Dogs – CRP as the Gold Standard

As a major APP, C-reactive protein (CRP) increases within 4–24 hours following the triggering insult, rising up to 50–100-fold, reaches its peak after 1–2 days, and declines rapidly with effective therapy.
Due to this dynamic behaviour, CRP is particularly well suited for early detection, monitoring disease progression, and assessing therapeutic success.

Elevated CRP levels are observed in a wide range of inflammatory and immune-mediated processes, including bacterial infections, parasitic diseases, autoimmune disorders, neoplasms, and post-traumatic or postoperative changes. In dogs with acute Babesia canis infection, CRP shows a strong correlation with clinical severity and haematological parameters.

Within the framework of antimicrobial stewardship, it has been shown that antibiotics can be discontinued once clinical improvement is observed and CRP concentrations have returned to normal, which significantly shortens the treatment duration for many diseases.

In systemic mycoses, such as pulmonary coccidioidomycosis, CRP in combination with haptoglobin (Hp) also demonstrated predictive value for remission. However, CRP can be elevated in the absence of inflammation, for example during extreme physical exertion or pregnancy, so its interpretation must always take the clinical context into account.

 

Acute-Phase Index (API) – a Combined Marker

Recent research has combined positive APPs (CRP, Hp) and negative APPs (albumin, optionally PON-1) into an acute-phase index (API). This index reflects the overall activity of inflammation. Dogs with malignant tumours and a high API had a significantly poorer prognosis.

 

Table 1: Overview of changes in acute-phase proteins and leukocyte counts over time following an inflammatory stimulus in dogs and cats.

Time after Inflammatory Stimulus	Dog – e.g., CRP, SAA	Dog – Leukocyte Count	Cat – e.g., SAA, AGP	Cat – Leukocyte Count
0–6 h	slight increase at the start (hepatic synthesis begins within a few hours)	usually still within the reference range	slight increase at the start	usually still within the reference range
6–12 h	marked increase measurable	first tendency to rise possible, often still borderline	marked increase measurable	first tendency to rise possible, often still unremarkable
12–24 h	strong increase, values usually clearly pathological	leukocytosis or leukopenia often visible	strong increase, clearly pathological	more frequent leukocytosis, sometimes stress leukogram
24–48 h	peak of APR – highest concentrations	further increase or plateau of leukocytes	peak of APR	further increase or plateau of leukocytes
2–5 days	beginning decline if inflammation is controlled	leukocytes often still elevated, slowly decreasing	decline with clinical improvement	leukocytes often still altered, normalising more slowly
> 5 days	return to or near reference range in resolving inflammation	normalisation, but may take longer with chronic processes	similar to dogs	similar to dogs

APR = acute-phase response, CRP = C-reactive protein, SAA = serum amyloid A, AGP = α1-acid glycoprotein

 

In chronic inflammatory diseases, such as canine leishmaniasis, CRP and Hp remain persistently elevated, while albumin and transferrin often decrease. Changes in the API correlate closely with treatment response and disease activity. Persistently high values indicate residual activity, co-infections, or treatment failure.

 

Cats – Focus on SAA and AGP

In cats, the dynamics and significance of APPs differ considerably from those in dogs. Serum amyloid A (SAA) is the most important major APP, while α1-acid glycoprotein (AGP) has particular diagnostic value in feline infectious peritonitis (FIP).

 

Serum Amyloid A (SAA)

SAA responds very early and sensitively, reaching high concentrations quickly, making it suitable for both early diagnosis and prognostic assessment. A rapid decline indicates a good response to therapy, while a stagnating value suggests persistent inflammation or secondary infection. Modern turbidimetric assays using monoclonal antibodies provide high diagnostic precision. Further studies demonstrate the usefulness of SAA, especially in bacterial infections such as pyelonephritis.

 

Alpha-1-acid Glycoprotein (AGP)

AGP is a moderately rising APP with high clinical relevance for FIP (Fig. 1). Elevated AGP serum levels support the presumptive diagnosis when interpreted alongside other findings. In particular, during antiviral therapy, AGP shows dynamic changes.

 

Haptoglobin (Hp)

In both dogs and cats, Hp is a moderate acute-phase protein synthesised in the liver. Its primary biological function is the high-affinity binding of free haemoglobin (Hb) from lysed erythrocytes, which reduces oxidative tissue damage and prevents the loss of Hb-bound iron. During acute inflammatory processes, both species exhibit a less pronounced and delayed increase in Hp concentration compared with major acute-phase proteins such as SAA or CRP. As in other mammals, intravascular haemolysis can lead to a decrease in haptoglobin concentration because the protein is rapidly consumed through binding large amounts of free haemoglobin.

 

Negative Acute-Phase Proteins

Albumin
Albumin decreases due to the redistribution of amino acids for the synthesis of positive APPs and increased capillary permeability. It is a valuable indicator of systemic inflammation but must be interpreted in the context of hydration status, protein loss, and liver function. In dogs, albumin is included in the calculation of the acute-phase index (API).

Transferrin
Transferrin, an iron-binding transport protein, decreases during the acute-phase response to reduce iron availability for microorganisms. In dogs, a marked decrease in transferrin has been observed during bacterial infections. In cats, a significant decline has also been documented in cases of chronic inflammation.

 

Acute-Phase Proteins in FIP

Feline infectious peritonitis (FIP) is an inflammatory disease that is generally associated with an increase in acute-phase proteins. Studies have shown that measuring AGP in effusions is the most informative method for distinguishing between cats with and without FIP. Different cut-off ranges with varying sensitivity and specificity have been defined (Table 2). Some of these cut-offs, for example in Helfer-Hungerbühler et al. (AGP > 2927), exhibit high specificity (97 %) and can therefore be strongly indicative of FIP. However, due to the relatively low sensitivity (54 %), almost half of the cats with FIP may not be detected. It should also be noted that APPs can increase in other diseases. Cats with a septic abdomen or disseminated neoplasms often show AGP concentrations similar to those observed in cats with FIP. Therefore, complementary cytological and bacteriological examinations are important to exclude differential diagnoses.
Measurement of AGP alone is insufficient for a definitive diagnosis and should be considered as one of many components in the diagnostic process.
AGP may also play an important role in therapy monitoring for cats with FIP. During treatment, AGP gradually decreases, although more slowly than SAA. This is likely due to the longer half-life of AGP, which means that AGP concentrations on day 2 of FIP therapy can be higher than before treatment began. A significant decline in AGP was observed from day 7 after the start of therapy. By day 28, AGP levels were within the normal range in almost all cats (Helfer-Hungerbuehler 2024: 17 of 18 cats, Zuzzi-Krebitz 2024: 37 of 39 cats), making it a reliable parameter for monitoring treatment success. In contrast, SAA showed a significant decline already by day 2, and most cats reached (almost) normal SAA concentrations within 4–7 days. Addie and colleagues (2022) use AGP as a marker to differentiate between remission and recovery. Recovery refers to complete healing from FIP, while remission is defined as an intermediate stage between recovery and death, still carrying a risk of relapse. Cats that fully recovered showed AGP values within the normal range, whereas cats in remission exhibited elevated AGP levels.
Therefore, a rise in AGP could also indicate a potential FIP relapse.

