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Laura Jackson

Case Studies, Toxicology

Case Study: Ethylene Glycol Toxicity

History

A 2-year-old dog presents with anuric acute kidney injury, renal failure and uraemic encephalopathy. There is no history of access to toxins. Urine analysis reveals SG 1.009, pH 5, protein ++, glucose +, haemoglobin-RBC ++++. On urine sediment examination, there are large numbers of calcium oxalate monohydrate crystals and occasional epithelial cells.

A kidney is collected at post-mortem and submitted in formalin for histopathology.

Examination of the gross specimen in the lab

The anatomic pathologist working in the laboratory grossly examines the kidney. No focal lesions are detected externally or on the cut surfaces after sectioning. Several representative sections are processed for histology.

Histology

 

 

Figure 1 – The overall architecture of the kidney is preserved, with distinct cortex and medulla visible. There is no evidence of hydronephrosis since the pelvis is narrow and not dilated.

 

 

Figure 2 – Even at low magnification, in the cortex, large numbers of crystals are noted within tubules.

 

 

Figure 3 – At high magnification, the crystals are colourless to faintly yellow, arranged in sheaves, rosettes, or prisms. When viewed under polarised light, the crystals are birefringent. Observe the relatively healthy glomerulus on the right in this image.

 

 

Figure 4 – Large numbers of tubules in the cortex show necrosis, in the form of hypereosinophilic (red staining) cells, fragmentation and lysis of the nuclei. Compare the two necrotic tubules (red arrow) adjacent to a relatively healthy tubule (green arrow).

 

What is your histological diagnosis?

Acute tubular necrosis with intratubular calcium oxalate crystals. This is consistent with ethylene glycol toxicity.

Ethylene glycol toxicity

Although there is no known history of exposure, it is highly likely that this dog has ingested ethylene glycol. Ethylene glycol is the main ingredient in antifreeze. Low numbers of calcium oxalate crystals can be observed histologically in kidneys in chronic renal failure of various causes, but the large number of calcium oxalate crystals observed here together with the observation of acute tubular necrosis is virtually pathognomonic for ethylene glycol toxicity.

Antemortem diagnosis of ethylene glycol toxicity requires a combination of history of exposure (if known), clinical signs, blood biochemistry, urinalysis and potentially toxicology testing. Common clinical signs are ataxia, depression and vomiting. Common pathologic findings are high anion gap metabolic acidosis (due to unmeasured anions, which are the metabolites – oxalic acid and others), hyperglycaemia, azotaemia and calcium oxalate monohydrate crystals in the urine (see figure 5). Less common/specific findings are hyperphosphataemia, hypocalcaemia, poorly concentrated urine, glycosuria, proteinuria.

 

 

Figure 5 – This is the usual appearance of calcium oxalate monohydrate crystals in urine in cases of ethylene glycol toxicity, and when detected they should be considered pathologic. The crystals have a flat picket fence board-like shape. The crystals may also form spindles and dumbbell shapes. Crystalluria can be observed within 3 hours of ethylene glycol ingestion in cats and within 6 hours in dogs.

 

Toxicology testing at VPG

VPG offers toxicology testing for a wide range of toxic compounds, including ethylene glycol using a technique called gas chromatography-mass spectrometry (GC-MS). The test is useful in the clinical setting to confirm clinical suspicion of ethylene glycol toxicity. Blood and urine are recommended samples and the test can be performed on as little as 5 μL of either. For more information about toxicology at VPG please click on this link to visit our toxicology page https://thevpg.co.uk/toxicology/

 

References:

Amoroso L, Cocumelli C, Bruni G, Brozzi A, Tancredi F, Grifoni G, Mastromattei A, Meoli R, Di Guardo G, Eleni C. Ethylene glycol toxicity: a retrospective pathological study in cats. Vet Ital. 2017 Sep 30;53(3):251-254. doi: 10.12834/VetIt.1159.6409.2. PMID: 29152707.

Iqbal A, Glagola JJ, Nappe TM. Ethylene Glycol Toxicity. 2022 Sep 26. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2025 Jan–. PMID: 30725694.

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Case Studies, Cytology

Eucoleus (Capillaria) Aerophila in a Corsac Fox

History

Toulouse – 1yr 6 month Male neutered Corsac Fox.
History of chronic coughing and weight loss with intermittent diarrhoea. Toulouse had previously been treated with 4 x weekly milbemycin. Chronic cough persisted. Previous faecal testing negative.

Xrays, blood profile and BAL performed. Bloods showed increase in ALT, BUN and Phos with evidence for haemoconcentration.

 

 

Figure 1:
X-rays revealed a diffuse bronchial pattern, possibly more alveolar pattern in cranial lung fields.

