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Laura Jackson

Articles, Case Studies

Case Study: Feline Herpes Virus-Induced Dermatitis: Clinical and Histological Insights

A biopsy from an external nose lesion on a 13-year-old domestic longhair male neutered cat was submitted to our laboratory. Histopathology identified a severe ulcerative necrotising dermatitis with numerous intranuclear inclusion bodies. These histological changes, in combination with the anatomical location are consistent with a herpesvirus infection.

Feline herpes virus ulcerative dermatitis is a rare feline skin disease caused by feline herpes virus 1. Feline herpesvirus 1 is known to cause mainly upper respiratory tract infection (Feline Viral Rhinotracheitis) and it is associated with many clinical symptoms which include: sneezing, nasal discharge, conjunctivitis, mouth ulcers and sometimes fever. In more severe cases breathing may be affected and secondary bacterial infection can develop.

This virus is highly contagious and spreads through direct contact between infected cats or via contaminated surfaces. Once the cat is infected, the virus remains in the body in a latent state, and presence of the virus has been demonstrated in the trigeminal ganglion. Following immunosuppression, the virus can start replicating again and lesions may occur.

In rare instances this virus can cause ulcerative dermatitis, particularly of the face, however, lesions have also been reported on the pinna and flank.

Clinically these lesions are characterised by vesicles, ulcers and crust formation which are variably pruritic. This is a non-specific clinical presentation, and differentials include: hypersensitivity to insect bites, eosinophilic granuloma complex lesions, pemphigus foliaceous, adverse drug reactions and even neoplasia.

Histopathology of this lesion can help distinguish between these entities. On histological examination these lesions are characterised by severe epidermal necrosis and ulceration which often extend into the wall of the underlying hair follicles. The inflammatory infiltrate is usually characterised by significant numbers of eosinophils, however, neutrophils may predominate on occasion. Intranuclear inclusion bodies, when identified, are diagnostic. These are usually observed in the areas of epithelial necrosis.

 

 

Figure 1: On low power the segmental epidermal necrosis and ulceration are evident (blue arrow).

 

 

Figure 2: The necrosis extends into the wall of hair follicles which are multifocally heavily infiltrated by mixed inflammation (blue circle area).

 

 

Figure 3: Large intranuclear inclusion bodies with associated chromatin margination are observed (blue arrow).

 

In the case presented here, inclusion bodies were readily identified, and the diagnosis was straightforward, however, herpes virus inclusion bodies may be difficult to detect or completely absent. In these cases, PCR testing may be useful. However, it is important to bear in mind that a previous vaccination with live vaccine may produce a false positive PCR result. This is due to virus latency and the fact that the PCR currently available in unable to distinguish between field and vaccination strains. Immunohistochemistry for feline herpes virus is also available commercially and it is considered the gold standard to confirm the diagnosis.

In summary, feline herpes virus 1 is a lifelong infection which is usually associated with mild disease, and it is largely prevented by vaccination. In rare cases infection becomes more severe and supportive care is required. Skin lesions associated with this infection are rare, however, they do occur, and, in these cases, histopathology is a very useful tool to identify it.

 

References

Persico, P., Roccabianca, P., Corona, A., Vercelli, A. and Cornegliani, L. (2011), Detection of feline herpes virus 1 via polymerase chain reaction and immunohistochemistry in cats with ulcerative facial dermatitis, eosinophilic granuloma complex reaction patterns and mosquito bite hypersensitivity. Veterinary Dermatology, 22: 521-527. https://doi.org/10.1111/j.1365-3164.2011.00984.x

Sánchez MD, Goldschmidt MH, Mauldin EA. Herpesvirus dermatitis in two cats without facial lesions. Vet Dermatol. 2012 Apr;23(2):171-3, e35. doi: 10.1111/j.1365-3164.2011.01031.

Porcellato I, Luciani L, Marenzoni ML, Santagostino SF, Sforna M, Mechelli L, Brachelente C. Feline herpesvirus ulcerative dermatitis: an atypical case? Vet Dermatol. 2018 Jun;29(3):258-e96. doi: 10.1111/vde.12537

Parzefall, Matiasek, Evidence of feline herpesvirus-1 DNA in the vestibular ganglion of domestic cats, The Veterinary Journal, Volume 184, Issue 3, 2010, Pages 371-372, ISSN 1090-0233, https://doi.org/10.1016/j.tvjl.2009.03.030.

 

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Feline Ventral Abdominal Wall Angiosarcoma
Articles, Case Studies

Case Study: Feline Ventral Abdominal Wall Angiosarcoma

Clinical history

Simba, 9 year old male intact domestic shorthair cat, presented with firm rapidly growing large subcutaneous ulcerated mass on the inguinal area.  The ventral abdominal haired skin containing a portion of the inguinal mass and the inguinal lymph node are submitted for histopathology.

Histology

The dermis and the subcutis are markedly expanded by a moderately cellular neoplasm. Neoplastic cells are either forming anastomosing poorly-organised vascular spaces and channels, supported by a collagenous stroma, or are arranged in streams and sheets, closely packed, supported by a delicate fibrovascular stroma (Fig. 1 and 2). Neoplastic cells are elongated to spindle-shaped, with a prominent ovoid nucleus and small amphophilic nucleoli, and show moderate anisocytosis and anisokaryosis. Ten mitotic figures are observed in 2.37 mm2 – equivalent to 10 high power (x400) fields.

This neoplasm has metastasized to the inguinal lymph node (Fig. 3).

 

Figure 1: Haired skin. The dermis and the subcutis are effaced by a malignant vascular neoplasm, consistent with angiosarcoma (Haematoxylin and eosin stain, 5x).

 

 

Figure 2: Haired skin. Higher magnification of the angiosarcoma. Note the anastomosing poorly-organised vascular spaces (clear spaces, highlighted with asterisks), lined by a single layer of endothelium and  supported by a collagenous stroma (pink areas) (HE stain, 10x).

 

 

Figure 3: Inguinal lymph node. The nodal architecture is effaced by a metastatic angiosarcoma (HE stain, 10x).

 

Interpretation
Angiosarcoma, with nodal metastasis: ventral abdominal haired skin and inguinal lymph node.

Comments
Histology of the ventral abdominal haired skin submitted captures a malignant vascular neoplasm, consistent with angiosarcoma. In this particular case, this angiosarcoma has metastasised to the inguinal lymph node.

Angiosarcoma is a general term for a highly malignant neoplasm arising from endothelial cells of blood vessels (haemangiosarcoma) or lymphatic vessels (lymphangiosarcoma). If the neoplastic vascular channels are filled with blood, this would provide further evidence supporting a diagnosis of haemangiosarcoma. However, immunohistochemistry using endothelial cell markers (factor VIII and CD31) and  a lymphatic vessel marker (PROX-1) may be necessary to further characterise the neoplastic cells and determine whether the angiosarcoma represents a haemangiosarcoma or a lymphangiosarcoma. It should be noted, however, that it is currently unknown whether the different phenotypes affect treatment response or prognosis.

Angiosarcoma is rare in cats; however, the most commonly reported primary sites are cutaneous and visceral locations. Angiosarcomas in cats show a predilection for the subcutis of the caudal ventral abdominal wall, as observed in this case, and this condition is known as ‘feline ventral abdominal wall angiosarcoma’ (FVAA).

FVAA primarily affects middle-aged to older cats, with no distinct breed or gender predisposition.  Clinically, it manifests as a rapidly enlarging, firm to soft mass located in the ventral abdominal region. The mass may appear subcutaneous or involve deeper tissues, such as the abdominal musculature, and is prone to ulceration or spontaneous bleeding due to its vascular nature. Some patients  may present with lesions resembling bruising or inguinal hernia, which may not initially raise suspicion of a neoplastic process.

In addition to the mass, affected cats may present with non-specific clinical signs such as lethargy, anorexia, weight loss, and general malaise. If the neoplasm has metastasised or invaded adjacent structures, clinical signs can include respiratory distress (if pulmonary metastases are present), pale mucous membranes (due to internal haemorrhage), or abdominal distension.