 

Table 2: Overview of recent publications on the use of AGP. Measured median values (including range) in cats with FIP compared with cats without FIP, defined cut-off values, and their sensitivity and specificity.

Study	Acute-Phase-Protein	Median Value in Cats with FIP (Range)	Median Value in Cats without FIP (Range)		Cut-Off	Sensitivity (%)	Specificity (%)
	Serum
Hazuchova 2017	AGP (µg/ml)	2900 (960-5040)	690 (120-4500)		2260	85	90
	SAA (µg/ml)	98,5 (1,3-163,4)	7,6 (0,1-163,8)		97,3	55	87
	Hp (mg/ml)	2,0 (2,0-9,0)	1,8 (0,0-2,0)		2,0	55	82
	Effusion
	AGP (µg/ml)	2570 (1300-5760)	480 (190-3800)		1550	93	93
	SAA (µg/ml)	80,4 (0,1-207,4)	0,1 (0,1-182,7)		43,6	71	91
	Hp (mg/ml)	2,2 (0,1-9,3)	0,8 (0,1-2,5)		2,1	79	87
	Serum
Helfer- Hungerbuehler 2024	AGP (µg/ml)	2954 (200-5861)	healthy	sick	2531	61	79
			235 (78-616)	1734 (305-3449)	2927	54	97
	Effusion
	AGP (µg/ml)	2425 (343-5611)	560 (83-3950)		1686	71	89
	Serum
Romanelli 2024	AGP (µg/ml)	1986 (405-4428)	296 (165-4254)		707	80	80
					>4099	–	100
					<438	100	–
	Effusion
	AGP (µg/ml)	1717 (549-3166)	233 (103-4099)		990	75	73
					>4254	–	100
					<296	100	–

 

 

Dr. Ruth Klein, Katharina Buchta

Survey-based Study at Laboklin

The aim of the study was to identify the reasons for discontinuation of ASIT in horses during or after the induction phase (initial treatment, starter set).
The induction phase lasts approximately six months, which is considerably shorter than the recommended 12-month period for assessing the response to therapy.

Horses were selected from the ASIT treatment order lists of the Laboklin laboratory for the years 2021–2023, specifically those for whom no further ASIT treatments were ordered after the starter set. Of 4,271 initial treatments, 1,475 cases (34.5 %) did not receive any follow-up treatments.
To determine the reasons for discontinuation of ASIT, the treating veterinarians were contacted using written questionnaires. They could select one or more possible reasons. The collected data were analysed descriptively.

 

Reasons for ASIT Discontinuation

A total of 171 responses reporting 204 reasons for why no follow-up ASIT treatment was ordered after the initial treatment were analysed. Patients who did not receive further ASIT due to death (n = 15) were excluded from the analysis.
The horses included in the study exhibited the following symptoms: asthma (n = 68, 39.8 %), pruritus (n = 40, 23.4 %), urticaria (n = 3, 1.8 %), headshaking (n = 3, 1.8 %), or a combination of these symptoms (n = 31, 18.1 %). In 26 horses (15.2 %), the specific symptoms were not reported (Fig. 1).

The most common reasons for discontinuation of ASIT (Fig. 2) were loss of contact with the owner (n = 55, 27 %), lack of owner compliance (n = 40, 19.6 %), insufficient treatment response (n = 39, 19.1 %), or a good treatment response (n = 27, 13.2 %).

These four reasons accounted for over 80 % of treatment discontinuations. Other reasons included cost or sale of the horse (each n = 13, 6.4 %), adverse effects (n = 4, 2 %), and lack of knowledge regarding continuation of ASIT (n = 4, 0.5 %).

ASIT was also discontinued due to symptom improvement following a change of stable or optimisation of management (n = 7, 3.4 %) or a dietary change (n = 1, 0.5 %). In three horses (1.5 %), the induction phase had not yet been completed due to protocol adjustment, and in one horse (0.5 %), ASIT was continued but the ASIT manufacturer was changed.

In 116 questionnaires, it was reported that ASIT was administered according to the manufacturer’s protocol, while in four horses the protocol was individually adjusted. No additional symptomatic therapy was given in 93 horses; 11 horses received mucolytics alongside ASIT, 15 underwent inhalation therapy with glucocorticoids and bronchodilators, and six patients were treated with systemic glucocorticoids. In eight horses, it was reported that symptoms worsened again after ASIT was discontinued.

 

How Can ASIT Discontinuation Be Reduced and Treatment Success Optimised?

Induction treatments usually last for six months, which is considerably shorter than the recommended 12-month period for evaluating therapeutic success.

It can take up to a year for the full clinical benefit of ASIT to become apparent. In this Laboklin study, follow-up treatment was not ordered for over one third of initial treatments, resulting in premature discontinuation of ASIT after the induction phase.

 

Loss of Contact with Owners and Lack of Owner Compliance
The most common reasons for ASIT discontinuation were loss of contact between veterinarian and owner (27 %) and lack of owner compliance (19.6 %), together accounting for nearly 50 % of all discontinuations. In a Laboklin study on ASIT discontinuation in dogs, these were likewise the most frequently reported reasons. Owner compliance – the cooperation of the owner in implementing the recommended therapeutic measures – is a crucial factor for treatment success. Owners should be thoroughly informed about the treatment protocol, duration of therapy, the delayed onset of ASIT effects, and the expected costs, in order to align their expectations appropriately. Continuous communication is key to ensuring good owner compliance, particularly during the first year of therapy. Regular check-ups or telephone contact with owners not only maintain communication but also ensure continuous monitoring of the patient.

 