 

Clinical Approach

Cytology reveals marked, predominantly eosinophilic inflammation with the presence of parasitic ova. Ova were ellipsoid and generally 60-65 µm in length. Occasionally the ova had distinct bipolar caps. Morphology raised significant suspicion for Eucoleus (Capillaria) spp. Given the location of the inflammation in the lower airways, presence of ova in the airways and morphology of the ova – Eucoleus (Capillaria) aerophila was suspected.

 

 

Figure 2: BAL cytology preparation showing high numbers of inflammatory cells which are mainly eosinophils with lesser numbers of neutrophils and occasional macrophages.

 

 

Figure 3: BAL cytology preparation showing marked predominantly eosinophilic inflammation and embedded helminth ova.

 

 

Figure 4: Eucoleus (Capillaria) aerophila egg in BAL from this case (left), and from a dog right (5)

 

Samples sent for PCR confirmed the presence of Eucoleus (Capillaria) aerophila. Culture also revealed a pure profuse growth of Bacillus which is generally considered of low pathogenicity. However, in the context of a small neutrophilic component to inflammation, absence of oropharyngeal contamination and noting this is a pure growth, the finding may be significant in this case.

Eucoleus (Capillaria) aerophila is a parasitic helminth found in the lower respiratory tract, primarily affecting wild carnivores however, it can also infect dogs and cats. Infections are often asymptomatic in otherwise healthy individuals but in young animals or immunosuppressed individuals, the disease can be more severe and can lead to cough, nasal discharge, dyspnoea, anorexia and debilitation.

 

 

Figure 5: Adult female Eucoleus aerophilus on the mucosa of the opened trachea of a fox  (1)

 

Life cycle

Eggs are released from infected animals in faeces and remain viable in the environment for up to 1 year. Earthworms may act as paratenic hosts. Infection is faecal oral or via ingestion of infected earthworms and ingested ova hatch in the small intestine, penetrate the mucosa and migrate via the blood or lymphatics to the lungs. Here they enter the alveoli and migrate up to the trachea, bronchi and bronchioles where they live beneath the epithelium and release eggs to be coughed up and swallowed (2).

 

 

Figure 6: Eucoleus (Capillaria) aerophile life cycle. Source VPG

 

Diagnosis

Diagnosis is either by detection of ova in BAL or sputum or in faecal analysis. Larvae are rarely seen allowing differentiation from other respiratory helminth infections. Main differentials for the ova are Eucoleus boehmi and if found in faeces, Trichuris vulpis (whipworm). Other lungworm species are generally seen as larvae rather than ova aiding identification.  Eucolues boehmi, Truchuris vulpis and Eucoleus aerophilia all have very similar appearing ova but location is important. Eucoleus boehmi is mainly nasal cavity and paranasal sinuses and Trichuris vulpis is an intestinal parasite.

 

Prevalence

Very wide geographical distribution virtually worldwide and across Europe and it primarily infects wild carnivores (foxes) but can infect domestic carnivores. It is also a potential zoonosis with occasional cases reported in humans (3). In UK, High prevalence in lungs of red fox population, average of 31%. No current studies to determine prevalence in UK cat and dog population but worldwide an average of 3% in dogs and up to 8% in cats (4).

 

(1) Lalošević, V., Lalošević, D., Čapo, I., Simin, V., Galfi, A. & Traversa, D. – Lalošević, V., Lalošević, D., Čapo, I., Simin, V., Galfi, A. & Traversa, D. (2013). High infection rate of zoonotic Eucoleus aerophilus infection in foxes from Serbia. Parasite 20: 3. doi:10.1051/parasite/2012003

(2) Traversa D, Di Cesare A, Lia RP, Castagna G, Meloni S, Heine J, et al. New insights into morphological and biological features of Capillaria aerophila (Trichocephalida, Trichuridae) Parasitol Res. 2011;109:S97–104. doi: 10.1007/s00436-011-2406-4. [DOI] [PubMed] [Google Scholar]

(3) Lalosević D, Lalosević V, Klem I, Stanojev-Jovanović D, Pozio E. Pulmonary capillariasis miming bronchial carcinoma. Am J Trop Med Hyg. 2008;78:14–16. doi: 10.4269/ajtmh.2008.78.14. [DOI] [PubMed] [Google Scholar]

(4) Samorek-Pieróg M, Cencek T, Łabuć E, Pac-Sosińska M, Pieróg M, Korpysa-Dzirba W, Bełcik A, Bilska-Zając E, Karamon J. Occurrence of Eucoleus aerophilus in wild and domestic animals: a systematic review and meta-analysis. Parasit Vectors. 2023 Jul 20;16(1):245. doi: 10.1186/s13071-023-05830-0. PMID: 37475031; PMCID: PMC10360280.

(5) Di Cesare, Angela & Castagna, Giuseppe & Otranto, Domenico & Meloni, Silvana & Milillo, Piermarino & Latrofa, Maria & Paoletti, Barbara & Bartolini, Roberto & Traversa, Donato. (2012). Molecular Detection of Capillaria aerophila, an Agent of Canine and Feline Pulmonary Capillariosis. Journal of clinical microbiology. 50. 1958-63. 10.1128/JCM.00103-12.