Diagnosis of FVAA typically involves a thorough clinical examination, coupled with imaging studies such as ultrasound or radiography to evaluate the extent of the mass and to check for signs of metastasis. Fine-needle aspiration or core biopsy of the mass may provide initial cytological findings, often showing spindle-shaped or pleomorphic cells with abundant mitotic activity. However, cytology alone may not be definitive due to the poorly exfoliative nature of the cells, and histopathology is ultimately needed to diagnose this condition.

The prognosis for cats diagnosed with ventral abdominal wall angiosarcoma is generally guarded to poor, due to the highly aggressive and invasive nature of the tumour. Angiosarcomas have a marked propensity for local recurrence, even after seemingly complete surgical excision, and a high likelihood of metastasis. The lungs and liver are the most common sites of metastasis, but the tumour can also spread to other organs, including the spleen and kidneys.

Surgical resection is the primary treatment option, and wide surgical margins are essential to reduce the risk of recurrence. However, achieving complete excision can be difficult due to the tumour’s infiltrative growth pattern. Even with aggressive surgical intervention, recurrence rates are high, and the median survival time post-surgery is often limited to a few months. In cases where metastasis has already occurred at the time of diagnosis, survival time may be significantly shorter.

Overall, while early detection and aggressive surgical management may provide temporary control of the disease, the long-term outlook for cats with ventral abdominal wall angiosarcoma remains poor, and most cases have a guarded prognosis. Owners should be informed of the tumour’s aggressive behaviour and potential for rapid progression, and monitoring for recurrence is essential post-surgery.

 

Reference

Bellamy E, Larsen Moberg H, Suárez-Bonnet A, Palma SD, Murgia D, Pittaway R, Verganti S. Feline ventral abdominal wall angiosarcoma: haemangiosarcoma or lymphangiosarcoma? Clinical and pathological characteristics in nine cases. J Feline Med Surg. 2024 Jan;26(1):1098612X231216636. doi: 10.1177/1098612X231216636. PMID: 38227337; PMCID: PMC10949878.

 

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Articles

Histology Tips and Tricks: Sampling of the Hepatobiliary Tree

General Information

Biopsy of the hepatobiliary tree is often required for investigation of hepatobiliary disease. Techniques for biopsy of the hepatobiliary tree include needle biopsy (Tru-cut biopsy), laparoscopic biopsy (cup forceps), surgical biopsy (i.e. wedge or guillotine biopsy), and liver lobectomy. Gall bladder sampling most often involves cholecystectomy, but incisional biopsies may also be taken.

  • Selection of the appropriate biopsy technique is at the veterinarian’s discretion. This decision should be made in conjunction with the patient’s clinical state, coagulation capability and results of other diagnostic testing, and clinical differentials.
  • In general, pathologists prefer biopsies that are obtained laparoscopically or surgically. These samples tend to have fewer artefacts, allow for enhanced assessment of the liver architecture, and provide sufficient tissue for additional testing (such as immunohistochemistry, PARR, and special stains including copper staining). However, needle biopsies are a less invasive sampling technique and may be more appropriate for certain patients.

Submission form and relevant patient history

The pathologist will interpret the biopsy histology in the context of the available clinical information.

  • Please include the full animal details and a detailed clinical history. Include a description of the lesions and their distribution, the timeframe and progression of disease, any previous or ongoing treatments (including antibiotics), and travel history.
  • Include the results of any bloodwork (including serum biochemistry, bile acid stimulation testing, and coagulation testing), including the reference ranges
  • If cytology or other testing has been performed, please include the previous submission number (if performed at VPG) or include a copy of the results (if performed elsewhere)
  • If photos or imaging results are available, please include these because they can aid the pathologist in accurate interpretation

Photos can be uploaded to PATHPORT, emailed to [email protected], or printed and attached to the submission form.

Sampling guidelines

Current guidelines for investigation of hepatobiliary disease recommends a minimum of 12-15 portal tracts be available for histologic examination. Artefacts introduced during sampling, such as crush, shearing, or puncture of the liver by sponges in cassettes, can reduce the number of intact lobules and inhibit histologic interpretation.

 

To maximise the pathologist’s ability to achieve a diagnosis and to have adequate tissue for additional tests, multiple needle biopsies (>4, ideally from a 14 gauge needle) or laparoscopic, cup forceps biopsies (2-4) are required.

  • Multiple lobes should be sampled and labelled according to the lobe from which they were obtained (if it can be identified). If specific lesions are sampled (for example masses), these should be placed in a separate pot and labelled.
  • Biopsies for histology should be placed in a labelled pot containing 10% neutral buffered formalin at a ratio of 10:1 (formalin : tissue).
    • Put needle biopsies in a mesh cassette, taking care not to crush the samples. Avoid using sponges, which can puncture the tissue and introduce artefact.

Large biopsies, including partial or complete lobectomies, may not fix fully. To enhance fixation, incise the lesional areas such as masses.

  • If margin assessment is required, avoid incising the margin. If you have histologic marking ink at your clinic, consider applying the ink to the margin prior to placing the sample in formalin and/or incising it.
  • Note: Staples must be cut out of the tissue before it can be processed. Consequently, if surgical margins are stapled, this may interfere with histologic margin assessment.

 

Liver lobe with 2 masses (yellow stars). To improve fixation, incise the lobe at approximately 2cm intervals. Avoid the surgical margins.

Sampling of patients with suspected canine chronic hepatitis

Laparoscopic biopsies are preferred for sampling patients with suspected chronic hepatitis. However, needle biopsies may also provide sufficient tissue for diagnosis, if enough biopsies are obtained. The current ACVIM consensus guidelines for investigation of canine chronic hepatitis recommend obtaining  biopsies for histology, culture, and copper quantification.

  • Histology: 3 laparoscopic biopsies or 4+ full length, 14 gauge (2cm long) needle biopsies, placed in labelled pot(s) of 10% neutral buffered formalin
    • Copper staining of formalin-fixed liver biopsies is recommended for patients with suspected chronic hepatitis, to allow for assessment of copper accumulation in relation to the pathologic changes. Copper staining may be performed as part of the initial histologic assessment or can be requested after the histology has been performed.
  • Copper quantification: 1 laparoscopic biopsy or 1 full length, 14 gauge (2cm long) needle biopsy in a sterile, liquid free, labelled pot
    • Note that copper quantification requires approximately 20-40mg of liver (wet weight). Insufficient amounts of tissue may yield inaccurate results. This equates to approximately 1 full length 14 gauge (2cm long) needle biopsy or ½ of a 5mm laparoscopic biopsy specimen. A full length 18 gauge needle biopsy provides only 3-5mg of tissue.
  • Culture: 1 laparoscopic biopsy in a sterile, liquid-free, labelled pot
    • In some cases (for example, sampling of liver abscesses), a swab may be indicated. These should be placed in the appropriate transport medium, labelled, refrigerated until transport, and transported to the laboratory as quickly as possible.

Sampling of patients with suspected biliary disease

In patients with suspected biliary disease (including cats with cholangiohepatitis), evaluation of bile, bile “sludge”, cholecystoliths/choledocholiths, and/or the gall bladder may be indicated in addition to liver histology

  • Culture can be performed on fluid, bile sludge and/or stones, and gall bladder and liver (swabs or tissue).
    • These should be placed in the appropriate transport medium or a sterile container. Culture samples should be refrigerated until they are sent via courier to the appropriate laboratory, as quickly as possible.
  • Cytology specimens of the liver and/or bile should be placed in a separate parcel from formalin samples because formalin fumes can alter cellular morphology, limiting cytologic interpretation.
  • The gall bladder has a thick wall, which can slow the rate of formalin penetration and inhibit fixation. To expedite fixation, it can be incised. If margin assessment is required, avoid cutting into the relevant margin.

Sampling of patients with suspected lymphoma

Formalin fixation can inhibit PCR including PARR (PCR for antigen receptor rearrangement), which may be recommended by the pathologist to investigate for a clonal lymphocyte population and the diagnosis of lymphoma.

  • If lymphoma is suspected clinically (particularly in cats with suspected lymphocytic cholangitis/cholangiohepatitis or small cell lymphoma), consider placing one biopsy sample in saline for PARR, if needed.

