) Changing Distribution Patterns of Canine Vector Borne Diseases in Italy: Leishmaniosis Vs. Dirofilariosis Parasit Vectors 2:1–8 Additional Declarations The authors declare no competing interests. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-6921015","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Case Report","associatedPublications":[],"authors":[{"id":472982653,"identity":"c99e6cf5-7e9e-4397-98ea-b56445c3189d","order_by":0,"name":"Angel Almendros","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA70lEQVRIiWNgGAWjYFAC5gYQwcDPwNgAETjAwAZj4gCMEC2SDYyNDaRpMTgAs4aQFv72g60bPu6wzjc+frj9MW8bgxzfjQS2hzPwaJE4k9h2c+aZdMttZxIbm4FajCVvJLAbbsDnrgOJbbd52w4bmN1gBGtJ3AC0RfIBHh3y5x+23f4L1GI8A6KlnqAWgxtAWxiBWgwkIFoSDEBa8DnM8MbDtpu9bekGQE81zpxzTsJw5pmHbZL4vC93PvnYjZ9t1gb87ccffHhTZiPPdzz5mGQPPu8jAyYeBgkGBgYCEYkCGH8Qr3YUjIJRMApGEAAAa9pXshh/VckAAAAASUVORK5CYII=","orcid":"https://orcid.org/0000-0002-2441-7850","institution":"Department of Veterinary Clinical Sciences, Jockey Club College of Veterinary Medicine and Life Sciences, City University of Hong Kong","correspondingAuthor":true,"prefix":"","firstName":"Angel","middleName":"","lastName":"Almendros","suffix":""},{"id":472982654,"identity":"5c4b5a36-2316-41d9-9160-759cc3723ec2","order_by":1,"name":"Stefan Hobi","email":"","orcid":"","institution":"Department of Veterinary Clinical Sciences, Jockey Club College of Veterinary Medicine and Life Sciences, City University of Hong Kong","correspondingAuthor":false,"prefix":"","firstName":"Stefan","middleName":"","lastName":"Hobi","suffix":""},{"id":472982655,"identity":"1ff3b0b3-75bf-4f3c-9771-086516978f27","order_by":2,"name":"Zhou You","email":"","orcid":"","institution":"Department of Infectious Diseases and Public Health, Jockey Club College of Veterinary Medicine and Life Sciences, City University of Hong Kong","correspondingAuthor":false,"prefix":"","firstName":"Zhou","middleName":"","lastName":"You","suffix":""},{"id":472982656,"identity":"7b76894e-d3c5-4c62-8c70-d56eae633f3a","order_by":3,"name":"Tiffany Wong","email":"","orcid":"","institution":"CityU Veterinary Medical Centre, City University of Hong Kong","correspondingAuthor":false,"prefix":"","firstName":"Tiffany","middleName":"","lastName":"Wong","suffix":""},{"id":472982657,"identity":"f0308cb0-4e2b-4194-84af-867fc9abce20","order_by":4,"name":"Rebecca Traub","email":"","orcid":"","institution":"Department of Infectious Diseases and Public Health, Jockey Club College of Veterinary Medicine and Life Sciences, City University of Hong Kong","correspondingAuthor":false,"prefix":"","firstName":"Rebecca","middleName":"","lastName":"Traub","suffix":""}],"badges":[],"createdAt":"2025-06-18 08:43:12","currentVersionCode":1,"declarations":{"humanSubjects":false,"vertebrateSubjects":true,"conflictsOfInterestStatement":false,"humanSubjectEthicalGuidelines":false,"humanSubjectConsent":false,"humanSubjectClinicalTrial":false,"humanSubjectCaseReport":false,"vertebrateSubjectEthicalGuidelines":true},"doi":"10.21203/rs.3.rs-6921015/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6921015/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":85305589,"identity":"3275deb7-5479-48bb-9e2e-e43ad5fb921c","added_by":"auto","created_at":"2025-06-24 12:40:01","extension":"jpeg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":59471,"visible":true,"origin":"","legend":"\u003cp\u003eStained blood smear (Diff-Quick). Microfilaria surrounded by red blood cells, neutrophils and an eosinophil from a cat blood sample (100x magnification).\u003c/p\u003e","description":"","filename":"floatimage1.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-6921015/v1/1bf8f2bc808ba7372dfd4763.jpeg"},{"id":85303961,"identity":"f2b6e11f-14d6-4c0e-a6ea-baa738fca00d","added_by":"auto","created_at":"2025-06-24 12:32:01","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":965583,"visible":true,"origin":"","legend":"\u003cp\u003eKnott’s test. Microfilaria fixed in 2% formalin and stained with methylene blue displaying curved tail, blunt head and 3 cephalic nuclei from a cat blood sample (40x magnification).\u003c/p\u003e","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-6921015/v1/8a8f339d85a238cbf2e9b0ad.png"},{"id":85306075,"identity":"70103bcd-2b58-4ec8-a99c-992af560e7d3","added_by":"auto","created_at":"2025-06-24 12:48:01","extension":"jpeg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":140232,"visible":true,"origin":"","legend":"\u003cp\u003eExploration of a nodule on the left pinna of a cat containing and adult filaroid.\u003c/p\u003e","description":"","filename":"floatimage3.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-6921015/v1/a0c9f6baa882eeda4172dd61.jpeg"},{"id":85303967,"identity":"5b201e15-e556-42b1-9917-7bcded78e88b","added_by":"auto","created_at":"2025-06-24 12:32:01","extension":"jpeg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":322013,"visible":true,"origin":"","legend":"\u003cp\u003eRadiographic interstitial lung pattern in a microfilaremic cat following moxidectin and doxycycline treatment. Right lateral view on the left and dorsoventral view on the right.