 

 

Thank you to Holly Faulkner at Ark Veterinary centre for collaborating on this case.

 

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News

VPG and Zoetis unite to advance veterinary diagnostics

We’re thrilled to announce that Veterinary Pathology Group (VPG) is now part of Zoetis, the world’s leading animal health company.

This partnership brings together VPG’s trusted, expert veterinary diagnostics with Zoetis’ global innovation and resources, a perfect match built on shared purpose and values. Together, we’re committed to delivering faster, more accurate, and comprehensive diagnostic solutions that empower veterinarians across the UK and Ireland to provide the best care possible.

With this collaboration, you’ll benefit from enhanced testing capabilities, access to cutting-edge technologies, and stronger local support, all while maintaining the personal service and “right-first-time” results you’ve come to expect from VPG.

Welcome to the next chapter of veterinary diagnostics excellence.

Read the press release here

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Case Studies, Diagnostics

MicroRNAs: Could Tiny Molecules Make a Big Difference in Diagnosing Feline HCM?

Hypertrophic cardiomyopathy (HCM) is the most common feline heart condition, with around 15% of cats, and nearly 1 in 3 cats over 9 years of age, being affected1. The disease is characterised by thickening of the left ventricular myocardium, resulting in impaired relaxation, diastolic dysfunction and, eventually, congestive heart failure or thromboembolic events.

Despite its prevalence and decades of research, feline HCM remains a challenging disease to diagnose and predicting how it will manifest in any individual cat can feel like educated guesswork at best.

 

The silent disease

Cats are, without doubt, masters of disguise. HCM often has a long subclinical phase, during which affected cats appear clinically well. For many, the first or only sign of disease may be a sudden onset of heart failure or aortic thromboembolism.

Even on clinical examination, HCM can be elusive. Some cats have a systolic murmur, gallop rhythm, or arrhythmia—but many do not. And to complicate matters further, not all murmurs in cats indicate structural heart disease.2

 

Diagnostic challenges

So how do we diagnose feline HCM with confidence?

Echocardiography remains the gold standard, but it’s far from perfect. Interobserver reliability of cardiac measurements remains a challenge, and both cost and specialist availability may limit how accessible echocardiography is for some owners.

Several “copy-cats” (such as hypertension, hyperthyroidism, and transient myocardial thickening) can mimic HCM on echo, and even minor technical deviations—like including moderator bands, or the endocardium in measurements, or obtaining slightly oblique imaging planes —can lead to overdiagnosis.

Then there’s the “cats will be cats” factor: tachycardia and hypertension, and movement artefacts caused by feline wriggliness and their reluctance to be examined, all affect reliability. For both feline and human participants, the process is rarely stress-free, and for affected cats, especially, is not without risk.

When we consider that 60–70% of cats with HCM will remain asymptomatic for life3, the benefits of investigation and frequency of follow-up must be carefully balanced against the cost, stress, and risks involved, and care taken not to over-diagnose.

 

Wouldn’t it be great if we had an easier test?

That’s exactly why there’s growing interest in biomarkers for HCM.

A biomarker capable of identifying disease before clinical signs or advanced structural change occur, could transform feline cardiology. A simple, accessible and cost-effective blood test to identify at-risk or affected cats would enhance screening programmes, refine case selection for referral, and support more confident diagnoses.

Commercially available markers such as NT-proBNP and cardiac troponin I (cTnI) are already in use and are valuable in distinguishing cardiac from non-cardiac causes of respiratory distress4-8. However, since they only rise after cardiac stretch, pressure overload, or myocyte injury has occurred, they are best applied to those with moderate to severe clinical disease, and their ability to detect subclinical HCM, is limited.

 

Enter microRNAs?

MicroRNAs (miRNAs) are tiny, non-coding RNA molecules that regulate gene expression – acting like molecular “project managers”, switching cellular pathways on and off. Many of these pathways are central to cardiac hypertrophy, fibrosis, and remodelling.

Changes in microRNA expression are associated with disease. Since these changes are seen before tissue damage has even occurred, miRNAs have fantastic potential as markers of early or subclinical disease.

It’s no surprise, therefore, that miRNAs are receiving a lot of attention in the scientific community. In human cardiology, miRNAs have already shown great promise as diagnostic and prognostic biomarkers for cardiomyopathies, including HCM. And now, thanks to pioneering team of veterinarians and scientists at MI:RNA, the same is now true for cats.

 

Cutting edge science

MI:RNA have combined the power of cutting-edge molecular science with AI-driven analysis to reveal distinct miRNA expression patterns in cats with HCM compared to healthy controls.

This raises an exciting possibility: a simple blood test capable of identifying cats with HCM—and perhaps even detecting disease in its earliest, subclinical phase, before myocyte damage occurs.