Interpreting the Pathology Report

We will always strive to give you a definitive diagnosis. However, in some cases, for various reasons this may not be possible. In those cases we will provide an opinion on the most likely diagnosis and possible differential diagnosis and discuss further diagnostic tests, if applicable. In particular in cases with some uncertainty regarding the diagnosis, it is important to compare to the clinical picture, list of clinical differential diagnoses, and results of other tests (e.g. imaging) that you may have done. We are always happy to discuss any questions you may have regarding the histopathology report.

Quick Facts

  • Aim for 4+ needle (14g) or 2-4 laparoscopic cup biopsies for liver histology.
  • Consider taking samples for liver cytology, culture, copper quantification, and PARR.
  • Bile, choleliths, bile sludge, and gall bladder sampling may also be indicated.
  • Clearly label all samples with the patient name and site.
  • Make incisions into larger samples (avoiding marginal areas) to aid fixation.
  • Histology samples should be put in labelled, leak-proof containers containing 10% neutral buffered formalin.
  • Cytology slides should be clearly labelled with the patient name and site and placed in break-free slide holders in a separately labelled bag.
  • Include the submission form in a labelled bag. The pathologist will incorporate any provided clinical information into their interpretation, so we thank you for your thoroughness.

For more information on our histopathology services please visit our Histopathology page or contact us by calling 0117 951 1283 or emailing [email protected].

Reference:

Webster CRL, Center SA, Cullen JM, et al. ACVIM consensus statement on the diagnosis and treatment of chronic hepatitis in dogs. J Vet Intern Med. 2019; 33: 1173–1200. https://doi.org/10.1111/jvim.15467

 

 

 

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Articles, Case Studies

Case Study: 3-Year-Old Crossbreed Dog with a Cutaneous Mass

Signalment

A 3-year-old male neutered crossbreed dog presented with a small, cutaneous mass located in the left axilla. The owner reported no noticeable changes in the dog’s behaviour, appetite, or activity level. Upon physical examination, the mass appeared raised, firm, mildly erythematous, and well-circumscribed, with no signs of ulceration. The dog did not seem bothered by the mass, and palpation elicited no apparent discomfort. No additional masses were observed, and the peripheral lymph nodes were unremarkable.

Cytologic Findings

A fine-needle aspirate (FNA) of the mass was collected and submitted for cytologic evaluation. Microscopic examination revealed a monomorphic population of round cells with uniform, round to reniform nuclei. The chromatin was finely stippled, and nucleoli were not clearly discernible. The cytoplasm of the cells was moderate in amount and pale blue in colour. These cytologic features suggested a round cell tumour, with characteristics most consistent with a histiocytoma.

 

 

The findings were suggestive of a round cell population, a broad category that includes tumours such as histiocytomas, mast cell tumours, lymphomas, and transmissible venereal tumours. However, based on the cytologic details and the patient’s age, a histiocytoma was considered the most likely diagnosis. Note the presence of a single mast cell and occasional neutrophils.

Discussion: Canine Histiocytomas

Histiocytomas are benign skin tumours that arise from Langerhans cells, which are specialised immune cells located within the skin’s epidermis. These tumours are common in younger dogs, generally appearing in animals under three years of age. Typically, histiocytomas are dome-shaped, small (usually less than 2 cm), and can occur on various parts of the body, including the head, ears, neck, and limbs, though they are not restricted to these areas and may also appear in other locations.

The typical course of a histiocytoma is benign and self-limiting. Most histiocytomas undergo spontaneous regression within one to three months, driven by the immune system’s recognition and clearance of the tumour. During this regression phase, the mass may become more erythematous or ulcerated before ultimately resolving on its own. When an FNA is obtained at this point, evidence of lymphocytic inflammation is often found. The cytologic appearance of histiocytomas can help distinguish them from other round cell tumours, such as lymphomas (which typically have larger nuclei and less cytoplasm), mast cell tumours (characterized by metachromatic granules), and plasma cell tumours (which often have greater pleomorphism and darker cytoplasm).

Prognosis for histiocytomas is excellent. Surgical removal is usually not required unless the tumour fails to regress spontaneously or is located in a problematic area that interferes with the dog’s movement or causes discomfort. However, some owners may opt for surgical removal for cosmetic reasons or if regression is prolonged. In cases where surgical removal is pursued, histopathology can be used to confirm the diagnosis.

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Articles

Histopathology: Sampling and shipment tips and tricks

Corresponding to other laboratory tests, the quality of histopathology reporting is directly related to the quality of the sample received. This document will explain common pitfalls in sampling and shipment of histopathology samples to help you avoid introducing pre-analytical artefacts.

Sampling

  • In particular when taking small biopsies, even with careful sampling, crush artefact cannot always be prevented. In addition, small biopsies may not always be fully representative of the underlying lesion. To mitigate this, we recommend taking multiple samples to increase the chance of obtaining representative tissue sections. For example, one study on gastrointestinal biopsy sampling reported a minimum of 6 mucosal biopsy specimen of adequate quality from the duodenum of cats are required for a reliable histopathological evaluation. For optimal reporting, 10-15 gastrointestinal biopsies per site (excluding ileum) are recommended1.
  • If you are using electrocautery, be aware that this causes coagulation artefact at the borders of the samples and may therefore impact histologic margin assessment (if required).
  • Transportation can cause fragmentation, especially with friable samples. To protect such samples during transit, it is advisable to use histology cassettes, which can be ordered via VPG via your local laboratory.
  • Avoid using foam inlays for fresh tissue, as they can cause crush artifact.

Histology cassettes

 

Inking

  • If histologic margin assessment is required, inking by the surgeon is very helpful for tissue orientation and to ensure that the tissue border on a histologic slide represents the surgeon-cut edge created at surgery (any samples that require histologic margin assessment that do not arrive in the lab pre-inked will be inked in the lab)
  • For best inking results, blot the tissue dry, apply surgical ink to the desired area, with a brush or cotton swab, wait for five minutes for the ink to dry, and then immerse the tissue in formalin.


‘Formalin pigment’ (acid hematin)

 

Fixation

  • To achieve optimal fixation, use 10% neutral-buffered formalin (NBF). The buffering agent reduces the formation of “formalin pigment,” which can interfere with accurate analysis. The histology containers that you can order via your local laboratory are already prefilled with 10% NBF.
  • Never freeze the sample before fixation or send it in alcohol or saline. These methods are not suitable and will cause significant tissue artefact.
  • Ideally, maintain a 10:1 formalin-to-sample volume ratio during fixation. However, if this is not feasible, ensure that the sample is at least covered with formalin and send it to the lab as soon as possible.
  • We recommend a fixation time of 24 to 48 hours. Prolonged fixation in particular of large samples and samples rich in adipose tissue (mammary strips) or blood (spleens) without cutting the sample further will not improve fixation in the centre.
  • For larger samples, carefully consider cutting them to enhance formalin penetration. However, exercise caution to avoid destroying the lesion or the margins that need assessment. For example, a large elliptical skin excision with a mass can be incised from the skin side into the mass without cutting through the mass or the deep and horizontal tissue borders


Incision into mass without cutting through tissue borders to enhance formalin penetration

 

  • If a container of adequate size is not available, samples can be halved. For large and elongate samples, such as mammary strips, cutting the sample into multiple sections is acceptable. Cutting a single mass into more than 2 sections should be a last resort, as it can make accurate margin assessment challenging. If you have surgical ink, apply the ink to the sample prior to sectioning to preserve the ability to perform accurate margin assessment. Any post-surgical alteration of a sample should always be noted down in detail on the submission form and containers must be labelled accordingly.

Sectioning a mammary strip

 

Shipment

  • Make sure no needles or other sharp materials are left in the sample/ histology container, as this is a significant health and safety risk for our staff!
  • Always use containers of appropriate size to prevent impairing fixation and deforming the sample. Squeezing samples into containers that are too small can adversely affect margin assessment.
  • Please do not use unsuitable containers (sharps containers, Tupper ware) or containers that put our staff at risk (e.g. glass containers).
  • Ensure that the container is leak-proof, shatter-proof, and include absorbent material in the shipment bag if possible.
  • Use individual containers for each sample or site to allow for correct sample/ site identification throughout the process; label the containers and make notes on the submission form accordingly.
  • Keep cytology samples separate from histology samples, as formalin fumes can cause severe artefact in cytology sample.