\u003c/p\u003e","description":"","filename":"floatimage4.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-6921015/v1/3ac53c71c61c0f0607cee12d.jpeg"},{"id":85306076,"identity":"96dfc871-5c92-412e-96e0-9ec3511db5f6","added_by":"auto","created_at":"2025-06-24 12:48:06","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1813190,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6921015/v1/bf3ce6f1-e9ab-4475-a78c-091051b791be.pdf"}],"financialInterests":"The authors declare no competing interests.","formattedTitle":"\u003cp\u003eUnveiling Dirofilaria asiatica sp. nov.: First Clinical Insights and Treatment Challenges for this Feline Zoonotic Filarioid Infection—A Cautionary Tale\u003c/p\u003e","fulltext":[{"header":"Background","content":"\u003cp\u003e \u003cem\u003eDirofilaria\u003c/em\u003e sp. Hong Kong genotype, recently classified as \u003cem\u003eDirofilaria asiatica\u003c/em\u003e sp. nov. [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e], is a putative species that was first identified in 2012 from human subcutaneous nodules and subsequently detected in dogs by PCR, suggesting a zoonotic cycle involving Culicinae vectors [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. Phylogenetically, it clusters within the \u003cem\u003eNochtiella\u003c/em\u003e subgenus, closely related to \u003cem\u003eD. repens\u003c/em\u003e, but exhibits distinct genetic markers such as cox1 and 12S rRNA sequences [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. While \u003cem\u003eD. repens\u003c/em\u003e primarily causes subcutaneous and ocular infections in dogs and humans across Europe and Asia, \u003cem\u003eDirofilaria asiatica\u003c/em\u003e sp. nov. appears restricted to Asia, with reports from India, Sri-Lanka, Bhutan [\u003cspan additionalcitationids=\"CR6\" citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e], Thailand, and more recently Hong Kong [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e], primarily affecting dogs and to a lesser degree cats. Zoonotic cases of \u003cem\u003eDirofilaria asiatica\u003c/em\u003e sp. nov. in humans have been reported in India and Thailand, and in returned travellers from Europe and Australia [\u003cspan additionalcitationids=\"CR11 CR12 CR13\" citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eRecent studies confirmed \u003cem\u003eDirofilaria asiatica\u003c/em\u003e sp. nov. within subcutaneous nodules of dogs and cats in Hong Kong [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Nodules were posteriorly located and filaroids were identified by histopathological assessment and species confirmed by PCR. The nematodes had longitudinal cuticular ridges typical of the subgenus \u003cem\u003eNotchiella\u003c/em\u003e, and the lesions resembled \u003cem\u003eD. repens\u003c/em\u003e, consisting of granulomatous to pyogranulomatous dermatitis with eosinophilic infiltrates [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Nevertheless, how \u003cem\u003eDirofilaria asiatica\u003c/em\u003e sp. nov. affects dogs or cats remains undocumented.\u003c/p\u003e \u003cp\u003eDiagnosis relies on PCR targeting cox1 and 12S rRNA due to morphological overlap of microfilaria with \u003cem\u003eD. repens\u003c/em\u003e [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. Treatment has never been described in dogs and cats but might present challenges mirroring those of \u003cem\u003eD. immitis\u003c/em\u003e in dogs, where potent microfilaricidal drugs such as diethylcarbamazine (DEC) can provoke fatal anaphylactic reactions owing to a sudden and significant destruction of circulating microfilariae [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e] and pulmonary inflammation [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThis case report offers novel insights into feline \u003cem\u003eDirofilaria asiatica\u003c/em\u003e sp. nov. infection, highlighting the challenges associated with its therapeutic management.\u003c/p\u003e"},{"header":"Case Presentation","content":"\u003cp\u003eAn 18-month-old spayed female Domestic Shorthair cat was referred to the Veterinary Medical Centre of City University of Hong Kong (VMC CityUHK) for further investigation of a 2-week history of subcutaneous nodules previously explored at the referring veterinarian, revealing the presence of a long, thin worm. A blood smear had identified microfilaria but no further investigation had been performed. Other than intermittent fur ball vomiting, the cat was bright and alert, had a good appetite and no other gastrointestinal signs were reported. On day 1 (D1) the body condition score (BCS) was 5/9 with a weight of 4.14 kg. Initial physical and dermatological examination revealed two palpable subcutaneous, intact and non-moveable nodules, including a small, raised nodule (2 mm) on the left elbow region, and a larger nodule (5 mm) on the left pinnae. The nodules were reported not to be pruritic or painful. Cardiothoracic auscultation was unremarkable, with a heart rate (HR) of 186 bpm, and a respiratory rate (RR) of 32 bpm. Her mucous membranes were pink and moist, and no obvious pulse deficits were detected. The rest of the physical exam was unremarkable. An in-house blood smear exam revealed numerous microfilariae morphologically suggestive of filarial species (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). Blood samples were submitted to the Veterinary Diagnostic Laboratory of City University of Hong Kong (VDL CityUHK) for a complete cell count and to the Department of Infectious Diseases and Public Health of CityUHK for a Knott\u0026rsquo;s test and PCR assay for identification of microfilariae.