So, while we won’t be putting down our stethoscopes or scanners any time soon, miRNA technology could soon add a powerful new tool to our diagnostic toolbox—offering molecular-level insight into what’s happening inside the feline heart.

If microRNAs can reveal what echocardiography and current biomarkers miss, they could transform how we detect, monitor, and manage feline heart disease. From the consultation room to the laboratory, these tiny molecules may well have a huge impact on feline health—and on how we care for our patients.

 

Excited to find out more?

Join us and the MI:RNA team at the London Vet Show to learn more about how this groundbreaking technology could change the future of feline cardiology. Stop by our stand (P45), meet the team behind the research, and discover how big a difference these tiny molecules might make.

 

References:

  1. Payne JR, Brodbelt DC, Luis Fuentes V. Cardiomyopathy prevalence in 780 apparently healthy cats in rehoming centres (the CatScan study). J Vet Cardiol 2015; 17 Suppl 1: 5244-5257.
  2. Paige CF, Abbott JA, Elvinger F, Pyle RL. Prevalence of cardiomyopathy in apparently healthy cats. J Am Vet Med Assoc. 2009 Jun 1;234(11):1398-403. doi: 10.2460/javma.234.11.1398. PMID: 19480619.
  3. Fox PR, Keene BW, Lamb K,et al. International collaborative study to assess cardiovascular risk and evaluate long-term health in cats with preclinical hypertrophic cardiomyopathy and apparently healthy cats: The REVEAL Study. J Vet Intern Med. 2018 May;32(3):930-943. doi: 10.1111/jvim.15122. Epub 2018 Apr 16. Erratum in: J Vet Intern Med. 2018 Nov;32(6):2310. doi: 10.1111/jvim.15285. PMID: 29660848; PMCID: PMC5980443.
  4. Ward JL, Lisciandro GR, Ware WA, et al. Evaluation of point-of-care thoracic ultrasound and NT-proBNP for the diagnosis of congestive heart failure in cats with respiratory distress. J Vet Intern Med 2018; 32: 1530-1540
  5. Wurtinger G, Henrich E, Hildebrandt D, et al. Assessment of a bedside test for N-terminal pro B-type natriuretic peptide (NT-proBNP) to differentiate cardiac from non-cardiac causes of pleural effusion in cats. BMC Vet Res 2017; 13:394.
  6. Connolly DJ, Soares Magalhaes RJ, Fuentes VL, et al. Assessment of the diagnostic accuracy of circulating natriuretic peptide concentrations to distinguish between cats with cardiac non-cardiac causes of respiratory distress. J Vet Cardiol 2009; 11: S41-S50.
  7. Wells SM, Shofer FS, Walters PC, et al. Evaluation of blood cardiac troponin I concentrations obtained with cage-side analyzer to differentiate cats with cardiac and noncardiac causes of dyspnea. J AM Vet Med Assoc 2014; 244: 425-430.
  8. Connolly DJ, Brodbelt DC, Copeland H, Collins S, Fuentes VL. Assessment of the diagnostic accuracy of cardiac troponin I concentration to distinguish between cats with cardiac non-cardiac causes of respiratory distress. J Vet Cardiol 2009; 11: 71-78.
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Case Studies, Histopathology

Case study: Mycobacterial Lymphadenitis in a Cat

A 12 year old female neutered domestic short hair cat presented with lymph node enlargement in the submandibular area.  Histological examination revealed a granulomatous lymphadenitis with necrosis.

 

 

Figure 1: Large areas of necrosis are present in the lymph node (stars). Magnification x 7.5. H&E stain. Scale bar =2.5 mm.

 

 

Figure 2: Granulomatous inflammation within the lymph node. Magnification x 200. H&E stain. Scale bar =100 µm

Figure 3: Acid-fast (pink) bacilli within the lymph node (arrows), consistent with mycobacteria. Magnification x 400. Ziehl-Neelsen stain. Scale bar = 50 µm.

 

PCR analysis revealed a mycobacterium of the Mycobacterium avium complex (MAC). Mycobacteria in this group are non-tuberculous mycobacteria. Typically, these are saprophytic and infections may occur via contaminated wounds.

 

Clinical signs of mycobacterial infections in cats:

The most common presentation of mycobacterial infection in cats is skin lesions. Cats may present with single or multiple nodular masses, non-healing wounds and granulomatous panniculitis, as well as regional lymph node enlargement. In some cases, such as in the case presented here, lymph node enlargement may be the only or most obvious clinical sign.

Most commonly nodular skin lesions are found in areas predisposed to bites during hunting, such as the face and extremities. Granulomatous panniculitis is most commonly found in the inguinal area, flanks and tail base.

Systemic dissemination may occur in particular to lungs, liver and spleen. An alimentary form has been associated with ingestion of M. bovis contaminated milk or pet food.