Treatment Success Lower than Expected
The third most common reason for ASIT discontinuation was insufficient therapeutic response (19.1 %). This was often reported in combination with poor owner compliance. Due to the delayed onset of ASIT effects, treatment success should be evaluated no earlier than one year after initiation.
Discontinuation of ASIT due to perceived lack of efficacy after the induction phase represents an issue of owner education, as these horses were prematurely classified as non-responders.
Veterinarians should clearly communicate the delayed onset of ASIT effects in order to manage owner expectations and prevent premature discontinuation during the first year of therapy.
In the current study, 93 horses reportedly did not receive any additional therapy, while only 15 horses were treated symptomatically during the induction phase of ASIT to alleviate allergic signs. Symptomatic treatments – such as glucocorticoids, antihistamines, bronchodilators, and mucolytics (in cases of equine asthma) – are often necessary during the first months of ASIT to rapidly reduce clinical symptoms until the effects of the immunotherapy take hold.
This approach is also an important factor in improving owner compliance. The duration and dosage of medications should be kept as low as possible. Symptoms should be reduced but not entirely suppressed, as complete suppression may obscure the need for protocol adjustment.
It is also important to correctly define treatment success. ASIT is considered successful if treated horses show an improvement of more than 50 % in clinical symptoms, or if the need for additional symptomatic medications can be reduced by more than 50 %. Before a horse is classified as a non-responder, it should be carefully evaluated whether there is truly no improvement. This requires precise documentation of the frequency of allergic episodes, as well as the duration and dosage of any additional symptomatic therapy. In this study, four respondents reported that ASIT was discontinued due to insufficient effect, but symptoms subsequently worsened after cessation. In these cases, it can be assumed that ASIT did provide clinical improvement that was not adequately documented, leading to the erroneous classification of these horses as non-responders.
The average success rate of ASIT in horses, according to a review by Herrmann et al. (2023), was 75 % for equine asthma, 88 % for urticaria, 59 % for pruritic dermatitis, and 36 % for sweet itch (ASIT using insect allergens only). Limited data are available regarding the efficacy of ASIT for allergy-in-duced headshaking; one study reported good to very good responses in five of the six horses included.
The reason why horses treated solely with insect allergens show lower success rates is not fully understood. One possible explanation is that, due to the simultaneous presence of different allergens, it is clinically difficult to distinguish between environmental pollen allergies and sweet itch (insect bite hypersensitivity). Many horses are polysensitised, and in these cases, additional allergens besides insects should be included in ASIT. Another theory is that insect allergens in ASIT may induce a weaker immune response compared with other allergens. This could be related to the fact that only whole-body extracts are currently available for ASIT, whereas treatments using pure insect salivary proteins might be more effective. This hypothesis is supported by studies using recombinant allergens in ASIT. A recent study by Graner et al. (2024) treated horses with ASIT consisting of recombinant Culicoides allergens. Clinical response was significantly higher than in the placebo control group, with almost 90 % of treated horses showing at least a 50 % improvement in symptoms in the second year of treatment.
The duration of the disease may also influence the success rate of ASIT. Hunsinger (2003) reported that horses treated with ASIT within two years of the onset of allergic symptoms responded significantly better to therapy. In horses with sweet itch, the success rate of ASIT was 75 % when treatment was initiated within the first two years after disease onset. This rate decreased considerably when the start of ASIT was delayed.
Another measure to optimise treatment success may be the individual adjustment of the protocol regarding injection volume and/or treatment intervals. In this study, protocol adjustment was reported in only four horses; all other patients were treated according to the manufacturer’s protocol.
Continuous patient monitoring is necessary to identify the need for protocol adjustment.

 

Discontinuation despite successful response
The fourth most common reason for stopping ASIT (13.2 %) was clinical improvement in the treated horses, leading to interruption of therapy. Most patients require long-term, often lifelong treatment to maintain control of allergic symptoms. Experience shows that symptoms typically recur after ASIT is discontinued. This was also observed in the present study in two patients whose ASIT was not continued due to marked improvement. Restarting ASIT can be demanding, often requiring repeated allergy testing, initiation of a new induction phase, and in some cases, a less favourable therapeutic response. Therefore, it is generally recommended not to interrupt ASIT in responders and to inform owners of the need for ongoing treatment. If clinical improvement is stable over several years during the maintenance phase, injection intervals may be gradually extended up to eight weeks.

 

Costs
In this study, costs were cited as the reason for ASIT discontinuation in 6.4 % of cases. For owners, the expenses associated with ASIT during the first year—including allergy testing and regular veterinary check-ups—may appear high.
However, in the long term, ASIT is considerably more cost-effective than purely symptomatic therapy, which can be very expensive in horses. Poorly controlled allergic horses often require more frequent veterinary visits, higher doses of symptomatic medications, and additional diagnostic or therapeutic interventions to manage potential side effects of these treatments. Providing this information can help motivate owners to continue ASIT throughout the first year of treatment and, during the maintenance phase, even in cases of moderate clinical response.

 

Side effects
In general, ASIT can be considered very safe in horses. In the present study, ASIT was discontinued due to adverse effects in four horses (2 %). In all cases, allergic symptoms worsened following the injections; additionally, one horse developed diarrhoea and another experienced circulatory problems. Worsening of allergic signs immediately after injections is one of the most common adverse effects, which was also observed in this study.
If this side effect occurs, the allergen extract dose should be reduced and the induction protocol individually adjusted. The importance of continuous communication between veterinarians and owners should be emphasised with regard to adverse effects. Owners should carefully observe their horses’ reactions to injections and immediately report any issues to the treating veterinarian to allow appropriate adjustments to the ASIT protocol.
The Laboklin team is available for consultation regarding protocol modifications.
Anaphylactic reactions (urticaria, angioedema, respiratory distress, circulatory collapse) are very rare. One horse in the current study experienced circulatory problems, leading to discontinuation of ASIT. The most common adverse effect during the induction phase is a self-limiting local reaction at the injection site; however, this was not cited as a reason for treatment discontinuation.

 

Conclusion

In summary, ASIT represents an important component of the multimodal management of allergic horses and is a lifelong therapy that requires close collaboration between owners and veterinarians. The first year of treatment necessitates intensive monitoring and constitutes the critical period for achieving therapeutic success. The most common reasons for discontinuation of ASIT are loss of contact with owners, poor owner compliance, and overly high expectations regarding rapid treatment effects. Improved education and communication, regular check-ups, and strict adherence to ASIT guidelines can increase the number of horses that respond successfully to ASIT and derive long-term benefit from the therapy.

 

Dr. Elisabeth Reinbacher

 

Our Services for Equine Allergies

• Screening tests
• Main tests for allergen differentiation (seasonal allergens, perennial allergens, insects, feathers/ hair/dander, feed)
• Screenings (allergy profiles – skin, allergy profile – respiratory)
• PAX complete (environmental allergens and/or food)
• Allergen-specific immunotherapy (ASIT)

The Diagnostic Pitfall: NTI and Methodological Limitations

In a typical case, the diagnosis of canine hypothyroidism is based on the combination of compatible anamnesis, clinical signs, haematological and biochemical findings, together with a T4 concentration below the reference interval and a simultaneous increase in TSH.

However, this pattern is not present in all cases in which hypothyroidism is suspected.
Many diseases can secondarily suppress thyroid hormone concentrations without any functional and/or structural thyroid abnormality. This so-called euthyroid sick syndrome (ESS or NTIS) is characterised by low T4 and/or fT4 concentrations with normal TSH. Interpreting such findings is problematic without further diagnostic work-up.

Additional challenges arise from breed-specific lower iodothyronine concentrations (e.g. Greyhounds, Basenjis) and from the interference of binding proteins or autoantibodies (thyroglobulin-, T3-, T4-autoantobodies) with routinely used immunoassay kits. Moreover, various medications (e.g. glucocorticoids, phenobarbital, sulphonamides) can also influence thyroid parameters.

 

LC-MS/MS: A Potential New Reference Method

LC-MS/MS has been established as a highly precise analytical technique. Unlike immunoassays, LC-MS/ MS enables the direct and specific quantification of T4, T3 and rT3 without interference from autoantibodies or medications. This makes it particularly valuable in diagnostically challenging cases. For example, while immunoassays can yield falsely low or falsely high results in the presence of thyroglobulin autoantibodies, LC-MS/MS provides a reliable representation of the actual hormone concentration.

At Laboklin, a method for the measurement of T4, T3 and rT3 using LC-MS/MS has been established and validated for veterinary medicine as part of a doctoral thesis. This allows for highly specific diagnostics, particularly in unclear cases. It can also be a reliable option for therapy monitoring or in cases of suspected hypothyroidism despite T4 concentrations within the reference range.