 

Example of well-packaged histology sample

Shipment of large samples

For large samples that do not fit in any suitable containers and cannot be cut into smaller sections (such as whole legs) consider:

  • Is margin assessment required for this sample? For example in a leg amputation where the lesion is located at or below the carpus/ tarsus, margin assessment is likely not relevant, as ‘clear’ margins are expected. In those cases, the portion of the sample with the lesion can be cut out and fixed and submitted as usual.
  • Note that margin tissue could also be submitted separately and clearly labelled as such.
  • Can parts of the tissue that are not relevant for the histologic assessment be discarded? For example in leg amputations with the lesion high on the leg, the lower extremity can be cut off and the remaining sample fixed and submitted as usual.
    • Keep any tissue that has been cut off in the practice in formalin until the final report has been issued.

If the whole sample is required for histologic assessment:

  • Fix the whole sample in a container of sufficient size with an adequate amount of formalin for 24-48h. If the day of shipment would fall on a Friday, then keep the sample in formalin over the weekend and ship the sample on Monday. Consider if incisions can be made that facilitate better formalin penetration without impairing any necessary margin assessment (see above).
  • Once fixed, wrap the sample in formalin-soaked gauze/ tissue papers, then double-bag in plastic bags of adequate size; consider adding absorbent material (tissue papers/ cotton wool) inside the second bag to prevent leakage.
  • Sending samples fresh without formalin-fixing them first should be an absolute last resort! Please contact VPG Histology prior to sending a fresh sample.

Sampling and shipment of spleens

  • Spleens have a thick capsule and are often markedly congested, both of which hampers proper formalin fixation. To enhance fixation, either several incisions through the capsule above a mass and in ‘normal’ areas of the spleen, at approximately 2cm intervals.

Entire spleen submitted to VPG Histology. Incisions can be made at approximately 2cm intervals (green lines) through normal spleen and masses to improve fixation.

 

  • If the whole spleen cannot be sent/ you wish to send incisional biopsies, please consider the following
    • It is critical that representative sections are taken from the transition zone between the mass and normal spleen. Splenic masses commonly rupture: sampling of the haematoma or necrosis often does not reveal the underlying cause of rupture.
    • Neoplasia (including haemangiosarcoma) may not be diagnosed if only the subcapsular haematoma is sampled or if fewer than 5 sections are taken.
      • In fact, in one study2, the likelihood of missing a haemangiosarcoma is 32% and 17% if only 1 or 2 sections, respectively, are examined.
      • If 5 sections are examined the likelihood of missing haemangiosarcoma decreases to 5%.
      • If 10 sections are examined, the likelihood of missing haemangiosarcoma decreases to 1%.
    • Consequently, we recommend taking 5 representative sections from the transition area between the mass and normal spleen and 1 additional representative section from the normal spleen.
      • Please take care to sample the transition area between a mass and ‘normal’ spleen, otherwise it is possible that only reactive lesions (e.g. the central haematoma in a haemangiosarcoma) are represented in the samples.

Obtaining representative sections from a spleen. Take a minimum of 5 sections between the transition zone of the mass and normal spleen (green boxes). Also take at least one section of normal spleen (orange box).

 

Sampling of the hepatobiliary tree

Biopsy of the hepatobiliary tree is often required for investigation of hepatobiliary disease. Techniques for biopsy of the hepatobiliary tree include needle biopsy (Tru-cut biopsy), laparoscopic biopsy (cup forceps), surgical biopsy (e.g. wedge or guillotine biopsy), and liver lobectomy. Gall bladder sampling most often involves cholecystectomy, but incisional biopsies may also be taken.

Current guidelines for investigation of hepatobiliary disease recommends a minimum of 12-15 portal tracts be available for histologic examination. Artefacts introduced during sampling, such as crush, shearing, or puncture of the liver by sponges in cassettes, can reduce the number of intact lobules and inhibit histologic interpretation.

To maximise the pathologist’s ability to achieve a diagnosis and to have adequate tissue for additional tests, multiple needle biopsies (>4, ideally from a 14 gauge needle) or laparoscopic, cup forceps biopsies (2-4) are required.

  • Multiple lobes should be sampled and labelled according to the lobe from which they were obtained (if it can be identified). If specific lesions are sampled (for example masses), these should be placed in a separate pot and labelled.
  • Biopsies for histology should be placed in a labelled pot containing 10% neutral buffered formalin at a ratio of 10:1 (formalin : tissue).
    • Put needle biopsies in a mesh cassette, taking care not to crush the samples. Avoid using sponges, which can puncture the tissue and introduce artefact.

Example of a mesh cassette

Large biopsies, including partial or complete lobectomies, may not fix fully. To enhance fixation, incise the lesional areas such as masses.

  • If margin assessment is required, avoid incising the margin. If you have histologic marking ink at your clinic, consider applying the ink to the margin prior to placing the sample in formalin and/or incising it.
    • Note: Staples must be cut out of the tissue before it can be processed. Consequently, if surgical margins are stapled, this may interfere with histologic margin assessment.

Liver lobe with 2 masses (yellow stars). To improve fixation, incise the lobe at approximately 2cm intervals. Avoid the surgical margins.

Sampling of patients with suspected canine chronic hepatitis

Laparoscopic biopsies are preferred for sampling patients with suspected chronic hepatitis. However, needle biopsies may also provide sufficient tissue for diagnosis, if enough biopsies are obtained. The current ACVIM consensus guidelines for investigation of canine chronic hepatitis recommend obtaining  biopsies for histology, culture, and copper quantification3.

  • Histology: 3 laparoscopic biopsies or 4+ full length, 14 gauge (2cm long) needle biopsies, placed in labelled pot(s) of 10% neutral buffered formalin
    • Copper staining of formalin-fixed liver biopsies is recommended for patients with suspected chronic hepatitis, to allow for assessment of copper accumulation in relation to the pathologic changes. Copper staining may be performed as part of the initial histologic assessment or can be requested after the histology has been performed.
  • Copper quantification: 1 laparoscopic biopsy or 1 full length, 14 gauge (2cm long) needle biopsy in a sterile, liquid free, labelled pot
    • Note that copper quantification requires approximately 20-40mg of liver (wet weight). Insufficient amounts of tissue may yield inaccurate results. This equates to approximately 1 full length 14 gauge (2cm long) needle biopsy or ½ of a 5mm laparoscopic biopsy specimen. A full length 18 gauge needle biopsy provides only 3-5mg of tissue.
  • Culture: 1 laparoscopic biopsy in a sterile, liquid-free, labelled pot
    • In some cases (for example, sampling of liver abscesses), a swab may be indicated. These should be placed in the appropriate transport medium, labelled, refrigerated until transport, and transported to the laboratory as quickly as possible.

Sampling of patients with suspected biliary disease

In patients with suspected biliary disease (including cats with cholangiohepatitis), evaluation of bile, bile “sludge”, cholecystoliths/choledocholiths, and/or the gall bladder may be indicated in addition to liver histology

  • Culture can be performed on fluid, bile sludge and/or stones, and gall bladder and liver (swabs or tissue).
    • These should be placed in the appropriate transport medium or a sterile container. Culture samples should be refrigerated until they are sent via courier to the appropriate laboratory, as quickly as possible.
  • Cytology specimens of the liver and/or bile should be placed in a separate parcel from formalin samples because formalin fumes can alter cellular morphology, limiting cytologic interpretation.
  • The gall bladder has a thick wall, which can slow the rate of formalin penetration and inhibit fixation. To expedite fixation, it can be incised. If margin assessment is required, avoid cutting into the relevant margin.

 

Sampling of patients with suspected lymphoma

Formalin fixation can inhibit PCR including PARR (PCR for antigen receptor rearrangement), which may be recommended by the pathologist to investigate for a clonal lymphocyte population and the diagnosis of lymphoma.

  • If lymphoma is suspected clinically (particularly in cats with suspected lymphocytic cholangitis/cholangiohepatitis or small cell lymphoma), consider placing one biopsy sample in saline for PARR, if needed.