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eOn day 2, the results of the haematological assessment revealed neutrophilia (11.35 \u0026times; 10\u0026sup3;/\u0026micro;L; reference interval - RI, 2.30\u0026ndash;10.29) with a left shift (bands: 0.15 \u0026times; 10\u0026sup3;/\u0026micro;L; RI, 0.00\u0026ndash;0.10), hyperproteinaemia (80 g/L; RI, 59\u0026ndash;75), and a normal eosinophil count (0.92 \u0026times; 10\u0026sup3;/\u0026micro;L; RI, 0.10\u0026ndash;1.80). For quantitation and species identification, 1 ml of whole blood was subjected to a Modified Knott's Test (MKT) stained with 0.1% methylene blue [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. The average count across three 10 \u003cem\u003e\u0026micro;l\u003c/em\u003e aliquots of the modified Knott\u0026rsquo;s test sediment revealed a microfilarial count of 36,907/ml of blood. Microfilariae were unsheathed, with a blunt head and curved tail (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e) as per previous descriptions of \u003cem\u003eDirofilaria\u003c/em\u003e sp. Hong Kong genotype\u003cem\u003e/ asiatica\u003c/em\u003e sp. nov. [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e].\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eFor molecular characterization, DNA was extracted from whole blood using the DNeasy Blood and Tissue Kit (Qiagen, Germany) according to the manufacturer\u0026rsquo;s protocols.\u003c/p\u003e \u003cp\u003eConventional PCR amplification was performed targeting a 763 bp region of the mitochondrial (mt) cytochrome oxidase \u0026ndash; 1 (cox-1) gene [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. PCR reactions were carried out in a final volume of 20 \u0026micro;l, containing 10 \u0026micro;l 2X Phanta Flash Master Mix (Vazyme, China), 1 \u0026micro;l of each primer (10 \u0026micro;M), 6 \u0026micro;l distilled water and 2 \u0026micro;l of template DNA. The mixtures were amplified using 35 cycles under the following conditions: denaturation at 98\u0026deg;C for 40 s, annealing at 58\u0026deg;C for 30 s, and extension at 72\u0026deg;C for 1 min, followed by a final extension at 72\u0026deg;C for 1 min using the SimpliAmp thermocycler (Thermofisher, USA). The PCR products were visualized after electrophoresis using 2% agarose gel and submitted for bidirectional Sanger sequencing at BGI Genomics (Guangdong, China). Clear and readable sequences of the 763 bp region of the cox gene were directly compared with reference sequences using BLASTn, which revealed between 99.86\u0026ndash;100% sequence identity with \u003cem\u003eDirofilaria\u003c/em\u003e sp. \u0026lsquo;\u003cem\u003ehongkongensis\u003c/em\u003e\u0026rsquo; over 88\u0026ndash;96% sequence coverage, isolated from cats, dogs and humans originating in India and in Hong Kong (GenBank accession numbers PQ327004-5, NC031365, OL744441).\u003c/p\u003e \u003cp\u003eThe PCR result was available on day 15 and the patient returned on day 17 for further evaluation. The cat was bright and alert, eating well, with a BCS of 5/9, and a body weight of 4.04 kg. A new nodule had developed on her right forelimb of approximately 5 mm, while the previously reported nodules on the left forelimb and the left pinnae remained unchanged. An echocardiograph was unremarkable with no visible adult parasites in the heart chambers or in the pulmonary artery as it might occur in cases of \u003cem\u003eD. immitis\u003c/em\u003e infection. An abdominal ultrasound showed mild reactive mid abdominal lymphadenopathy but there was no evidence of visceral parasites. Haematology and biochemistry were unremarkable and a urine analysis revealed urine dilution (SG 1.016), protein traces and a pH of 6 (RI, 6\u0026ndash;7).\u003c/p\u003e \u003cp\u003eFor further exploration of the cutaneous nodules, the patient was sedated intramuscularly with butorphanol (Torbugesic, Zoetis, Australia) 0.3 mg/kg and dexmedetomidine (Dexdomitor, Zoetis, Australia) 6 \u0026micro;g/kg. One large live filaroid was extracted from the ear nodule and was submitted for pathological examination (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). After confirming the absence of worms within the cardiovascular system, the patient was prescribed a topical heartworm preventative (Advocate, Elanco, USA), containing imidacloprid 10% and moxidectin 1% to reduce the microfilaria burden. Additionally, oral doxycycline (Vibramycin, Pfizer, USA) was prescribed for 30 days at 10 mg/kg q 24h alongside the topical treatment.