When interpreting T3 and T4 values measured by LC-MS/MS the method-specific reference intervals must be taken into account, as they differ from those obtained using other techniques.

 

Reverse T3 (rT3): Differentiating NTI from Hypothyroidism

rT3 is primarily produced when there is an excess of T4 or when conversion to active T3 is downregulated. In hypothyroidism, only minimal amounts of T4 are available for conversion, resulting in reduced rT3 concentrations. In contrast, NTI typically leads to normal or elevated rT3 levels: sufficient T4 is available, but the physiological demand for T3 is simultaneously markedly reduced (Fig. 1).

Initial studies confirm the diagnostic value of this parameter:

• rT3 < 50 pg/ml, in combination with a low T4 concentration, strongly suggests hypothyroidism.
• rT3 > 109 pg/ml makes hypothyroidism highly unlikely, even when T4 levels are reduced (Fig. 2).

 

Indication

This parameter is particularly valuable when T4 or fT4 concentrations are low, but no elevated TSH concentration is present to confirm the suspicion of hypothyroidism. This situation may occur in 20–30% of dogs with hypothyroidism, but it more frequently indicates a non-thyroidal illness (NTI).

 

Previous Experience with the Parameter

An initial multicentre study on rT3 was presented at an international congress (ECVIM) in 2024.
The study demonstrated that healthy dogs, dogs with hypothyroidism, and those with low T4 concentrations secondary to a non-thyroidal illness (NTI) could be reliably distinguished. A cut-off of 50 pg/ml was identified as highly specific for the presence of hypothyroidism, whereas concentrations above 109 pg/ml were incompatible with hypothyroidism. Values between these thresholds represented a grey zone. To establish a reference interval for rT3, Laboklin conducted a study in a larger population of clinically healthy, euthyroid dogs. This study determined a reference interval of 109–533 pg/ml.

In addition to the conducted studies, highly valuable field data have now become available.

Between March 2024 and June 2025, the rT3 parameter was requested 3,052 times at Laboklin, of which 1,887 requests were for rT3 alone and 1,165 were performed as part of a profile or as an add-on to initially requested thyroid parameters.

Among the 491 cases with an rT3 concentration below 50 pg/ml, T4 was also measured in our laboratory in 289 cases. Of these, 92% of the dogs exhibited T4 concentrations below the reference range, confirming the close functional relationship between reverse triiodothyronine and thyroxine.

Only in 22 cases (7.6%) with an rT3 concentration below 50 pg/ml was the T4 concentration within the reference range. This phenomenon can primarily be attributed to the limitations of the immunoassays routinely used for T4 measurement (e.g., Immulite 2000; Siemens, Germany). Falsely elevated T4 results may arise due to the presence of autoantibodies or suboptimal sample quality (haemolytic or lipaemic samples). These interfering factors are well recognised and can complicate the interpretation of results in routine diagnostics.

In 570 dogs with simultaneously low T4 concentrations, rT3 levels were clearly within the normal range (> 109 pg/ml). Such a result does not support a diagnosis of hypothyroidism, but rather indicates a non-thyroidal illness (NTI). Of course, a single measurement of even highly specific laboratory parameters cannot definitively determine that these dogs did not actually suffer from hypothyroidism, but rather from a non-thyroidal illness (NTI). In all cases that we were able to review in personal discussions with the attending veterinarians, the assessment was, however, confirmed.

Previous studies and clinical experience suggest a major advance in canine hypothyroidism diagnostics. The parameter appears to fulfil its promise. It should be regarded as an additional tool in the “hypothyroidism” toolbox. rT3 is not intended as an initial screening parameter, but rather as an add-on to an existing thyroid profile. Accordingly, it should be interpreted in the context of clinical findings as well as other thyroid parameters (Fig. 3).

For further diagnostics, determination of thyroglobulin-, T3-, T4-autoantibodies is available through extended thyroid profiles. Another option is the measurement of T3 and T4 concentrations using liquid chromatography–tandem mass spectrometry (LC-MS/MS), which is not affected by interfering factors such as sample quality or autoantibodies, allowing for more specific and precise results.

 

Conclusion

The precise diagnosis of canine hypothyroidism requires more than the assessment of T4 and TSH alone. In particular, an rT3 measurement and LC-MS/MS-based analyses provide crucial information in cases of diagnostic uncertainty.

 

Dr. Jennifer von Luckner, Niklas Wiesner, Dr. Ruth Klein

 

Our Services Related to Hypothyroidism

• T4, TSH, fT4
• T3, fT3
• rT3
• Thyroid Profile (T4, fT4, T3, fT3, TSH, ATG, T4-AK, T3-AK)
• Hypothyroidism/NTI Profile (T4, fT4, reverse T3, TSH)
• Thyroid Monitoring (T4, TSH, Creatinine, SDMA, ALT, AP, Troponin I)
• Thyroid Mass Spectrometry (T4, T3, rT3 by HPLC-MS/MS)

Clinical Signs of Liver Disease

A large proportion of liver diseases are subclinical. This is due to the liver’s substantial capacity for cellular regeneration. Liver failure occurs only when the loss of hepatocyte function exceeds their regenerative capacity, which generally requires damage to around 80% of the organ. This highlights the importance of recognising and interpreting non-specific clinical signs at an early stage, such as lethargy, inappetence, icteric sclerae and mucous membranes, weight loss, reduced performance, or episodes of colic. Early identification of a mild hepatopathy is associated with a positive prognosis.

More severe signs include hepatic encephalopathy, hepatocutaneous syndrome, bleeding tendency, photosensitivity, and diarrhoea.

 

Diagnostics

The laboratory diagnostic evaluation of liver diseases in horses is primarily based on the interpretation of liver-specific enzyme activities and functional parameters in serum. Additional laboratory results provide valuable information on the extent of liver damage as well as on possible underlying systemic diseases. Often, further investigations such as sonography and liver biopsy are required to clarify the aetiology (Fig. 1).

 

Blood Tests – Enzymes

Blood tests often provide the first indications of liver disease through the detection of elevated liver enzyme activities. The parameters are distinguished according to their localisation within the liver (hepatocellular, biliary). They can also be classified based on whether they are liver-specific or ubiquitous.

1. Hepatocellular Enzymes

a. Glutamate dehydrogenase (GLDH)
GLDH is an enzyme specifically found in hepatocytes. It is rapidly released into the serum during acute, cellular liver damage.
Due to its short half-life (approximately 14 hours, with complete decline after 3–5 days), it is particularly useful for detecting acute hepatocellular damage and is considered the most sensitive marker.

b. Aspartate aminotransferase (AST)
AST is not liver-specific, as it is present not only in hepatocytes but also in muscle cells and erythrocytes. Isolated elevations should always be interpreted in the context of GLDH, creatine kinase (CK), and LDH.

c. Lactate dehydrogenase (LDH)
LDH is present in numerous tissues, including cardiac and skeletal muscle as well as erythrocytes. Because of this widespread distribution, its diagnostic value for the liver is meaningful only when interpreted alongside other enzymes.