 

Submission form and relevant patient history

  • You can book in any collected samples onto Pathport at: https://pathport.thevpg.co.uk
  • Along with the submission form containing the animal details, for optimal reporting, it is essential to provide the relevant patient history. Include a description of the lesions, their distribution, timeframe of occurrence/ progression, international travel history, and any previous or ongoing treatment, particularly antibiotic and steroid usage.
  • Include any relevant photos, imaging results, blood results, and cytology results from other labs, as they can aid in accurate interpretation. Photos can be uploaded directly into Pathport, emailed to [email protected], or printed and affixed to the submission form.
  • Specify your clinical differential diagnoses and the specific questions you want the histopathology report to address.

 

Interpreting the Pathology Report

We will always strive to give you a definitive diagnosis. However, in some cases, for various reasons this may not be possible. In those cases we will provide an opinion on the most likely diagnosis and possible differential diagnosis and discuss further diagnostic tests, if applicable. In particular in cases with some uncertainty regarding the diagnosis, it is important to compare to the clinical picture, list of clinical differential diagnoses and results of other tests (e.g. imaging) that you may have done. We are always happy to discuss any you may have regarding the histopathology report.

For more information on our histopathology services, including comprehensive guides on sampling spleens and the hepatobiliary tree, please visit our website or contact us by calling 0117 951 1283 or emailing [email protected].

 

References:

  1. Willard MD, Mansell J, Fosgate GT, et al. Effect of sample quality on the sensitivity of endoscopic biopsy for detecting gastric and duodenal lesions in dogs and cats. J Vet Intern Med. 2008;22:1084-1089.)
  2. Herman EJ (2019). Understanding the efficiency of splenic hemangiosarcoma diagnosis using Monte Carlo simulations. Vet Pathol 56(6): 856-859 doi:10.1177/0300985819868732
  3. Webster CRL, Center SA, Cullen JM, et al. ACVIM consensus statement on the diagnosis and treatment of chronic hepatitis in dogs. J Vet Intern Med. 2019; 33: 1173–1200. https://doi.org/10.1111/jvim.15467

 

 

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Articles, Case Studies

Cutaneous Sarcomas In Pet Degus

Introduction

Neoplasia is considered uncommon in degus, with previous research indicating a low incidence in this species. A study by Jekl et al. (2011) found that only six out of 300 pet degus examined showed evidence of neoplastic disease. Among the reported cases, fibrosarcomas have been identified as the most prevalent cutaneous neoplasm, as highlighted in a study by Svara et al. (2020). This study aims to retrospectively analyse cutaneous neoplasms in pet degus within the United Kingdom, providing further insight into their occurrence and characteristics.

Results

A systematic review of archived case material identified 35 skin neoplasms in pet degus, with 31 of these diagnosed as sarcomas, making them the most prevalent tumour type in this species. The average age at diagnosis was five years, with cases ranging from two to nine years old. Among the affected degus, 16 were male, 11 were female, and for four cases, age details were not recorded on the submission form. Sarcomas most commonly occurred on the rump or legs, with fewer cases involving the head or neck. Histopathological examination revealed a range of sarcoma types, from well-differentiated spindle cell sarcomas to pleomorphic sarcomas exhibiting multinucleated cells.

Figure 1: Age distribution of the patients.

Figure 2: Sites of cutaneous sarcomas.

 

Immunohistochemistry confirmed vimentin expression, supporting a mesenchymal origin. Further analysis of selected cases for the histiocytic marker Iba-1 and the muscle marker desmin showed that all tested samples were negative for Iba-1, while some exhibited desmin expression in a subset of neoplastic cells. Regarding surgical excision, 22 tumours were submitted as excisional samples, but only six cases achieved complete excision. Additionally, four sarcomas were submitted following limb amputation, with three showing no evidence of residual tumour tissue.

 

Discussion

This study confirms that the majority of cutaneous neoplasms in pet degus are sarcomas, reinforcing previous findings on neoplastic trends in this species. Interestingly, the results indicate that these tumours were more commonly found in male degus, which contrasts with earlier research by Svara et al. (2020), where cutaneous sarcomas were reported as more prevalent in females. This discrepancy highlights the need for further investigation into potential sex-related predispositions or underlying factors influencing tumour development.

 


Figure 3: Histopathological features of cutaneous sarcomas in degus.
Left: well-differentiated sarcoma forming streams and exhibiting low mitotic activity.
Right: Poorly-differentiated sarcoma exhibiting multinucleated cells (asterisk) and frequent mitotic figures (arrows). H&Estain. Scale bar = 100μm

 

Histological and immunohistochemical analysis confirmed that these neoplasms were consistent with soft tissue sarcomas. The expression of desmin in some cases suggests a possible smooth muscle, skeletal muscle, or myofibroblast origin. Additionally, the absence of Iba-1 expression indicates that none of the tumours represented histiocytic sarcomas. Notably, no cases of cutaneous hemangiosarcomas or extra-osseous osteosarcomas were identified within this study population. Despite attempts at complete surgical excision in 26 out of 31 cases, histological examination confirmed that only nine achieved lesion-free tissue borders, emphasising the challenges of achieving clear surgical margins in these tumours.

 


Figure 4: Immunohistochemistry analysis.
Left: Neoplastic cells exhibit expression of vimentin.
Right: Occasional neoplastic cells exhibit expression of desmin. Scale bar = 100μm

 

Materials and methods

Study population

A systematic review of archive material of skin masses from degus submitted for histopathological evaluation to the authors’ laboratories between 2012 and 2024 was carried out. The age, sex , site and surgical procedure (excisional or incisional biopsy) were analysed based on the information given on the submission form.

Histopathology and immunohistochemistry analysis

For a review of all sarcoma cases the formalin-fixed paraffin-embedded tissue samples were sectioned at a thickness of 2-3 μm and stained with haematoxylin and eosin (H&E). For immunohistochemistry analysis antigen retrieval, labelling and counterstaining were performed on an Autostainer Link 48 (Agilent, Stockport, UK) using the En Vision Flex detection system (Agilent).

Primary antibody dilutions were 1:1000 for vimentin (Agilent), 1:800 for Iba-1 (EMD Millipore, Darmstadt, Germany) and 1:200 for desmin (Agilent) Target retrieval for all three antibodies was performed using Target retrieval solution high pH (Agilent). Sections were counterstained using haematoxylin.

Conclusions

Soft tissue sarcomas were the most common skin masses identified in degus in this study. On histological examination excision was often incomplete; however follow-up studies are required to determine the clinical outcome and prognosis of these tumours.

References

JEKL, V., HAUPTMAN, K. & KNOTEK, Z. (2011) Diseases in pet degus: a retrospective study in 300 animals. Journal of small animal practice 52 (2), 107-12 .

ŠVARA T., GOMBAČ M., POLI A., RAČNIK J. & ZADRAVEC M. (2020). Spontaneous Tumors and Non-Neoplastic Proliferative Lesions in Pet Degus (Octodon degus). Veterinary Sciences.13;7(1):32.

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Articles

Understanding Blood Smears

The Power of the Blood Smear: A Simple Tool That Tells Big Stories

If there’s one thing I’ve learned during my residency in Veterinary Clinical Pathology, it’s this: never underestimate the humble blood smear. While automated analysers can crunch numbers and spit out results quickly, they can’t capture the whole story. That’s where a smear comes in – it’s a window into the microscopic world of your patient, full of clues and surprises you might otherwise miss.

Why Bother with a Blood Smear?

Analysers are impressive and mighty useful pieces of machinery, but they’re not foolproof. A blood smear gives you the chance to verify results and identify morphological irregularities, which is especially important when things just don’t add up.

  1. White Blood Cells (WBCs): More Than Just Numbers
  • Double-Checking the Analyser: Sometimes, the analysers get confused. Toxic neutrophils might be counted as monocytes, or a reactive monocytosis might be mistaken for a lymphocytosis. Seeing the cells under a microscope clears up the confusion.
  • Spotting Left Shifts and Toxic Change: Even in the absence of a neutrophilia, the finding of bands or toxic change signals inflammation, which might otherwise be missed.
    • Morphology of Left Shifts: When referring to a ‘left shift’, we mean to say that immature, non-segmented neutrophils have been identified and are circulating. The further left, the less mature the cells are. Myeloid cells will begin as myeloblasts, the earliest form, which then mature through promyelocytes, myelocytes, metamyelocytes, bands, and finally mature neutrophils. This process usually happens in the bone marrow, but when the need for neutrophils increases – for example during active infection – these cells might be released from the bone marrow a little earlier than planned. Depending on the magnitude of inflammation, a left shift could mean the odd circulating band, or in severe cases, circulating myelocytes or even rare promyelocytes. It is important not to confuse these cells with neoplastic cells.


