Cytology cases

Kathleen Tennant BVetMed Cert SAM Cert VC FRCPath MRCVS

Clinical LeadDiagnostic Laboratories

Langford Veterinary Services

Aims

• Case based • Mixture of levels• Some recurring problems• Practice ‘screening’ slides

First, the set-up

• A major reason many people won’t screen their cytology is because they can’t see down the microscope properly....

Which of these would apply to you?

1. Microscope’s always covered in oil2. Forgotten what I’m supposed to do with

the condenser3. Our Diff-Quik’s scummy and won’t stain

right4. All of the above5. None of the above

Quick fixes

Microscope’s always covered in oilClean it! Ethanol, methanol, not acetone. You can take

the lenses off to make it easier but be careful not to get anything inside it...

Forgotten what I’m supposed to do with the condenserUp – just below the stage, if it will move, usually openOur Diff-Quik’s scummy and won’t stain rightYou can scrape the scum off the last one with filter paper

– use small narrow upright jars so you are more willing to throw it away

Which of the following can not be successfully used as immersion oil?

1. Liquid paraffin2. A mix of two different commercial

immersion oils3. Perfume grade cedar oil4. Coverslipping glue

Immersion oils

• You cannot mix different oil types – each has its own refractive index and a combination will be impossible – clean the oils off both the lens and the slide to fix it

• Coverslipping glue works suprisingly well for approximately 20 seconds. Takes about 2 hours to mend the microscope (sorry, Nikon technician....)

Which of these is not a ‘cytology sin’?

1. Putting thick samples in a slide holder before they are dry

2. Putting the slides in the same bag as the histopath sample

3. Staining a slide with Diff-quik for a look and then sending it to an external lab

4. Staining a slide with Diff-quik then not including it with the rest of the slides to go out

Cytology ‘sins’

• Slides going into holders stay wet in the humid environment for a long time and this can affect morphology

• Even a small amount of formalin affects the way the cells stain and drastically cuts down detail

• External labs prefer to have all slides – the one that you have stained may be the only one with material on it

Case 1

• 5 year old Great Dane from a rescue centre with cough, pyrexia and anorexia

• Radiographs show widely distributed alveolar pattern

• Tracheal wash taken as initially unresponsive to antibiotics

• Gelatinous material retrieved – direct smears made

Direct smear, 100x lens oil immersion – what is this?

Is this/are these....?

1. Bacteria2. Stain precipitate3. Necrosis4. Inflammation5. KY gel6. Mucus

KY/ ultrasound gel

• Pink to deeply purple staining, irregular flecked material

• Stain precipitate looks very similar but (hopefully) should not be this thick

Mucus

Case 1

• Same dog, second sample (one reason it’s good to screen a sample before waking up – can hair dry them if in a hurry, pre- and post staining)

Tracheal wash # 2

Is this...

1. Normal2. Neutrophilic inflammation3. Septic4. Mast cell metastasis

Normal tracheal wash

• When screening your samples, consider what the normal cell population in that area should be

• Airways have regular columnar epithelial cells – sometimes the cilia are visible

• Goblet cells (mucus producing) look very similar to mast cells

Tracheal wash # 2

Same dog, BAL left

Closer up

What’s your diagnosis?

1. Fungal pneumonia2. Gram positive bacterial pneumonia3. Gram negative bacterial pneumonia4. Mixed bacterial pneumonia

Same dog, BAL left, neutrophils

Septic neutrophilic inflammation

• The demonstration of organisms INSIDE neutrophils is needed to confirm sepsis

• Use neutrophil size to compare organisms to – bacteria rarely > 1/20th neutrophil diameter

• Don’t forget that Diff-Quik tells you nothing about the Gram characteristics of bacteria...

• Neutrophils will often degenerate in the presence of bacterial toxins, or irritants such as pancreatic fluid, bile or urine

Non – degenerate/ degenerate

Aspirate from a nodule on a 2 y.o. DSH’s head

What’s your diagnosis?

1. Fungal infection2. Gram positive bacterial infection3. Gram negative bacterial infection4. Mycobacterial infection5. Mast cell tumour

Aspirate from a nodule on a 2 y.o. DSH’s head

• The cell here is a macrophage

• The white (non- staining) linear stripes within it are Mycobacteria

• Macrophages are very good at ‘presenting’ the cause of some infections – keep an eye out for what’s inside them

Peritoneal fluid from an ascitic dog

• The gross appearance of the fluid is serosanguinous

• The nucleated cell is a macrophage

When did the bleeding occur?

1. Current/ iatrogenic2. Minutes to hours ago3. Last week

Peritoneal fluid from an ascitic dog

• There are no platelets present, which makes current/ iatrogenic bleeding less likely (not impossible)

• The macrophage has had time to phagocytose a red cell – minutes to hours

• Might expect a week old bleed to have cleared, or macrophages to contain haem pigment breakdown products

Recent haemorrhage - cytology

Long standing haemorrhage - cytology

Longer standing haemorrhage

Tracheal wash from a 7 y.o. Coughing Yorkshire Terrier

Is this...

1. Normal2. Squamous cell carcinoma3. Oropharyngeal contamination and

neutrophilic inflammation4. Septic neutrophilic inflammation

Oropharyngeal contamination

• Oropharynx contains nucleated squamous cells

• The ‘striped’ organisms are Simonsiella, which are particular to the oropharynx

• Neutrophilic inflammation also present – but interpreting any culture results will be difficult

Pollen and mucus

• Other ‘interlopers’ in airway washes include plant, food and pollen material

• Anything green on a stained slide has to be self pigmented and is often of plant origin

Nasal flush from a 3y.o. GSD

Is this...

1. Bacterial infection2. Yeast/fungal infection3. Plant contamination4. Nematode infection

Yeast infection

• Yeast are generally larger than bacteria and often have a non- staining capsule

• Look for septae• Aspergillus usually large

and may have long hyphae

Aspergillus

Yeast from laboratory cactus!

BAL from a 4y.o. Coughing cat

Similar BAL from a coughing dog

Is this...

1. Eosinophilic inflammation2. Basophilic inflammation3. Neutrophilic inflammation4. Normal

Eosinophils

• Round granules in dogs, elongated granules (more difficult to see) in cats

• Look for potential causes

• Pulmonary infiltrate with eosinophils in dogs, ‘asthma’ in cats

Angiostrongylus

• Note the size of the organism versus the dots in the background, which are neutrophils

• Neutrophilic inflammation can be associated with this as much as eosinophils