 

Diagnosis:

  • Ziehl-Neelsen (ZN) stain on cytology samples or histological sections may detect the presence of acid-fast rods. Mycobacteria may be present in low numbers and ZN stains may therefore be false negative in some cases.
  • Mycobacterial culture – please note that not all mycobacteria can be cultured and more than 50% of ZN positive lesions in cats were found to be negative by culture in one study (Gunn-Moore et al. 2011).
  • PCR analysis may be performed on non-fixed and formalin-fixed samples. Please note that formalin-fixation may inhibit PCR analysis.

If a mycobacterial infection is a potential differential diagnosis, and tissue samples are taken, preparation of an impression smear should be considered.  Ziehl-Neelsen staining of the impression smear will be performed, and if found positive for mycobacteria, the sample can be prioritised for specialist culture and PCR.

Tissue samples may be divided, with one third submitted in formalin for histopathological analysis and the remainder frozen in 2 separate containers for subsequent culture and/or PCR. This may not be an option for small samples.

  • The interferon-gamma release assay uses peripheral blood mononuclear cells obtained from a blood sample of the patient. The cells are stimulated with synthetic peptides representing mycobacterial epitopes. In cats with a mycobacterial infection, this will lead to a release of interferon-gamma, which is measured in an ELISA assay.

Please contact us prior to sampling, if you wish to discuss sample handling, sample submission and test options.

 

 

Zoonotic risk:

Most mycobacteria have zoonotic potential. If the presence of mycobacteria is confirmed by ZN stain, stringent precautions should be taken, until further characterisation of the causative agent has been performed. Cats with ulcerated wounds or draining tracts pose an increased risk of exposure. If samples are submitted to diagnostic laboratories, any suspicion of a mycobacterial infection should be clearly stated on the submission form to prevent exposure of laboratory staff.

Mycobacteria of the M. tuberculosis complex (including M. tuberculosis, M. bovis and M. microti) pose the most significant risk to all in contact persons, not only those that are immunocompromised.

Non-tuberculous mycobacteria cause opportunistic infections in immunocompromised human hosts, including those with underlying disease, undergoing chemotherapy, pregnant women and the elderly. M. avium (subspecies avium) poses the greatest risk of those NTM found in cats.

The identification of M. bovis in cats (and any other non-human mammals) is notifiable to the APHA.

Reference:
Gunn-Moore DA, McFarland SE, Brewer JI, Crawshaw TR, Clifton-Hadley RS, Kovalik M, Shaw DJ. (2011). Mycobacterial disease in cats in Great Britain: I. Culture results, geographical distribution and clinical presentation of 339 cases. J Feline Med Surg. 13(12):934-44.

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Histopathology

Supporting Vets with Post Mortem Histopathology

At VPG, we understand that post mortem submissions can be an essential part of veterinary diagnostics, helping to provide answers in difficult cases and contributing to wider clinical understanding. Our Histopathology team is here to support you with a straightforward submission process and accurate interpretation of findings.

When to Submit Tissues or Cadavers

In some circumstances, transporting a full cadaver to our laboratories is not practical. To support you, we provide a flexible approach:

  • Tissue submissions (PM1 – PMC 6+ tissues): for samples taken during a full or partial post mortem performed in practice.

  • Small cadaver submissions (HPMC): we can accept cadavers no greater than 10 cm in length (e.g. fish, mice or invertebrates) for necropsy on site at our Bristol laboratory. These must be fully submersed in formalin before submission.

Both tissue and cadaver submissions can be sent through the same service you already use for histopathology work.

What Information to Include

To ensure our pathologists can provide the most accurate and valuable interpretation, we ask that you include as much relevant information as possible on the submission form. This should cover:

  • A detailed clinical history, including presenting symptoms, observed lesions, and any treatment or medication given.

  • If euthanasia was performed, the method used.

  • Any recent changes in environment or housing.

  • Whether other animals are affected.

  • A short summary of the post mortem examination findings.

  • Images (ante-mortem or post-mortem), or a complete post mortem report with relevant laboratory or imaging results.

  • Clinical differential diagnoses you are considering.

  • Please also note if there is any suspicion of mycobacteriosis or other potentially zoonotic disease.

This level of detail allows our laboratory technicians and pathologists to incorporate clinical and post mortem data into their interpretation, strengthening the diagnostic outcome.

New Post Mortem Form

We have introduced a new post mortem submission form, now available to download from our website. This ensures that all essential details are recorded consistently and can be incorporated into your case report.

📄 [Download the new Post Mortem form here]

If you are unsure about how best to proceed with a submission, or how a set of samples will be priced, our Histopathology team is happy to advise.

📧 [email protected]
📞 0117 951 1283

Our anatomical pathologists and laboratory staff are here to support you every step of the way.

💡 Excellence. Everywhere.

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Webinar

Webinar: Could microRNAs transform the future of cardiac diagnosis?

VPG, in partnership with MI:RNA, is evaluating a cutting-edge microRNA-based diagnostic for veterinary cardiology.

In this session, you’ll hear from leading experts on how microRNAs could complement existing diagnostics such as NT-proBNP and Troponin I, and what this might mean for decision-making in practice.