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe following day (D18), prednisolone (Macrolone, Mavlab, Australia) 2 mg/kg was additionally prescribed due to concerns regarding the effects of microfilarial death causing the increased respiratory rate reported by the owner, but the drugs were never collected. The cat presented to the emergency department on the evening of day 18, approximately 30 hours post-treatment, after developing an onset of acute respiratory distress with a RR of 92 bpm, oxygen saturation (SpO₂) of 83%, and diffuse interstitial lung patterns on radiography (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). The cat was admitted to the hospital and was initially managed with oxygen therapy (15 L/min), dexamethasone (Dexafort, Merck, USA) 0.1 mg/kg IV q12h, and ampicillin-sulbactam (Unasyn, Pfizer, USA) 30 mg/kg IV q8h.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eInitial blood work upon admission revealed mild hyperglycaemia (11.34 mmol/L;RI, 4.11\u0026ndash;8.84), likely due to stress, along with a mild decrease in urea (4.7 mmol/L; RI, 5.7\u0026ndash;12.9). The white blood cell count was normal, but the patient was mildly neutropenic (2.11 \u0026times; 10\u0026sup3;/\u0026micro;L; RI, 2.30\u0026ndash;10.29) and exhibited mild eosinopenia (0.08 \u0026times; 10\u0026sup3;/\u0026micro;L; RI, 0.17\u0026ndash;1.57). Blood gas (venous) values included pH 7.31 (RI, 7.22\u0026ndash;7.38), HCO3 19.5 mmol/L (RI, 15.4\u0026ndash;23.4), pCO2 41.1 mmHg (RI, 32.8\u0026ndash;50.8), pO2 33.3 mmHg (27.6\u0026ndash;49.6). Oximetry parameters in venous blood included cHb 10.9 g/dL (9.8\u0026ndash;15.4), sO2 43.4% (62.4 +/- 13.5), and the metabolite Lactate measurement was 2 mmol/L [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eDuring hospitalization, a nasogastric tube (NGT) was inserted for nutritional support. Supportive treatment included 8 ml/hr IV Ringer\u0026rsquo;s lactate infusion solution (LRS), 0.1 mg/kg dexamethasone (Dexafort, Merck, USA) q12h IV, 30 mg/kg ampicillin-sulbactam (Unasyn, Pfizer, USA) q8h IV, 10 mg/kg doxycycline (Vibramycin, Pfizer, USA) q24h via NGT, 0.02 mg/kg buprenorphine (Zorbium, Elanco, USA) q6h IV, and 1 mg/kg maropitant (Cerenia, Zoetis, USA) q24h IV. The oxygen level was adjusted hourly, ranging from 10 to 15 L/min.\u003c/p\u003e \u003cp\u003eThe cat was discharged 7 days after admission when the RR returned to normal.\u003c/p\u003e \u003cp\u003eOn day 41, the patient returned for a blood test. Samples were collected in EDTA for a blood smear examination and a microfilaria test. A microscopical assessment in-house revealed absence of microfilaria under stained and fresh blood smear exams. The sample was submitted for further analysis to a referral laboratory, and resolution of infection was confirmed by a clear Knott\u0026rsquo;s test and a negative PCR on day 65. Nodules regressed by day 70. A final check-up on day 72 revealed no abnormalities on echocardiogram or abdominal ultrasound. The patient was discharged from the clinic with instructions to continue long-term application of imidacloprid 10% and moxidectin 1% spot-on formulation (Advocate, Elanco, USA). No recurrence of clinical signs was observed over 3 months after the last check-up.\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eThis case unveils for the first time the effect of \u003cem\u003eDirofilaria asiatica\u003c/em\u003e sp. nov.on a feline patient (\u003cem\u003eFelis catus\u003c/em\u003e) describing detailed clinical signs, diagnostic confirmation and treatment outcome. Other than the description of histopathological lesions and molecular confirmation, little has been reported about this parasite and its effect on dogs and cats [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e]. Weight loss and vomiting were mentioned in the history notes in one cat in a previous study, but no diagnostic tests to investigate the potential association were available [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. The present case additionally highlights potential life threatening complications after the use of a macrocyclic lactone, a common treatment for gastrointestinal nematodes and prophylaxis for heartworm (Imidacloprid 10% and moxidectin 1% spot-on formulation), that was used for its known microfilaricidal and long-term adulticidal effects.