 

2. Biliary Enzymes

a. γ-Glutamyltransferase (γ-GT)
γ-GT is a liver-specific enzyme with a half-life of 3–4 days, although in some cases it may remain elevated for several weeks. This enzyme is mainly produced in the bile duct epithelium. Increased serum activity occurs primarily in cholestasis and biliary diseases. While also present in the kidney and pancreas, it is mainly liver-specific. In sport horses, isolated γ-GT elevations (>50–150 U/l) can occur during intensive exercise.

b. Alkaline phosphatase (AP)
AP also shows increased activity in cholestatic processes but is less organ-specific, as it is present in bone, placenta, and intestine. In young animals, elevated AP due to bone growth should be considered physiological.

 

Proper interpretation of values is essential to guide further diagnostics or determine the optimal timing for re-testing. For γ-GT, in cases of mild elevation (Table 1), re-evaluation after 2–4 weeks, possibly including additional herd mates, is recommended to clarify a toxic or infectious aetiology and to assess disease progression.

 

Table 1: Classification of liver enzyme elevation by degree

Degree of Elevation
Mild	2–3 × upper cut-off
Moderate	4–5 × upper cut-off
High	10 × upper cut-off

 

 

Blood Tests – Liver Function

Changes in liver enzyme levels provide only limited information on disease severity, prognosis, or aetiology. Additional liver function parameters can help to better assess the extent of damage and prognosis. Bile acids constitute the primary parameter for assessment.

1. Bile Acids
   Bile acids are produced in hepatocytes, continuously secreted into the duodenum, and 90–95% are reabsorbed. When cell function is impaired, reabsorption is reduced or absent, leading to accumulation and increased serum concentrations. Values above 25 μmol/l are regarded as pathological and indicate a poor prognosis in chronic cases. In acute cases, high values are of less prognostic concern, but require close surveillance. Bile acids are a highly sensitive marker for functional liver insufficiency, particularly in chronic diseases.
2. Bilirubin
   Bilirubin is derived from degradation of haemoglobin, conjugated in the liver, and excreted via bile. Values above 75 μmol/l are associated with the characteristic yellow discoloration of the sclera and mucous membranes (icterus, jaundice). Differentiating between conjugated and unconjugated bilirubin in the blood allows the cause to be classified as pre-hepatic, intra-hepatic, or post-hepatic.
   A conjugated fraction of more than 25% indicates a hepatocellular or hepatobiliar origin. In chronic conditions, however, serum bilirubin concentrations may remain within the normal range.
3. Ammonia
   Elevated blood ammonia levels indicate advanced liver insufficiency and can lead to hepatic encephalopathy. Measurement is challenging due to its low stability (maximum 30 minutes).

 

Imaging / Biopsy

Ultrasonography of the liver is a useful supplementary diagnostic tool, even if only a portion of the organ is accessible for imaging due to anatomical reasons. Although many changes are diffuse, ultrasonography can still provide valuable information. The absence of pathological findings does not rule out liver disease. Liver biopsies are indicated when clinical signs and laboratory values do not allow a definitive diagnosis. Samples can be submitted for histology, bacteriology, and pathogen detection via PCR.

 

Aetiology

Intoxication

Intoxications are among the most common causes of hepatopathies. They can arise from, for example, mycotoxins or moulds in roughage, microcystins from algae-contaminated water, or excessive iron intake.

Particular attention should be paid to poisonous plants in pastures and hay. Although horses usually avoid them, ingestion can occur under certain conditions (e.g. in young animals, during feed shortages, in the presence of exotic plants, or when plant parts are fragmented).

Species of Senecio, such as ragwort, can lead to altered liver enzyme activities even in the early stages of intoxication, often without clinical symptoms. The toxic agents are pyrrolizidine alkaloids (PAs), which can cause irreversible liver fibrosis with chronic exposure.

LC-MS analysis of urine allows detection of senecionine/senecionine-N-oxide and indicates toxin exposure in the preceding hours to days. Additionally, roughage analysis (e.g., through agricultural testing facilities) is recommended.

 

Infections

Viruses

Viral hepatitis in horses is increasingly well studied. Of particular relevance are equine hepacivirus (EqHV) and equine parvovirus-hepatitis virus (EqPV-H).

EqPV-H is associated with Theiler’s disease – an acute hepatitis with fulminant liver necrosis and usually fatal outcome. Transmission is likely via blood products (e.g., tetanus antitoxin, stem cell products, equine plasma), and possibly via vectors. The virus is widespread worldwide, with seroprevalences in healthy horse populations (e.g., Germany, Austria) ranging between 15% and 34.7%, though only about 2% develop clinical disease. EqPV-H should be considered in the differential diagnosis when relevant symptoms are present.

EqHV, first described in 2012, can cause acute or chronic persistent infections. Symptoms range from weight loss, anorexia and jaundice to neurological abnormalities.

Both viruses are detectable by PCR in blood or liver tissue during the acute stage. Histopathological examination of liver biopsies can additionally help to assess the severity and prognosis of liver damage.

 

Bacteria

Bacterial liver disease is rare and usually secondary. When it occurs, it is often severe. It typically involves ascending bacterial infections, e.g., with Streptococcus equi or Staphylococcus aureus. In foals, liver abscesses can be caused by Rhodococcus equi and young animals may develop Tyzzer’s disease due to Clostridium piliforme. Clinically, bacterial hepatitis usually presents with jaundice, fever and colic. Pathogen detection can be performed on liver biopsy samples, which can be evaluated microbiologically and histologically. PCR-based detection is also an option.

 

Parasites

Parasitic liver damage occurs due to migrating stages of, for example, Strongylus spp. and Parascaris equorum. Liver fluke (Fasciola hepatica) is rare in horses but may occur in mixed pastures with ruminants or in wetland areas where water snails are present. Lesions can affect both liver parenchyma and bile ducts. In suspected cases, faecal samples can provide proof of infection; for some parasites (Fasciola hepatica, small strongyles/ small redworm), serology is more sensitive.

 

Conclusion

Hepatopathies are often detected late due to non-specific symptoms and subclinical cases. Diagnosis is based on the determination of liver enzyme activities and functional parameters, supported by diagnostic imaging techniques and, if necessary, liver biopsy. Toxic and infectious causes are most common, while environmental and dietary factors should also be considered when investigating the aetiology. Treatment efficacy is monitored through regular assessment of liver enzyme activities.
Recovery, however, can take weeks to months. As a preventive measure, depending on the aetiology, feed and pasture hygiene as well as evidence-based parasite control management can be helpful.