Maturation sequence from myeloblast to mature neutrophils.

 

    • Toxic change: The term toxicity is a bit of a misnomer. When discussing toxicity, we are referring to increased cytoplasmic basophilia, foamy vacuolation and presence of Dohle bodies (aggregates of retained rough endoplasmic reticulum) that neutrophils quite commonly exhibit under strong inflammatory conditions. Toxic granulation is again slightly different, referring to the retention of pink primary granules seen in promyelocytes (see above image) – a rare sign of very severe inflammation. Don’t confuse these with the pale pink staining ‘secondary granules’ seen in normal mature neutrophils (they are called granulocytes for a reason!)


 High numbers of neutrophils in a blood film. Neutrophils have foamy basophilic cytoplasm (toxicity) and reduced nuclear segmentation (bands).

 

  • Catching Atypical Cells: Some of the most fascinating cases come from spotting atypical cells. For instance, with or without a lymphocytosis, an increase in the proportion of small granular lymphocytes could point to developing chronic lymphocytic leukaemia (CLL), which you might not know about without peeking at the film. Likewise, myeloid precursors can only be spotted down the scope, and in the eyes of an analyser, rare lymphoblasts may masquerade as monocytes.

Low numbers of these cells were found to be circulating, even though the total WBC was within reference interval. The nucleus is slightly larger than the neutrophil next to it and contains several pale nucleoli. The patient was known to have lymphoma.

 


Two atypical large lymphocytes are seen next to a small lymphocyte. A neutrophil and a platelet is also present.

 

  1. Platelets: More Than Meets the Analyser
  • Clumps Happen: Platelet clumping can lead to falsely low platelet counts, when in reality there are plenty of platelets – this is especially common in cats. Assessing the blood film for platelet clumps is quick and easy, and a manual minimum estimate can be performed at the same time.

Platelet clumps at the feathered edge of the smear.

 

  • Platelet Count: Without clumping, most cats and dogs will have around 10-30 platelets per 100x field of view. To perform a manual count, the average number of platelets in ten 100x fields of view should be counted and an average calculated. This number is then multiplied by 15,000 to get the number of platelets per microlitres of blood.

There are 7 platelets in this 100x field of view

 

  • Macroplatelets and Breed Quirks: Some breeds, like Cavalier King Charles Spaniels, naturally have larger platelets due to hereditary traits. A smear can confirm this and prevent unnecessary concern.
  • Parasites: A very lucky pathologist might spot a morula from Anaplasma platys lurking in a platelet – a discovery no machine will make for you. This is something to consider in our well-travelled canine patients.


Platelet clumps

 

  1. Red Blood Cells (RBCs): The Subtle Storytellers
  • Signs of Regeneration: Spotting polychromasia and nucleated RBCs on a smear is a sign that the bone marrow recognises the need for more RBCs and is trying to keep up with demand.
  • Hypochromia: The MCHC value is a calculation derived from HCT and Hb, which are prone to error, and is also lower when increased numbers of reticulocytes are present. Confirming hypochromia (and a microcytosis) on a blood smear will help you know when to be worried about genuine iron restricted haematopoiesis.


Increased central pallor suggesting hypochromia. Occasional reticulocytes are also present.

 

  • Shape and Size Matter: A smear can reveal changes such as hereditary macrocytosis in a poodle or microcytosis from iron deficiency. Other changes such as Heinz bodies (denatured precipitated haemoglobin), spherocytes (usually associated with IMHA in dogs) or schistocytes (fragmented due to shear trauma with many possible causes) may also point to pathology requiring exploration.089

An erythrocyte impacts a fibrin strand under flow conditions, forming a schistocyte and a micospherocyte. Image adapted from Harvey’s Veterinary Haematology

 

  • Parasites and Bacteria: A smear may be key to identifying parasites or bacteria hiding out in or on RBC’s. Haemotrophic mycoplasmas and babesiosis are two such examples that can result in severe anaemia in cats and dogs.


Haemoproteus parasite in the erythrocyte of an owl. Parasitised birds rarely show serious disease and are therefore subclinically infected.

 

Making the Most of a Blood Smear

Smears are simple, but not all are created equal. If you want your smear to tell its full story, here are a few tips:

    • Fresh is Best: Time takes its toll on cell morphology. Smears made from aged blood can make cells look toxic or vacuolated, so we prefer to assess morphology from fresh submitted smears. It is also important to ensure smears are not refrigerated or exposed to excessive heat.
    • Capture the Feathered Edge: This is necessary for identifying platelet clumps and large neoplastic cells or parasites.
    • Perfect Your Technique: A clean, thin smear with an evenly spread monolayer is best. DiffQuik if reviewing in house or send unstained to your laboratory.

 

The Challenges of Blood Smears (and Why They’re Worth It)

Let me be honest: blood smears aren’t always easy. Even now, I sometimes struggle to identify subtle toxic changes in neutrophils or decide if a lymphoid population is reactive or neoplastic. It’s humbling. But it’s also where the learning happens.

One thing that helps is correlating findings with the patient’s story. For example, a young dog with a lymphocytosis and small cells on the smear probably doesn’t have CLL -context matters. And when in doubt, talking things over with colleagues is a lifesaver and a great learning opportunity. Follow-up tests like flow cytometry can confirm or refute your suspicions, giving you more confidence next time.

 

 


A moderate lymphocytosis consisting of small to medium lymphocytes with oval to indented nuclei and expanded amounts of pale blue cytoplasm. This was interpreted as a monocytosis by the in-house analyser. Flow cytometry confirmed T-lymphocyte origin, with moderate loss of the pan-lymphocyte marker CD45 supporting a leukaemic phase of T-zone lymphoma.

 

Why We Love Blood Smears

There’s something deeply satisfying about finding an answer hidden in a blood smear. It’s like solving a puzzle no one else even knew was there. And while I know I have a long way to go before I feel like an expert, every case teaches me something new.

So, the next time you’re drawing blood, consider adding a smear to the mix. It’s a small step that can make a huge difference.

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Articles, Insights

Revolutionising Dog Gut Health: BIOME9 Microbiome Testing with VPG

Introduction 

Imagine walking your dog through a forest. The soil nourishes the trees, and the diversity of species creates a rich and balanced ecosystem. This harmony mirrors the gut microbiome of a dog, where each tree represents a bacterial species. A diverse and balanced microbiome ensures resilience, allowing the system to recover from disruptions. However, when balance is lost, and one bacterial species overgrows, the gut environment destabilises, leading to a condition known as dysbiosis. 

Dysbiosis triggers inflammation in the gut lining, which, if untreated, can spread systemically, affecting the gastrointestinal system, skin, joints, cardiovascular health, energy levels, and even neurological functions. The gut microbiome does much more than digest food; it plays a vital role in overall health by producing cytokines, neurotransmitters, and hormones. 

Maintaining a rich and balanced gut microbiome is essential for canine well-being. BIOME9 is making this easier than ever with advanced microbiome testing designed to identify and address imbalances at their source.


GutDiscovery ® Microbiome Profiles and Insights 

BIOME9’s GutDiscovery® platform has analysed microbiome profiles from 1,000 dogs between March and November 2024, collecting approximately 40 billion bacterial sequences. Each faecal sample contained an average of six bacterial phyla, 12 classes, 17 orders, 37 families, 161 genera, and 1,543 species, showcasing the incredible diversity of the canine gut microbiome. 

Key Findings: 

  • Healthy dogs exhibit greater microbial diversity, with the four most abundant bacterial species accounting for 50% of their microbiome. 
  • Dogs with gastrointestinal (GI) unrest or skin irritation experience a significant loss in diversity, with the same four species dominating up to 75% of their microbiome. This imbalance reduces ecological space for beneficial probiotic species critical to overall health. 