\u003c/p\u003e \u003cp\u003eThis case supports previous findings where felines expanded the known host range beyond humans and dogs for \u003cem\u003eDirofilaria asiatica\u003c/em\u003e sp. nov. [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e]. A clinicopathological overlap was demonstrated by the cat\u0026rsquo;s subcutaneous nodules and microfilariae, mirroring \u003cem\u003eD. repens\u003c/em\u003e infections and further emphasizing the need for molecular differentiation in endemic regions [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]. In the present case, PCR unequivocally identified \u003cem\u003eDirofilaria\u003c/em\u003e sp. \u003cem\u003e\u0026lsquo;hongkongensis\u0026rsquo;/ Dirofilaria asiatica\u003c/em\u003e sp. nov.. Unlike \u003cem\u003eD. immitis\u003c/em\u003e, \u003cem\u003eDirofilaria asiatica\u003c/em\u003e sp. nov. exhibited tropism for subcutaneous tissues but absence of cardiac involvement, corroborating prior reports [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eSerum chemistry was unremarkable in this case, with marginal hyperglycemia and hyperproteinemia likely due to stress and an ongoing inflammatory process. Neutrophilia and a left shift were present and were interpreted as an active inflammatory response. Other than occasional furball-associated vomiting, no clinical signs were associated with this cutaneous filarioid infection. The presence of mid abdominal reactive lymph nodes was non-specific, however, previous reports of \u003cem\u003eD. repens\u003c/em\u003e and \u003cem\u003eDirofilaria asiatica\u003c/em\u003e sp. nov. have also identified lymphadenopathy [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e, \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]. Reactive lymphadenopathy is not uncommon and could be expected in young cats due to antigenic stimulation with or without gastrointestinal signs such as intermittent vomiting in this case [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e]. Feline immunodeficiency virus (FIV) and feline leukaemia virus (FeLV) tests were negative in our cat, which did not show any obvious co-morbidities or co-infections as it might be seen in other cases of \u003cem\u003eDirofilaria\u003c/em\u003e infection [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe location of the skin lesions in this case were different to a previous study, where all 5 cases including 4 male cats and a female dog presented lesions posteriorly [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Different to previous reports the cat affected here was a female. In this case the skin lesions were anterior, at the left pinna and both elbow regions, which would align with the biting preferences of mosquitos. Interestingly, in the current case the nodules were neither pruritic nor painful, which is in contrast to other cases of cutaneous dirofilariasis where pruritus is commonly seen and may even mimic allergic dermatitis [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e]. Whether this is associated with a reduced movement of \u003cem\u003eDirofilaria asiatica\u003c/em\u003e sp. nov., a modified expression of secretion products or individual factors, needs to be further elucidated.\u003c/p\u003e \u003cp\u003eTreatment complications occurred due to an immunologic reaction demonstrated by the severe respiratory distress following moxidectin and doxycycline application. This has been observed with \u003cem\u003eD. immitis\u003c/em\u003e treatment in dogs linked to a sudden and severe destruction of circulating microfilaria, most frequently seen with the administration of DEC [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. Corticosteroids were critical to mitigating this reaction. Although macrocyclic lactones are significantly safer than DEC in attaining a more gradual reduction in microfilarial clearance, the extremely high microfilarial count in this cat coupled with the use of doxycycline in combination with moxidectin, could have potentiated this reaction. Macrocyclic lactones in combination with doxycycline are known to have significant higher microfilaricidal effects compared to macrocyclic lactones alone [\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]. This warrants caution due to the risk of anaphylaxis. Treating with macrocytic lactones alone and pretreatment with prednisolone or dexamethasone is advisable in future cases.\u003c/p\u003e \u003cp\u003eThis case alerts clinicians for potential acute respiratory crises following the treatment with preventative care drugs such as moxidectin that can be further exacerbated with the administration of concurrent doxycycline. Since antigen tests are not available for this nematode species, microfilarial identification by microscopy might be a sensible consideration.\u003c/p\u003e \u003cp\u003eThere are several limitations of this study, including the design being retrospective in nature and including only one case. Avoidance of infection would be better achieved by having a confirmed vector. While Culicinae vectors such as \u003cem\u003eAedes\u003c/em\u003e spp. are suspected, vector competence studies are needed. Another limitation was the lack of skin biopsies due to owner refusal, although a nematode could be directly isolated from associated lesions. Nevertheless, a granulomatous inflammation seen in previous studies was suspected based on the appearance of lesions but could not be confirmed histopathologically.