 

Dominika Wrobel-Stratmann, Dr. Svenja Möller,
Dr. Michaela Gentil

 

 

Our Services in Equine Liver Diagnostics

Profile	Parameters	Sample Material
Liver 1	AST, GLDH, γ-GT, bile acids	Serum
Liver 2	GLDH, AST, AP, γ-GT, total bilirubin, cholesterol, urea, bile acids, protein, albumin, globulins, albumin/globulin ratio, glucose, Na, K, Cl	Serum and NaFB
Hepatotropic Viruses	PCR: equine parvovirus, equine hepacivirus	Serum, EDTA blood, liver tissue
Parasite Profile	Flotation, SAFC, modified McMaster method	Faeces
Coagulation	PT, PTT, thrombin time, fibrinogen	Citrate plasma
Bilirubin	Total and direct	Serum, EDTA plasma, heparin plasma
Single Analyses
Bile Acids		Serum
Serum Protein Electrophoresis	Albumin, α-globulins, β-globulins, γ-globulins, total protein	Serum
Liver Fluke (Antibody Detection)		Serum
Small Redworm Test (Antibody Detection)	Detects infection levels of all small redworm stages	Serum
Meadow Saffron	Colchicine	Urine
Ragwort	Senecionine, senecionine-N-oxide	Urine
Bacteriology	Aerobes, anaerobes	Swab with medium, tissue (native)
Histopathology		Tissue (fixed)

Further Reading

Tallon R, McGovern K. Equine liver disease in the field. Part 1: approach. UK-Vet Equine. 2020;4(1):14-18. doi:10.12968/ukve.2020.4.1.14

Tallon R, McGovern K. Equine liver disease in the field. Part 2: causes and management. UK-Vet Equine. 2020;4(1):71-76. doi.org/10.12968/ ukve.2020.4.3.71

Ramsauer AS, Badenhorst M, Cavalleri JV. Equine parvovirus hepatitis. Equine Vet J. 2021 Sep;53(5):886-894. doi: 10.1111/evj.13477

Lymphomas

Lymphomas also occur regularly in guinea pigs. In addition to lymphoma, a form of leukaemia has been described, particularly affecting young adult animals (under 3 years of age). These animals usually die within a few weeks.

The 46 lymphomas included in this study were predominantly located in the lymph nodes (n = 31), but also in the skin (n = 11), internal organs (n = 3), and the eye (n = 1).

A special form of cutaneous lymphoma is the cutaneous epitheliotropic lymphoma (syn. mycosis fungoides). This is a progressive tumour disease, characterised by infiltration of tumour T lymphocytes (memory T cells) into the epidermis and adnexa (Moore & Olivry, 1994). The aetiology of this disease is not yet fully understood. This form of lymphoma also occurs in guinea pigs and is often not initially recognised as a tumour. Clinically, erythema, alopecia, and scaling may be observed, often leading to an initial suspicion of a skin disease. Nodular proliferations may only develop at later stages of the disease (Fig. 6).

 

Mammary Tumours

In contrast to other animal species, mammary tumours are also regularly observed in male guinea pigs (Table 1) (Schöniger et al., 2025). During the study period, 157 tumours were submitted, of which 75 originated from female animals and 58 from male animals; the sex of 24 animals was unknown. A total of 78 adenomas, 7 fibroadenomas, 71 adenocarcinomas, and 1 undifferentiated carcinoma were diagnosed.

Table 1: Distribution of mammary tumours by sex

Diagnosis	Number	F	FS	M	MS	U
Total	157	74	1	44	14	24
Benign	85	47	–	17	7	14
Malignant	72	27	1	27	7	10

Legend: F: female, FS: female spayed, M: male, MS: male castrated, U: sex unknown.

 

Uterine Tumours

In contrast to rabbits, guinea pigs exhibit a wide range of uterine changes, which can be non-neoplastic or proliferative. Simple benign or malignant tumours may occur, and mixed tumours can also be diagnosed (Laik-Schandelmaier et al., 2017).

Between 2013 and 2020, 60 tumours from the uterine region were submitted, of which 37 were benign and 23 malignant. The tumours were classified as 28 epithelial, 27 mesenchymal, and 5 mixed tumours. Among the epithelial neoplasms, 19 adenomas were observed compared with 9 adenocarcinomas. Among the mesenchymal tumours, 17 leiomyomas, 8 leiomyosarcomas, and 2 undifferentiated sarcomas were identified. Of the mixed tumours, only one was benign, whereas four were malignant.

 

Thyroid Tumours

In contrast to other small mammals, guinea pigs frequently develop thyroid tumours. During the study period, 15 adenomas and 14 carcinomas were diagnosed.

 

Conclusion

In the present study of spontaneous tumours in guinea pigs, benign neoplasms of the skin and subcutaneous tissue predominated. Lipomas were by far the most common tumours, followed by trichofolliculomas. Cutaneous epitheliotropic lymphomas can be clinically difficult to distinguish from inflammatory skin conditions.
Mammary tumours occurred in both female and male guinea pigs. In this study, castrated females and males were less frequently affected than intact animals.
Histopathology is an important tool for the prognostic assessment of tumours, as neoplasms of the mammary gland, uterus, and thyroid can be either benign or malignant.
This study demonstrates that a tumour diagnosis in guinea pigs does not necessarily constitute a death sentence. In many cases, the underlying lesion is benign, and timely veterinary intervention with removal of the mass allows the animal to continue a normal, healthy life.

 

Dr Claudia Schandelmaier

 

Services offered
– Histopathology
– Cytology

What Is the Quality of Drinking Water?

Legal Considerations

From a legal perspective, drinking water is classified as an animal feed (Regulation EC No. 178/2002) and is subject to the Feed Hygiene Regulation (Regulation EC No. 183/2005). Drinking water must be suitable for the species in question and the drinking systems must be freely accessible. They should be designed to minimise the risk of contamination. Regular cleaning and maintenance of the systems is mandatory.

There are only recommendations from the BMLEH (Federal Ministry for Agriculture, Food and Home Affairs) regarding the quality of drinking water in livestock production. Unlike the Drinking Water Ordinance, these recommendations are not legally binding.

 

The Importance of Biofilms in Drinking Troughs and Water Lines

The quality of drinking water can be affected by the materials used and the formation of biofilms. Daily emptying of drinking troughs is not sufficient to ensure cleanliness. The most effective way to remove a biofilm is through drying or exposure to sunlight. In any case, regular mechanical cleaning with a brush should be performed.

 

Laboratory Diagnostic Analysis of Drinking Water

When is drinking water considered to be of good quality, and what do the individual parameters indicate? The following discussion addresses these points:

Criteria for Suitability as Drinking Water (BMLEH):

• Palatability is a fundamental prerequisite for adequate water intake.
• Safety/Compatibility ensures that all constituents are present only at concentrations that are not harmful to the animal.
• Usability ensures that no adverse effects occur on the watering equipment.

Correct Sampling

For microbiological testing, water samples should be collected under sterile conditions. A distinction is made between samples taken directly from drinking troughs and samples from inlet taps.

For general assessment of drinking water quality, it is advisable to take samples from an inlet tap or the source supplying the drinking system. In cases where herd problems are associated with insufficient water intake, additional samples should be collected directly from the animals’ drinking troughs and evaluated for suitability. The installed pipework can play a decisive role in water quality.

• Before sampling, the collection point should be sterilised by flaming the Alternatively, the tap can be immersed in an alcohol solution for several minutes.
• The clearly labelled sample container (Fig. 1) should be sterile; a mineral water bottle may also be suitable. It should be rinsed several times with the water to be sampled before use.
• Allow water to run for 2–3 minutes before taking the sample.
• Avoiding Contamination: unscrew the lid immediately before filling and close it immediately afterwards. Do not touch the interior of the container, and wear disposable gloves.
• Transport: keep refrigerated, protected from light, and transport as quickly as possible.

Sampling from a hose should be avoided, as effective chemical or thermal disinfection is not possible. Biofilms often develop inside hoses.