What You’ll Learn 

  • Discover why microRNAs react early in cardiac disease
  • Understand how this testing could enable earlier detection and accurate staging in dogs and cats
  • Explore how microRNA diagnostics could integrate seamlessly into your workflow

Who Should Watch? 

  • Veterinary cardiologists 
  • Internal medicine specialists with a cardiology interest 
  • General practitioners managing cardiac cases 

Simply fill out the short form below to watch the recording of the presentation:


 

 

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Case Studies, Histopathology

CASE STUDY: Pneumocystosis in a Dog

Clinical history:

A 7-year-old, male neutered, Cavalier King Charles Spaniel presented with a four-week history of progressively worsening tachypnoea, developing into dyspnoea. CT imaging under sedation revealed generalised ground glass appearance throughout the lung fields, deemed to be consistent with non-cardiogenic oedema. The dog was treated with trimethoprim-potentiated sulfonamides and oxygen supplementation, but deteriorated and sadly died from cardiopulmonary arrest. A post-mortem sample of lung was submitted to VPG for histopathology.

Histopathology:

The lung tissue was diffusely consolidated. The alveoli were lined by hyperplastic type II pneumocytes and the interstitium was fibrotic. Alveolar spaces were flooded with oedema, fibrin or surfactant-type material, and large numbers of inflammatory cells, mainly macrophages and neutrophils. There were accumulations of foamy eosinophilic material visible in some areas within the alveolar spaces. When Grocott-Gomori methenamine silver (GMS) stain was applied, the foamy eosinophilic material was shown to be composed of numerous 3-4 micron diameter fungal organisms. This appearance was diagnostic for Pneumocystis sp. infection (pneumocystosis).

 

 

Figure 1.

This low-power image (H&E) shows lung consolidation. Alveolar spaces are no longer filled with air (should show as clear space). The bronchioles are still visible containing clear space though.

 

 

Figure 2.

This high-power view (H&E) shows the alveoli infiltrated with inflammatory cells (arrow head), multifocally filled with foamy eosinophilic material (horizontal arrows), and lined by type II pneumocytes (vertical arrows).

 

 

Figure 3.

With GMS silver stain, in the areas of foamy material, there are numerous round organisms (arrow heads), typical of Pneumocystis sp. Note that most of the foamy material is non-staining, which corresponds to the trophozoites. The silver stain only stains the cyst walls.

 

Histological diagnosis:

Interstitial pneumonia with intra-alveolar Pneumocystis sp

 

More information on Pneumocystis:

Clinical cases of canine pneumocystosis are rare. The clinical history of gradual onset respiratory disease, usually longer than 4 weeks, is common. The other most common clinical signs are exercise intolerance, cough, and weight loss despite normal feed intake. It is thought that underlying immune impairment is required for the causative agent, Pneumocystis carinii, to cause clinical disease, and this may be inherited or induced by immunosuppressive therapy. The Cavalier King Charles Spaniel is a predisposed breed for this infection owing to a suspected underlying immune deficiency. There is a known common variable immunodeficiency inherited in the Miniature Dachshund, making this the other most commonly affected dog breed. Foals with severe combined immunodeficiency (SCID) are also predisposed. At 7 years of age, this spaniel was older than most affected dogs (median 1 yr).

 

Pneumocystis carinii is a yeast-like fungus that is highly adapted to infect type I pneumocytes that line the alveoli in health. The typical histological picture is the presence of foamy material filling alveoli. This material is composed of numerous fungal cysts and trophozoites. The individual organisms are difficult to appreciate without a special silver stain (GMS) and can be overlooked if they are present in low numbers and only standard H&E stain is used.

 

This patient unfortunately succumbed to pneumocystosis. Dogs can recover but early treatment is generally needed to achieve this. Obtaining an antemortem diagnosis can be problematic, but bronchio-alveolar lavage fluid cytology is an effective and noninvasive method.

 

 

Alex Civello DipACVP FRCPath MRCVS

Board Certified Anatomic Pathologist

 

References:

1. Weissenbacher-Lang C, Fuchs-Baumgartinger A, Guija-De-Arespacochaga A, Klang A, Weissenböck H, Künzel F. Pneumocystosis in dogs: meta-analysis of 43 published cases including clinical signs, diagnostic procedures, and treatment. J Vet Diagn Invest. 2018 Jan;30(1):26-35. doi: 10.1177/1040638717742429.

2. Lobetti R. Common variable immunodeficiency in miniature dachshunds affected with Pneumonocystis carinii pneumonia. J Vet Diagn Invest. 2000 Jan;12(1):39-45. doi: 10.1177/104063870001200107.