\u003c/p\u003e \u003cp\u003eThe blood gas analysis was from venous origin in this case, hence the ranges are adjusted. This might explain the oxygenation discrepancy in blood gas PO2 versus sPO2 where the latter is more reliable in venous samples. However, ventilation and acid-base can be estimated and evaluated respectively in venous samples when arterial samples are not available [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eIn this case we emphasize the zoonotic and epidemiological significance of felines as potential reservoirs for \u003cem\u003eDirofilaria asiatica\u003c/em\u003e sp. nov. Given the high microfilarial counts, surveillance and monthly prophylaxis for Hong Kong's feline population are crucial to mitigate risks to the public. The summer onset of symptoms in this case report coincides with the peak of the mosquito activity. Vector dynamics therefore are important and suggest summer months to be a higher risk for transmission [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e, \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e].\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThis case highlights \u003cem\u003eDirofilaria asiatica\u003c/em\u003e sp. nov. as an emerging feline pathogen with zoonotic potential. Affected individuals may only show subcutaneous nodules, mimicking neoplastic, inflammatory or other infectious pathologies, underlining the need for further diagnostics. Treatment requires multimodal therapy to address both parasitosis and immunologic complications. Public health messaging should emphasize on year-round parasite and vector prevention in pets, particularly in endemic regions like Hong Kong.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u0026nbsp;I hereby confirm that the owner and legal guardian of the patient consented to participate in this clinical case and its publication.\u003c/p\u003e\n\u003cp\u003eThe authors confirm that the ethical policies of the journal, as noted on the journal’s author guidelines page, have been adhered to. No ethical approval was required as this is a retrospective study with no original research data but analysing only history records. Informed owner consent was obtained.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors acknowledge the help of clinicians at VMC CityUHK and pathologists at VDL CityUHK for their help with data collection and laboratory interpretation.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTW was the primary clinician of the case and collected clinical data, together with AA provided care and treatment for the cat. AA and TW collected nematode samples. AA drafted the initial manuscript and TW, SH and RT reviewed and edited the manuscript accordingly. YZ carried out the PCR assay and DNA sequencing analysis. RT performed and interpreted results of the Knott’s test and supervised the molecular diagnostics . All authors read and approved the final manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis research received no external funding. This study was funded with a grant from City University of Hong Kong to AA (Project No. 9610596)\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eData are available upon request, with patient identifiers redacted.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll authors consent to this manuscript’s publication.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors disclose no conflicts of interest.\u0026nbsp;\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eColella V, Young ND, Manzanell R, Atapattu U, Sunita B, Sumanam LG, Huggins AV, Koehler, Robin B, Gasser (2025) Dirofilaria Asiatica Sp. Nov.(Spirurida: Onchocercidae)\u0026ndash;Defined Using a Combined Morphological-Molecular Approach. Int J Parasitol\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCapelli G, Genchi C, Baneth G, Bourdeau P, Brianti E, Cardoso Lu\u0026iacute;s, Danesi P, Fuehrer H-P (2018) Alessio Giannelli, and Angela Monica Ionică. Recent Advances on Dirofilaria Repens in Dogs and Humans in Europe. Parasit Vectors 11:1\u0026ndash;21\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYilmaz E, Fritzenwanker M, Pantchev N, Lendner M, Wongkamchai S, Otranto D, Kroidl I, Dennebaum M (2016) Thanh Hoa Le, and Tran Anh Le. The Mitochondrial Genomes of the Zoonotic Canine Filarial Parasites Dirofilaria (Nochtiella) Repens and Candidatus Dirofilaria (Nochtiella) Honkongensis Provide Evidence for Presence of Cryptic Species. PLoS Negl Trop Dis 10(10):e0005028\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePradeep RK, Murikoli Nimisha V, Pakideery J, Johns G, Chandy S, Nair L, Chandrasekhar, Karapparambu G, Ajithkumar CK, Deepa, Anju Varghese (2019) Whether Dirofilaria Repens Parasites from South India Belong to Zoonotic Candidatus Dirofilaria Hongkongensis (Dirofilaria Sp. Hongkongensis)? Infect Genet Evol 67:121\u0026ndash;125\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGowrishankar S, Aravind M, Sastya S, Latha BR, Azhahianambi P, Vairamuthu S, Jayanthy C (2019) Dirofilaria Hongkongensis\u0026ndash;a First Report of Potential Zoonotic Dirofilariosis Infection in Dogs from Tamil Nadu. Vet Parasitol Reg Stud Rep 18:100326\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAtapattu U, Koehler AV, Huggins LG, Wiethoelter A, Traub RJ, Colella V (2023) Dogs Are Reservoir Hosts of the Zoonotic Dirofilaria Sp.