If sampling from a hose is unavoidable, it must be flushed thoroughly for an extended period to remove stagnant water with very high bacterial counts (Fig. 2).

Sampling directly from the animals’ drinking troughs is only appropriate in cases of acute symptoms (e.g., diarrhoea) when testing for specific pathogens (e.g., Salmonella) (Fig. 3). Such troughs often show elevated colony counts and contamination with, for example, enterococci or E. coli/coliform bacteria.
This type of sampling corresponds to purpose C in the drinking water sector (“as it is consumed”) and does not provide information on the suitability of the water supplied to the troughs as drinking water.

When taking samples for the analysis of chemical parameters, a sterile sampling container is not strictly required. However, water should be allowed to run for 2–3 minutes before sampling. During collection, care must be taken to prevent dirt or rust from entering the sample. Unlike microbiological sampling, the container should be filled completely without air bubbles and sent to the laboratory as soon as possible, preferably chilled.

 

Guidance Framework for the Feed Law Assessment of Drinking Water (BMLEH)

Microbiological Parameters

Parameter	Orientation Value
Salmonella	0/100 ml
Campylobacter	0/100 ml
E. coli	0/100 ml (ideally absent)
Heterotrophic plate count at 20 °C	< 10.000 CFU/ml
Heterotrophic plate count at 37 °C	< 1000 CFU/ml

 

Recommended physico-chemical parameters

Parameter	Unit	Guideline value	Notes	Drinking Water Ordinance
pH value		> 5, < 9	may cause pipe corrosion and heavy metal release	6,5 – 9,5
Electrical conductivity	µS/cm	< 3000	Higher values may impair palatability and induce diarrhoea	2790
Total soluble salts	(g/l)	< 2,5	Refers primarily to NaCl content
Oxidisability	(mg/l)	< 15	Indicates the presence of oxidisable substances / biofilm burden	5

 

Recommended chemical quality parameters for drinking water

Parameter	Recommendation (mg/L)	Possible disturbances	Drinking Water Ordinance (mg/L)
Ammonium (NH4+)	< 3	Indicator of contamination	0,5
Arsenic (As)	< 0,05	Health disorders, reduced performance	0,01
Lead (Pb)	< 0,1		0,01
Cadmium (Cd)	< 0,02		0,005
Calcium (Ca)	500	Functional disorders, lime deposits in pipes and valves	No limit value
Chloride (CI-)	< 250 (poultry) < 500 (others)	Wet droppings	250
Iron (Fe)	< 3	Antagonist to other trace elements, deposits in pipes, biofilm formation, taste alteration	0,2
Fluoride (F)	< 1,5	Disturbances of teeth and bones	1,5
Potassium (K)	< 250 (poultry) < 500 (others)	Wet droppings	No limit value
Copper (Cu)	< 2	Total intake must be considered in sheep and calves	2
Manganese (Mn)	< 4	Precipitations in the distribution system, possible biofilms	0,05
Sodium (Na)	< 250 (poultry) < 500 (others)	Wet droppings	200
Nitrate (NO3-)	< 300 (cattle) < 200 (others)	Risk of methaemoglobin formation, total intake must be considered	50
Nitrite (NO2-)	< 30	Risk of methaemoglobin formation, total intake must be considered	0,5
Mercury (Hg)	< 0,003	General disturbances	0,001
Sulphate (SO42-)	< 500	Laxative effect	250
Zinc (Zn)	< 5	Mucosal alterations	No limit value

 

 

Conclusion

In addition to microbiological parameters, such as pathogenic E. coli or Salmonella, chemical parameters such as sulphate and nitrate in drinking water play a particularly important role in cattle health. Calves are generally more susceptible than adult animals. Sulphate concentrations of 500–600 mg/L in calves can negatively affect faecal consistency, while higher concentrations (> 2500 mg/L) may cause severe clinical symptoms resembling vitamin B1 deficiency, which can be explained by polioencephalomalacia (Kamphues et al., 2007).

These examples demonstrate that providing cattle with ad libitum access to suitable drinking water is crucial for maintaining health and performance. At Laboklin, we are happy to assist you in assessing the quality of your drinking water and to provide expert guidance on its interpretation.

Dr. Martin Felten, Dr. Anna-Linda Golob,

Swanhild Wagenfeld

1.   α1-Antitrypsin

α1-Antitrypsin (α1-AT) is a protein belonging to the serine protease inhibitor (serpin) family. It is primarily synthesised in the liver and circulates in the blood plasma at relatively stable concentrations. Its physiological function is to inhibit proteolytic enzymes — particularly neutrophil elastase — in order to prevent tissue damage caused by excessive inflammatory responses. α1-AT is of particular diagnostic relevance in cases of protein-losing enteropathy (PLE). Unlike many other proteins, α1-AT is largely resistant to enzymatic degradation within the gastrointestinal tract. When intestinal barrier function is compromised — e.g. due to inflammation or ulceration — α1-AT may leak from the plasma into the intestinal lumen and can be detected intact in faeces. This makes α1-AT an ideal faecal marker for identifying plasma protein loss via the gastrointestinal tract. Detection in faeces indicates impaired intestinal barrier integrity and is considered a marker of intestinal protein loss.

 

Indication
In dogs, faecal α1-antitrypsin has been particularly well studied as an early marker for developing protein-losing enteropathy (PLE) in predisposed breeds (e.g. Soft Coated Wheaten Terrier). Studies indicate that the marker can detect intestinal barrier dysfunction at an early stage – before clinical signs or detectable hypoalbuminaemia in blood are present. Another potential application is in the evaluation of hypoproteinaemia or hypoalbuminaemia. Since PLE does not always present with diarrhoea, faecal α1-AT can be a useful tool to help identify the cause of such abnormalities in bloodwork. In patients with chronic enteropathy, increased faecal α1-AT concentrations are indicative of more severe intestinal disease. Additionally, the marker can be used for therapy monitoring: decreasing values during treatment suggest an improvement in intestinal barrier function.

 

Points to Consider
Measurement is typically performed via ELISA. Even small amounts of α1-antitrypsin escaping through a damaged mucosa can be detected. However, both diurnal variation and uneven distribution within a single faecal sample are known issues. This may result in low or undetectable concentrations being reported despite the presence of disease. Testing three consecutive faecal samples increases diagnostic accuracy.

Interpretation of faecal α1-AT concentrations should always be made in the clinical context. The reference interval is broad, and overlap with healthy control animals is possible. Diurnal fluctuations and uneven distribution in the faeces can lead to both false-positive and false-negative results. It is also important to consider that concentrations may be elevated in the presence of gastrointestinal bleeding or increased mucus production in the intestines.

α1-antitrypsin does not provide information on the aetiology of a PLE – it is a quantitative marker of protein loss, not specific for inflammation or neoplasia.

 

α1-Antitrypsin in Cats

Data are also available for cats, indicating that α1-antitrypsin concentrations can be significantly elevated in cases of chronic enteropathy. As with dogs, interpretation should always be made in the clinical context. Overall, chronic enteropathy in cats appears to be more frequently associated with intestinal protein loss than in dogs. As a result, defining a protein-losing enteropathy (PLE) in this species is more challenging.