 

Figure 1: Relative abundance profiles for 1,000 GutDiscovery ® microbiomes, generated using Oxford Nanopore 16S rRNA v1-9 sequencing and the Kraken2 reference database. Data were analysed and visualised using the R packages phyloseq and ggplot2.

 

Condition-Specific Microbial Differences 

A groundbreaking insight from this dataset is the statistically significant differences in specific bacterial species between dogs with GI conditions and those with skin irritation. While healthy dogs maintain a balance of species, those with conditions often experience either an overabundance or underrepresentation of key microbes. 

For example: 

  • Dogs with GI conditions often display an overgrowth of particular bacterial species, contributing to symptoms such as diarrhoea or bloating. 
  • Dogs with skin irritation show a reduction in beneficial microbes, exacerbating inflammatory responses. 

Understanding these differences helps veterinarians and pet owners target treatments more effectively, tailoring care to the specific needs of the animal.

Figure 4: Differential abundance analysis using a generalised linear model (GLM) based on the negative binomial distribution (P < 0.01) with Benjamini-Hochberg correction (FDR < 0.05). Normalisation was applied to correct for the overestimation of low-abundance species. Data were processed using the R packages DESeq2 and phyloseq.

 

Biome9 Testing: A Game-Changer for Veterinary Clinics 

Through our collaboration, VPG and BIOME9 offer cutting-edge microbiome testing kits designed for ease of use and maximum impact. These kits provide: 

  • Comprehensive Analysis: Advanced sequencing technologies deliver actionable insights into microbial imbalances. 
  • Convenience: Pet owners collect samples at home, eliminating the need for multiple clinic visits. 
  • Tailored Recommendations: Reports include nutritional and supplement advice to help restore balance and improve overall health. 

By integrating BIOME9 testing into your practice, you can offer clients innovative solutions for managing common conditions such as GI unrest and skin irritation, while positioning your clinic as a leader in advanced diagnostics. 

 

Why Choose VPG and Biome9? 

  1. Scientific Excellence: Powered by BIOME9’s state-of-the-art technology and supported by VPG’s trusted diagnostic network. 
  2. Ease of Access: Order microbiome testing kits through VPG, streamlining the process for veterinary clinics. 
  3. Client Engagement: Enhance trust and satisfaction by offering precise, science-backed insights that empower pet owners.

Conclusion 

BIOME9’s GutDiscovery® platform reveals the critical role of the gut microbiome in canine health. With clear distinctions in microbial composition between healthy dogs and those suffering from conditions like gastrointestinal unrest or skin irritation, this groundbreaking testing offers a transformative approach to understanding and addressing these challenges. 

By partnering with BIOME9, VPG is set to revolutionise how veterinarians tackle gut health, providing cutting edge tools to diagnose, treat, and prevent conditions more effectively. With microbiome testing kits now available through VPG, the future of veterinary diagnostics is within reach. 

Comprehensive Faecal Testing Tailored to Your Practice 

At VPG, we deliver comprehensive and often bespoke faecal testing solutions to meet both practice and pet parent needs. Our routine faecal testing is designed to detect common parasites and bacterial infections, all carried out by our team of extraordinary experts. 

Our services go beyond routine faecal flotation and filtration examinations. With antigen and PCR testing, we ensure the most accurate results, empowering you to deliver the best outcomes for your patients. 

No matter your faecal testing requirements, our team is here to help. Give us a call today to discuss your needs and discover how we can support your practice. 

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Articles, General

Complete Blood Count (CBC): Decoding the Numbers and What They Mean for Your Patients

The CBC is more than just numbers on a report — it’s a snapshot of your patient’s cellular health. It represents the dynamic interplay between peripheral cellular demand and the bone marrow response, mediated by various hormonal, neural and cytokine stimuli. And as a veterinary clinical pathology resident, I’ve come to appreciate the depth of information the humble CBC provides. Let’s break it down and explore how to interpret these values with confidence.

 

 

 

What Makes Up a CBC, and Where Do These Cells Come From?

A CBC evaluates the three major cellular components of blood: red blood cells (RBCs), white blood cells (WBCs), and platelets. But understanding these cells starts with their origins in the bone marrow.

  • Red Blood Cells (RBCs): These originate from erythropoietic precursors in the bone marrow, maturing under the influence of erythropoietin (EPO). EPO production ramps up in response to hypoxia, regulated by hypoxia-inducible factors. Immature RBCs, or reticulocytes, are released into circulation, where they complete maturation. While dogs, cats, and pigs readily release reticulocytes into circulation, horses retain them in the marrow, and healthy cattle and goats rarely have circulating reticulocytes.

 


RBC maturation. Image adapted from Veterinary Haematology, A Diagnostic Guide and Colour Atlas, John W. Harvey.

 

  • White Blood Cells (WBCs): These include:
    • Neutrophils: Mature from myeloblasts through promyelocytes, myelocytes, metamyelocytes and bands, becoming segmented neutrophils. This process takes 6–9 days, with bone marrow storing ~7x the number of circulating neutrophils.
    • Eosinophils and Basophils: Similar maturation pathways to neutrophils, with lineage distinctions becoming apparent at the myelocyte stage. Interleukin-5 released by activated T-helper cells drives eosinophil maturation.

 

Granulocyte maturation. Image adapted from Veterinary Haematology, A Diagnostic Guide and Colour Atlas, John W. Harvey.

 

    • Lymphocytes: Derived from common lymphoid progenitors. B cells mature in the bone marrow, while T cells migrate to the thymus. Both undergo rigorous selection to ensure self-tolerance; failures here can lead to immune-mediated diseases.

 

  • Platelets: Produced by megakaryocytes in the bone marrow, each generating 1,000–3,000 platelets. Their production is stimulated by thrombopoietin (TPO), and their appropriate function is critical for primary haemostasis.

 

What Does My CBC Tell Me?

The CBC provides quantitative data about the blood’s cellular components. Paired with a blood smear, it paints a complete picture of haematological health, offering insights into immune function, oxygen-carrying capacity, clotting ability, and bone marrow activity.

Key Parameters and Their Clinical Significance

  1. Red Blood Cell Indices

 

 

 

  • RBC Count, Haemoglobin (Hb), and Haematocrit (HCT):
    • HCT and PCV should give a similar result, but they are obtained differently. HCT is calculated by the analyser as ([RBC x MCV] / 10) and is therefore susceptible to artifacts affecting these indices (see below). PCV is the direct measure of the packed RBC fraction and is performed manually. This will still be affected by haemolysis (due to loss of the cells you are counting!)
    • These values indicate whether anaemia is present. HCT is typically ~3x Hb, and any mismatches exceeding 5% will be picked up and explored by your pathologist. Discrepancies are often caused by sample interference (e.g., lipemia or haemolysis).
    • Histogram analysis can identify whether analysers are misclassifying giant platelets as small RBCs or microcytes as platelets.
  • Mean Corpuscular Volume (MCV):
    • We use the Sysmex XN-V analyser. The Sysmex directly measures MCV using hydrodynamic focusing, forcing individual RBCs into a single file line which is passed through a laser beam. Increased MCV can indicate regeneration (reticulocytes are bigger than mature RBCs) or breed-specific macrocytosis (e.g., poodles). Decreased MCV may suggest iron deficiency, portosystemic shunts, or breed-specific traits (e.g., Akitas), and can also reflect in vitro cell shrinkage in hyponatraemic patients (think osmosis).
  • Mean Corpuscular Haemoglobin (MCH) & Mean Corpuscular Haemoglobin Concentration (MCHC):
    • MCH is the amount of Hb per red blood cell [(Hb/RBC) x10], and so does not account for the volume of the cell.
    • MCHC is calculated as [(Hb/HCT) x 100], which accounts for the cell volume (since HCT is calculated using MCV). Both MCH and MCHC are susceptible to the interferences which affect Hb and RBC count (e.g., lipemia falsely elevates Hb).
  • Red Cell Distribution Width (RDW):
    • Reflects anisocytosis. Elevated RDW often accompanies regenerative anaemia or recent blood transfusions, for example.
  • Reticulocyte Count:
    • A critical marker of bone marrow activity. In mild anaemia, even a “normal” reticulocyte count might signal regeneration. Manual reticulocyte counts are often necessary for cats due to the persistence of punctate reticulocytes; aggregate reticulocytes provide a clearer indication of recent bone marrow activity.
  • Nucleated RBCs (nRBCs):
    • Indicates accelerated turnover. Their presence in peripheral blood reflects the bone marrow’s attempt to meet increased demand, but it may also signify marrow disruption or extramedullary haematopoiesis.
  • Retic-He:
    • A marker of iron availability. This is an early warning for iron restricted haematopoiesis and would be low in developing iron deficiency anaemia. However, it is relatively non-specific and is commonly low in patients with neoplasia, immune mediated disease and inflammation.