\u0026lsquo;Hongkongensis\u0026rsquo; and Potentially of Brugia Sp. Sri Lanka Genotype in Sri Lanka. One Health 17:100625\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHuggins LG, Ugyen Namgyel P, Wangchuk U, Atapattu R, Traub, Colella V (2024) Metabarcoding Using Nanopore Sequencing Enables Identification of Diverse and Zoonotic Vector-Borne Pathogens from Neglected Regions: A Case Study Investigating Dogs from Bhutan. One Health 19:100839\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYilmaz E, Wongkamchai S, Ram\u0026uuml;nke S, Koutsovoulos GD, Blaxter ML, Poppert S, Schaper R (2019) Georg von Samson-Himmelstjerna, and J\u0026uuml;rgen Kr\u0026uuml;cken. High Genetic Diversity in the Dirofilaria Repens Species Complex Revealed by Mitochondrial Genomes of Feline Microfilaria Samples from Narathiwat, Thailand. Transbound Emerg Dis 66(1):389\u0026ndash;399\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eManathunga T, Tse M, Perles L, Beugnet F (2024) Vanessa Barrs, and Domenico Otranto. Zoonotic Dirofilaria Sp.Hongkongensis in Subcutaneous Nodules from Dogs and Cats, Hong Kong Sar. Parasit Vectors 17:469\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKumar A, Sreedhar A, Biswas L, Prabhat S, Suresh P, Asokan A, Tomy RM, Vinod V (2021) Bindu Lakshmanan, and Ajit Nambiar. Candidatus Dirofilaria Hongkongensis Infections in Humans During 2005 to 2020, in Kerala, India. \u003cem\u003eAm J Trop Med Hyg\u003c/em\u003e 104, no. 6 : 2046\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWinkler S, Pollreisz A, Georgopoulos M, Bago-Horvath Z, Auer H (2017) J\u0026uuml;rgen Kr\u0026uuml;cken, Sven Poppert, and Julia Walochnik. Candidatus Dirofilaria Hongkongensis as Causative Agent of Human Ocular Filariosis after Travel to India. Emerg Infect Dis 23(8):1428\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTirakunwichcha S, Sansopha L, Putaporntip C, Jongwutiwes S (2021) Case Report: An Eyelid Nodule Caused by Candidatus Dirofilaria Hongkongensis Diagnosed by Mitochondrial 12s Rrna Sequence. Am J Trop Med Hyg 106(1):199\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSchroeder J, Rothe C, Hoerauf A, Kroidl I, Pfarr K, Marc PH\u0026uuml;bner (2023) First Case of Dirofilaria Hongkongensis Infection in Germany Presenting as a Breast Tumour. J Travel Med 30:8\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCope ED, Nishant Gupta AV, Koehler RB, Gasser, Crowe A (2024) Ocular Dirofilariasis in Migrant from Sri Lanka, Australia. Emerg Infect Dis 30(4):829\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAtwell RB, Boreham PFL (1983) Adverse Drug Reactions in the Treatment of Filarial Parasites: Clinical Reactions to Diethylcarbamazine Therapy in Dogs Infected with Dirofilaria Immitis in Australia. J Small Anim Pract 24(11):695\u0026ndash;701\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCong W, Meng Q-F, Blaga R, Villena I, Zhu X-Q (2016) Toxoplasma Gondii, Dirofilaria Immitis, Feline Immunodeficiency Virus (Fiv), and Feline Leukemia Virus (Felv) Infections in Stray and Pet Cats (Felis Catus) in Northwest China: Co-Infections and Risk Factors. Parasitol Res 115:217\u0026ndash;223\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGenchi C, Venco L, Genchi M (2007) Guideline for the Laboratory Diagnosis of Canine and Feline Dirofilaria Infections\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ede Morais H, Autran, Stephen PD (2017) Advances in Fluid, Electrolyte, and Acid-Base Disorders, an Issue of Veterinary Clinics of North America: Small Animal Practice, vol 47. Elsevier Health Sciences\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTamura J, Itami T, Ishizuka T, Fukui S, Miyoshi K (2015) Tadashi Sano, and Kazuto Yamashita. Central Venous Blood Gas and Acid-Base Status in Conscious Dogs and Cats. J Vet Med Sci 77(7):865\u0026ndash;869\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTo KKW, Samson SY, Wong RWS, Poon, Nigel J, Trendell-Smith, Antonio HY, Ngan JWK, Lam, Tommy HC, Tang (2012) Ah-Kian AhChong, Joshua Chi-Hang Kan, and Kwok-Hung Chan. A Novel Dirofilaria Species Causing Human and Canine Infections in Hong Kong. J Clin Microbiol 50(11):3534\u0026ndash;3541\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eFerri E, Barbuto M, Bain O, Galimberti A, Uni S, Guerrero R, Fert\u0026eacute; H, Bandi C (2009) Coralie Martin, and Maurizio Casiraghi. Integrated Taxonomy: Traditional Approach and DNA Barcoding for the Identification of Filarioid Worms and Related Parasites (Nematoda). Front Zool 6:1\u0026ndash;12\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePampiglione S (2000) and Francesco Rivasi. Human Dirofilariasis Due to Dirofilaria (Nochtiella) Repens: An Update of World Literature from 1995 to 2000. \u003cem\u003eParassitologia\u003c/em\u003e 