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Pathology

Smart Diagnostics Part 2: AI to Improve Prognostic Accuracy in Ki67 staining of Canine Cutaneous Mast Cell Tumours

Cutaneous mast cell tumours (MCTs) are amongst the most common malignant skin tumours in dogs. Their biological behaviour can be variable and as a consequence the prognosis can be unpredictable. Whilst well-established grading systems have been developed and are routinely used to assess these tumours, namely the ‘two-tier’ (Kiupel et al 20I0) and older ‘three-tier’ (Patnaik et al 1984) grading systems, these do not correctly predict clinical outcomes in all cases. A subset of MCTs, identified as low/intermediate grade examples (Figure 1) using these grading schemes, do not act as indicated by the histologic grading. This subset of tumours can display more aggressive behaviour, including recurrence, nodal metastasis (Figure 2) and distant metastasis, ultimately resulting in euthanasia or death of some of the affected patients.

 

 

Figure 1. Typical histological appearance of a Grade II/low grade mast cell tumour,  exhibiting well-differentiated mast cell amongst a fibrous stroma, with accompanying eosinophilic inflammation.

 

 

Figure 2. Image of a nodal metastasis of a well-differentiated MCT. Neoplastic mast cells are seen to fill sinuses and efface the nodal architecture.

 

Multiple additional prognostic markers have been investigated to more precisely predict the prognosis of canine cutaneous MCTs. These tests, such as the Ki67 index, may help to identify those tumours likely to be in the aggressive subset of apparently histologically ‘low/intermediate’ grade MCTs.

 

What is the Ki67 index?

The Ki67 index is the most commonly requested prognostic marker for mast cell tumours and melanocytic neoplasms. It essentially measures the proportion of tumour cells that are actively dividing in a given snap shot of time. Elevated expression of Ki67 in mast cell tumours has been shown to be statistically associated with more aggressive tumour behaviour and poorer patient outcomes (Scase et al 2006; Maglennon et al 2008). However, formulation of this index typically requires manual counting of large numbers of individual mast cells under the microscope – a method that is inherently time-consuming. Other cells, including stromal cells and inflammatory cells, can also display positivity for this marker, adding to the complexity of interpretation of the test. At VPG, Ki67 stained slides are interpreted by anatomical pathologists experienced with the methodology and analysis. For inexperienced pathologists interpreting Ki67 stained sections, the test can be time consuming, and potentially susceptible to interobserver and intraobserver variability.

To address these limitations, a recent VPG study explored the use of artificial intelligence (AI)-assisted analysis to automate Ki67 cell counting in canine cutaneous MCTs. The goal was to increase efficiency, reduce interobserver and intraobserver variability, and ultimately support more accurate prognostication for clinical practice.

 

Development of the AI-Assisted Tool

The study utilised QuPath, a platform for digital pathology image analysis (Bankhead et al 2017). VPG pathologists trained a machine learning algorithm to identify and distinguish between Ki67-positive and Ki67-negative mast cells, as well as other neighbouring cells including inflammatory leucocytes, which may complicate the interpretation of the immunohistochemical staining (Figure 3).

Training was conducted using a dataset of 244 representative MCT images, allowing the machine to learn the relevant digital and microscopic features of positive and negative staining (this could include, for example, staining intensity, ‘roundness’ of the cells and other colour/shape parameters). To evaluate its performance, the trained AI system was tested on a separate set of 77 images, which were also reviewed manually by three board-certified veterinary histopathologists. This allowed for direct comparison between ‘machine’ and human in terms of accuracy and reproducibility.

 

 

Figure 3. Example of a Ki67-stained mast cell tumour (left). Dark brown round cells are considered to be positive mast cells in this image. A machine-interpreted image (right) highlights positive mast cells (red outline) and negative mast cells (blue outline). Other cells include stromal cells and eosinophils.

 

Key Findings

The AI-assisted method demonstrates good levels of accuracy, closely matching the results obtained by manual cell counting. Importantly, the automated system significantly reduces the time required to complete the analysis. It also allows for greater consistency across samples, helping to minimise interobserver variability. Remarkably, the time required to complete an image analysis is now less than 1 second, rather than the average time of a ‘manual count’, which is often several minutes.

These findings demonstrate that AI can be a practical advancement in the diagnostic workflow of veterinary pathology providers, particularly for specific tasks that are labour-intensive and potentially subject to interobserver and intraobserver variation. While AI does not aim to replace the expertise of a trained pathologist, it can serve as a valuable aid, particularly in high-throughput and laborious settings, or when standardisation is a priority. Incorporating this AI tool into diagnostic pathology offers benefits for both pathologists and clinicians; More efficient and reproducible Ki67 scoring allows for improved prognostic assessments.

 

Conclusion

The study aimed to validate the use of AI as a tool to assist with Ki67 cell counting in canine cutaneous mast cell tumours. Whilst traditional manual cell counting to attain a Ki67-index provides clinically useful prognostic information, AI-assisted counts enhance the process with more rapid and reproducible counts, reducing the physical ‘burden’ of a manual cell count. Whilst offering significant improvements in efficiency, all AI-assisted counts are verified carefully by a board-certified pathologist to ensure their accuracy.