 


RBC histogram created by direct current sheath flow detection. Cells which are between 40 and 150fL are classified as RBCs. The small peak to the left of the x-axis represents platelets.

 

 

  1. White Blood Cell Indices
  • Total WBC Count and Differential:
    • Many laboratory analysers will use more than one method to generate accurate total WBC counts, but nRBCs can falsely elevate these if not excluded manually. A blood smear review remains vital, and manual differential count would be performed if there are unusual changes such as neutrophil toxicity or atypical cells.
  • Neutrophil Count:
    • Neutrophilia suggests inflammation or stress (corticosteroid response). The presence of band neutrophils (left shift) signals increased inflammatory demand and can make it challenging for the analyser to accurately gate these populations.
  • Monocyte Count:
    • Elevated counts often accompany chronic inflammation but can also reflect stress.
  • Eosinophil Count:
    • Eosinophilia may point to parasitic, allergic, or paraneoplastic conditions.

 

WBC DIFF plot generated by the Sysmex XN-V. The plot on the left is from a healthy dog whilst the plot on the right is from a dog with marked toxic change and a left shift. The overlapping of the clouds indicates that the analyser is not able to confidently gate the different cell populations, and we should interpret the cell counts with caution.

 

 

  1. Platelet Parameters
  • Platelet Count:
    • Our analysers use two methods to count platelets: impedance (PLT-I) and optical (PLT-O). PLT-O uses fluorescent markers to identify platelets and tends to better mitigate the effects of platelet clumping, which can cause pseudothrombocytopenia. A blood smear review is essential to confirm platelet numbers and evaluate morphology.
    • Differentials for thrombocytopenia include consumption (usually mild reductions), immune-mediated destruction (marked reductions) and decreased production (e.g. bone marrow or infectious diseases).
  • Platelet Volume (MPV):
    • Larger platelets suggest increased turnover, as seen in recovery from thrombocytopenia or bone marrow stimulation.

 

PLT histogram illustrates the impedance method. Cells which are between 10 and 40fL are classified as PLTs. The large peak to the right of the x-axis are RBCs.

 

Why Is the CBC Useful?

A CBC helps to answer critical questions like:

  • Is the anaemia regenerative or non-regenerative?
    • Regenerative anaemia favours underlying haemorrhage or haemolysis (e.g. IMHA).
    • Non-regenerative anaemia often indicates chronic disease, iron deficiency, or bone marrow disorders (e.g., precursor-targeted immune-mediated anaemia [PIMA]).
  • What’s driving the leukocyte changes?
    • Inflammatory leukograms feature a neutrophilia and monocytosis, often with toxic change and left shifts. Stress leukograms may have a concurrent lymphopenia, and toxic changes are not present. Eosinophilia prompts investigation for parasites, allergies, or neoplasms.
  • What about platelets?
    • Mild thrombocytopenia could reflect consumption, whilst severe thrombocytopenia raises suspicion for immune-mediated thrombocytopenia (IMTP). Thrombocytosis might accompany inflammation or iron deficiency.
  • Are multiple lineages affected?
    • Bicytopenia or pancytopenia signals potential bone marrow involvement. Neutropenia and thrombocytopenia typically precede anaemia due to the longer lifespan of RBCs. Bone marrow aspiration is helpful to identify whether the marrow is responding appropriately, and may reveal underlying causes, such as leukaemia, infection (e.g., Ehrlichia, Leishmania), or immune mediated destruction (e.g., PIMA).
    • Concurrent anaemia and thrombocytopenia can create a chicken or egg dilemma – which came first? This is why a detailed clinical history is vital.

Personal Reflections on the CBC

The CBC is a powerful diagnostic tool, but the analyser is not foolproof. Throughout my residency I have learned to identify and flag erroneous results caused by factors such as RBC agglutination, hyponatraemia or sample lipemia. Identifying these unusual results and working out the mechanisms behind the error has encouraged me to gain a deeper understanding of how haematology analysers work, as well as the importance of assessing dot plots and the meaning behind the lesser-known indices which are often overlooked. Each case builds a richer understanding of the nuances in haematology.

Final Thoughts

Interpreting a CBC is both an art and a science. Each value tells a story, and together, they reveal the body’s dynamic response to health and disease. By combining analyser results, blood smear reviews, and clinical context, we can uncover the deeper narratives within our patients and provide them with the best possible care.

Next time you receive a CBC report, take a moment to appreciate the wealth of information it offers. And remember, when in doubt, look at the smear — it might just surprise you.

 

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Articles, Case Studies

Case Study: 15-Year-Old Domestic Shorthair Cat with Multiple Myeloma

Signalment

A 15-year-old domestic shorthair cat, neutered male, presented to the clinic for evaluation of lethargy, weight loss, and reduced appetite. The cat had a known history of heart failure, which was managed with medication. During physical examination, mild dehydration was noted along with pallor of the mucous membranes. There was no evidence of lymphadenopathy, however osteolytic bone lesions of the spinal vertebrae were present on radiography.

Clinical Presentation and Laboratory Findings

The initial laboratory workup revealed several abnormalities. The cat had a marked hyperglobulinemia with a globulin level of 160 g/L, accompanied by a markedly low albumin-to-globulin (AG) ratio of 0.1, indicating a hyperproteinaemia state driven by an elevated globulin fraction. Blood tests also revealed moderate azotaemia, which could be a result of renal impairment either due to hyperproteinaemia or as an independent comorbidity in this older cat. Additionally, a moderate neutrophilia was noted, alongside mild non-regenerative anaemia, which is often associated with chronic inflammation or neoplasia.

Cytologic Findings

Fine-needle aspirates were taken from the liver and spleen to further investigate the hyperglobulinemia and other laboratory abnormalities. Cytologic evaluation of the spleen revealed a large population of discrete cells with moderate pleomorphism, eccentric nuclei, and deeply basophilic cytoplasm. These cells exhibited moderate anisocytosis and anisokaryosis and occasional binucleation. Occasional mitotic figures were seen. A similar cytologic picture was observed within the liver. The cytological findings were consistent with plasma cell neoplasia.

 

 

Diagnosis and Discussion: Plasma Cell Neoplasia in Cats

Multiple myelomas are plasma cell neoplasms that originate in the bone marrow and infiltrate other organs, whilst solitary plasma cell tumours in organs other than bone marrow are plasmacytomas.  Neoplastic plasma cells secrete abnormal immunoglobulins which appear as a monoclonal spike on protein electrophoresis. Diagnostic criteria for multiple myeloma, initially outlined by MacEwen and Hurvitz in 1977 for dogs, require at least two of four specific indicators: (1) paraproteinemia or monoclonal gammopathy, (2) radiographic evidence of osteolytic bone lesions, (3) Bence Jones proteinuria (abnormal immunoglobulin light chains in urine), and (4) greater than 5% neoplastic plasma cells in the bone marrow. In this case, cytologic evidence of an atypical plasma cell proliferation in both the liver and spleen, alongside a marked hyperproteinaemia and presence of osteolytic lesions is strongly suggestive of multiple myeloma, although bone marrow samples were not obtained for confirmation.

Clinical signs of multiple myeloma are non-specific and can include lethargy, anorexia and weight loss. Initial work up may reveal renal impairment, anaemia and osteolytic lesions. However, osteolytic bone lesions, a hallmark of multiple myeloma in dogs, are less frequently observed in cats. An azotaemia (as seen in this case) may reflect existing renal compromise or reduced glomerular filtration rate secondary to hyperviscosity syndrome.

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