42, no. 3\u0026ndash;4 : 231\u0026thinsp;\u0026ndash;\u0026thinsp;54\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePerlini M, Bugbee A (2018) Computed Tomographic Appearance of Abdominal Lymph Nodes in Healthy Cats. J Vet Intern Med 32(3):1070\u0026ndash;1076\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGriffin S (2021) Feline Abdominal Ultrasonography: What\u0026rsquo;s Normal? What\u0026rsquo;s Abnormal? Abdominal Lymph Nodes, Peritoneal Cavity and Aorta. J Feline Med Surg 23(9):835\u0026ndash;849\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCarbonara M, Perles L, Venco L, Gabrielli S, Vanessa R, Barrs G, Mir\u0026oacute; E, Papadopoulos (2025) Clara Lima, Emilie Bouhsira, and Gad Baneth. Dirofilaria Spp. Infection in Cats from the Mediterranean Basin: Diagnosis and Epidemiology. Int J Parasitol\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTarello W (2011) Clinical Aspects of Dermatitis Associated with Dirofilaria Repens in Pets: A Review of 100 Canine and 31 Feline Cases (1990\u0026ndash;2010) and a Report of a New Clinic Case Imported from Italy to Dubai. \u003cem\u003eJ Parasitol Res\u003c/em\u003e no. 1 (2011): 578385\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKramer L, Serena Crosara G, Gnudi M, Genchi C, Mangia A, Viglietti, Quintavalla C (2018) Wolbachia, Doxycycline and Macrocyclic Lactones: New Prospects in the Treatment of Canine Heartworm Disease. Vet Parasitol 254:95\u0026ndash;97\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eOtranto D, Capelli G, Genchi C (2009) Changing Distribution Patterns of Canine Vector Borne Diseases in Italy: Leishmaniosis Vs. Dirofilariosis Parasit Vectors 2:1\u0026ndash;8\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"City University of Hong Kong","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Dirofilaria, hongkongensis, asiatica, feline dirofilariasis, zoonosis, moxidectin, subcutaneous nodules, acute pulmonary inflammation, respiratory distress","lastPublishedDoi":"10.21203/rs.3.rs-6921015/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6921015/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eBackground:\u003c/strong\u003e \u003cem\u003eDirofilaria asiatica\u003c/em\u003e sp. nov. (syn. \u003cem\u003eDirofilaria \u003c/em\u003esp. Hong Kong genotype) is an emerging zoonotic filarioid nematode, initially described in human subcutaneous nodules in Hong Kong and later demonstrated in dogs and cats. This report includes the first description of clinical signs, diagnostic findings, including comparative clinicopathology, treatment and associated clinical complications in a feline infection.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods:\u003c/strong\u003e An 18-month-old, spayed female Domestic Shorthair cat presented with subcutaneous nodules and microfilariae in blood smears. Diagnostic work-up included PCR, a quantitative modified Knott’s test, haematology, echocardiography, and abdominal ultrasonography.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults:\u003c/strong\u003e PCR confirmed the presence of \u003cem\u003eDirofilaria asiatica \u003c/em\u003esp. nov. Haematology revealed neutrophilia (11.35 × 10³/µL) with left shift, and hyperproteinaemia (80 g/L). A quantitative modified Knott’s test revealed a microfilaremia of 36,907 per ml. Treatment with oral doxycycline and topical moxidectin triggered an acute onset of respiratory distress, managed with oxygen and dexamethasone. Nodules regressed, and microfilariae cleared by day 70.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusions:\u003c/strong\u003e This case supports that indoor feline pets are susceptible to infection with \u003cem\u003eDirofilaria asiatica \u003c/em\u003esp. nov. In addition,\u003cem\u003e \u003c/em\u003ewe\u003cem\u003e \u003c/em\u003edescribe for the first time, the clinical and clinicopathological findings associated with infection and highlight the risks of immunologic reactions to microfilaricidal therapy. Moxidectin and doxycycline were critical for resolution, but pre-treatment with corticosteroids is recommended. Exorbitant microfilaremia might be a critical feature in cats. The zoonotic potential of this parasite warrants heightened surveillance in endemic regions.\u003c/p\u003e","manuscriptTitle":"Unveiling Dirofilaria asiatica sp. nov.: First Clinical Insights and Treatment Challenges for this Feline Zoonotic Filarioid Infection—A Cautionary Tale","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-06-24 12:31:56","doi":"10.21203/rs.3.rs-6921015/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"83f29b7a-7944-4b22-909a-5c61ff4eb931","owner":[],"postedDate":"June 24th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-06-24T12:31:56+00:00","versionOfRecord":[],"versionCreatedAt":"2025-06-24 12:31:56","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6921015","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6921015","identity":"rs-6921015","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}