 

References

Bankhead, P., Loughrey, M.B., Fernández, J.A., Dombrowski, Y., McArt, D.G., Dunne, P.D., McQuaid, S., Gray, R.T., Murray, L.J., Coleman, H.G. and James, J.A., 2017. QuPath: Open source software for digital pathology image analysis. Scientific Reports, 7(1), pp.1-7.

Kiupel M, Webster JD, Bailey KL, et al. Proposal of a 2-Tier Histologic Grading System for Canine Cutaneous Mast Cell Tumors to More Accurately Predict Biological Behavior. Veterinary Pathology. 2010;48(1):147-155.

Patnaik AK, Ehler WJ, MacEwen EG. Canine cutaneous mast cell tumor: morphologic grading and survival time in 83 dogs. Vet Pathol. 1984 Sep;21(5):469-74.

Maglennon GA, Murphy S, Adams V, Miller J, Smith K, Blunden A, Scase TJ. Association of Ki67 index with prognosis for intermediate-grade canine cutaneous mast cell tumours. Vet Comp Oncol. 2008 Dec;6(4):268-74. doi: 10.1111/j.1476-5829.2008.00168.x. PMID: 19178685.

Scase TJ, Edwards D, Miller J, Henley W, Smith K, Blunden A, Murphy S. Canine mast cell tumors: correlation of apoptosis and proliferation markers with prognosis. J Vet Intern Med. 2006 Jan-Feb;20(1):151-8. doi: 10.1892/0891-6640(2006)20[151:cmctco]2.0.co;2. PMID: 16496935.

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Diagnostics

How Targeted Microbiome Support Is Transforming Dog Health

Introduction 

Retesting is a vital part of how long-term gut health is supported through the BIOME9 supplement plan. Just like in human medicine, adjusting and optimising based on results is key to sustainable change. With over 100 individual retests now completed, BIOME9 has gathered robust evidence showing how precision microbiome support, guided by data and tailored supplements, can lead to measurable, wide-ranging improvements in dogs.

The Power of Retesting and Adjustment

Microbiome modulation isn’t a one-off fix. It’s a journey. BIOME9 recommends retesting after dogs have been on the supplement plan for 8 to 12 weeks. This gives both practitioners and owners a clear window into what’s changing and where further support might be needed.

In 101 individual retests, 83% of dogs showed significant improvements across core gut health scores and functions. These changes were not only reflected in lab data but also matched the improvements reported by owners: better digestion, calmer behaviour, fewer symptoms, and increased vitality.

Microbial Community Shifts: Diversity, Stability, and Beneficial Bacteria

Across the retests, the average increase in microbial diversity was striking. Diversity more than doubled, with an average of 167 additional bacterial species detected post-supplementation. Balance and stability—two hallmarks of a resilient microbiome—also improved by 39% on average.

Additionally, 68% of dogs saw an increase in all twelve core commensal (beneficial) bacteria. The most notable increases were seen in Megamonas, Fusobacterium, and Faecalibacterium—species associated with healthy digestion, anti-inflammatory functions, and immune regulation. This shift suggests that targeted supplementation supports broad microbial health and helps re-establish key populations that may have been lost or suppressed.

 

Digestive Function: Precision Nutrition at Work

Improving digestive efficiency is one of the core aims of microbiome modulation. After completing the supplement plan and retesting, dogs showed an average 12% improvement in digestive function. These gains weren’t just about symptom relief—they reflected better nutrient absorption and energy extraction from food:

  • +6% protein digestion

  • +9% carbohydrate digestion

  • +11% fibre digestion

  • +13% lipid digestion

  • +20% increase in vitamin and mineral absorption

By optimising the existing diet rather than replacing it, the BIOME9 plan supports precision nutrition, helping each dog get more out of the food it already eats.

Beyond Digestion: Whole-Body Benefits

While gut health is the foundation, the benefits of a balanced microbiome extend across multiple systems. In 91% of retests, dogs showed improved health markers beyond digestion.

For example, breath odour (a common complaint) improved by 28%, reflecting internal metabolic improvements rather than superficial masking. Neurological health scores rose by 12%, supporting what broader data has shown: nearly one-third of dogs with gut issues also experience behavioural symptoms.

Perhaps most striking was the impact on immune-related functions. Retests showed:

  • +8% improvement in coat and skin health

  • +16% increase in immune and cardiovascular markers

  • +17% reduction in markers of gut inflammation

  • +27% improvement in joints and mobility

These findings reinforce the microbiome’s central role in systemic health and highlight the importance of personalised, science-based support.

What We’re Learning, and What’s Next

With over 100 retests and counting, the data speaks for itself. Personalised microbiome support doesn’t just shift numbers—it transforms health outcomes. As BIOME9’s database continues to grow, it’s providing even more confidence in this precision-based approach and expanding understanding of how to support different breeds, life stages, and health challenges.

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