 

2.    Calprotectin

Calprotectin is a calcium-binding protein from the S100 protein family, primarily found in neutrophilic granulocytes. It is released in increased amounts during inflammatory processes. In cases of gastrointestinal inflammation, calprotectin is secreted across the intestinal mucosa into the lumen and can therefore be detected in faeces.
Its measurement in faecal samples allows for a non- invasive assessment of inflammatory activity in the gastrointestinal tract – a method that is well established in human medicine and increasingly applied in veterinary medicine.

 

Indication
In dogs, faecal calprotectin is used as a marker for chronic inflammatory bowel disease. Studies have shown that elevated concentrations correlate with both the severity of histological changes and the clinical activity index (CCECAI). It can be helpful for differentiating between inflammatory and non-inflammatory diarrhoea, assessing disease activity, and monitoring treatment response.

The significance of calprotectin lies particularly in its ability to reflect the severity of chronic enteropathy. Studies have demonstrated that dogs with a higher CCECAI (e.g. ≥12) exhibit significantly increased values. The parameter can provide insights into the need for immunosuppressive therapy and support prognostic evaluations. The higher the faecal calprotectin concentration, the more likely it is that the patient will require immunosuppressive treatment. In follow-up assessments, a failure of levels to normalise may indicate incomplete remission. A renewed increase can occur even before clinical deterioration becomes apparent.

 

Points to Consider
As with all biomarkers, calprotectin is not disease- specific. It merely indicates the presence and extent of inflammation. Elevated levels may also be observed in bacterial infections, parasitic infestations, or neoplastic diseases. Interpretation should therefore always occur within the clinical context and be supported by further diagnostic investigations (e.g. imaging, endoscopy, histology). Faecal calprotectin concentrations within the reference range do not exclude the presence of enteropathy. This is particularly relevant in dogs, where the predominant inflammatory response tends to be lymphoplasmacytic rather than neutrophilic. Additionally, many enteropathies in dogs appear to be food-responsive, and in such cases, a strong neutrophilic inflammatory response – and therefore elevated calprotectin levels – is not expected.

 

Calprotectin in Cats
Calprotectin has also shown promising results in cats. It is used as a supportive marker in cases of chronic diarrhoea. Particularly relevant are investigations into its potential for differentiating between inflammatory and neoplastic processes (e.g. low-grade lymphoma). Faecal calprotectin concentrations may be significantly higher in cases of lymphoma; however, a reliable differentiation has not yet been clearly demonstrated.

 

3.    Zonulin

Zonulin is an endogenous protein that plays a central role in regulating intestinal permeability. It controls the permeability of the tight junctions – those cellular connections that link intestinal epithelial cells and thus prevent the intrusion of unwanted substances. Increased zonulin release leads to the loosening of these junctions, resulting in increased intestinal permeability – a condition commonly referred to as “leaky gut”.
Under physiological conditions, zonulin allows a temporary opening of the intestinal barrier, e.g. for immune surveillance or transport processes. This regulation is finely tuned and normally reversible. Pathological conditions arise when this opening is prolonged or excessive, permitting the passage of bacterial components, toxins, or incompletely digested dietary proteins into the tissue.

 

Indication
Zonulin has been identified as a potential marker for the integrity of the intestinal barrier. Elevated faecal or serological zonulin levels have been described particularly in dogs with chronic enteropathy, food allergies, or inflammatory bowel disease. Studies indicate that dysregulated zonulin expression is associated with increased intestinal permeability, i.e. a disturbed barrier mechanism. The determination of zonulin therefore offers a non-invasive insight into the functionality of the intestinal mucosa. Despite the currently limited amount of clinical data available for small animals, the measurement of zonulin opens up new diagnostic and therapeutic perspectives for a holistic approach to gastrointestinal diseases.

 

What should be considered
Zonulin is not a disease-specific marker.
As with many newer biomarkers, results must be interpreted in the overall clinical context.
Elevated values alone do not allow a diagnosis but can provide indications of functional disturbances of the barrier. Factors such as stress, medication, age, or diet may influence its concentration.

 

Zonulin in cats
Initial pilot studies in cats suggest that the protein may also play a role in feline chronic enteropathies (e.g. lymphoplasmacytic enteritis or low-grade lymphomas).

 

4.    Faecal secretory immunoglobulin A (sIgA)

Immunoglobulin A (IgA) is a central component of mucosal immune defence. It is produced as secretory IgA (sIgA) by plasma cells in the lamina propria of the intestine and actively transported across the epithelium into the intestinal lumen. There, it forms a first line of defence against pathogenic microorganisms without triggering inflammatory responses. sIgA thus acts as a ‘bouncer’, protecting the mucosa from bacterial adherence, toxin activity, and invasion. Secretory IgA binds to surface antigens of bacteria, viruses, or toxins and neutralises them before they come into contact with epithelial cells. Unlike IgG or IgM, IgA does not activate the complement system and is therefore non-inflammatory. This is essential for immunological tolerance in the intestine, where numerous harmless antigens (e.g. food components, commensals) are constantly present.

 

Indication
A reduced concentration in faeces may indicate a functional immunodeficiency of the intestinal mucosa. Reduced sIgA production has been particularly described in German Shepherds with chronic enteropathy.

 

What should be considered
sIgA is regarded as an indicator of mucosal immunocompetence in the intestine, although interpretation must be approached with caution, as it can be influenced by age, stress, diet, and even sample handling. Measurement should be performed on the freshest faecal samples possible.

 

sIgA in cats
Data on faecal sIgA in cats are still limited. However, initial studies suggest that reduced sIgA concentrations in faeces can occur in chronic intestinal diseases, such as inflammatory enteritis or intestinal lymphoma.

 

5.    Canine pancreatic elastase 1

Elastase is a proteolytic enzyme produced in the exocrine pancreas and released into the small intestine with pancreatic juice. Its main function is to break down elastin, a structural protein in connective tissue. In diagnostics, however, its enzymatic activity is of less importance than the detection of stable amounts of elastase in faeces, which allows conclusions to be drawn about exocrine pancreatic function. In contrast to many other pancreatic enzymes, elastase in the intestinal lumen is largely resistant to enzymatic degradation, bile acids, and bacterial influences. It is excreted unchanged in faeces and can be detected using immunological tests (ELISA), making it a non-invasive marker of exocrine pancreatic function. In dogs, measurement of pancreatic elastase 1 is a useful screening tool, particularly in patients with non-specific gastrointestinal signs. It can serve as a supplement or preliminary test to determine specific trypsin-like immunoreactivity (cTLI). Normal elastase values generally rule out clinically relevant exocrine pancreatic insufficiency (EPI). Significantly reduced values may indicate exocrine dysfunction, but confirmation with serum cTLI is required.
Low faecal elastase concentrations can occur, for example, due to the dilution effect in diarrhoea and are therefore not conclusive for EPI on their own.
Low concentrations can also be observed in healthy dogs. A cTLI concentration within the reference range argues against the presence of EPI, even if faecal elastase is very low. Only in very rare cases is EPI accompanied by low faecal elastase but normal or low-normal cTLI concentrations.
This can occur in situations such as occlusion of the pancreatic duct (the pancreas itself remains functional, but the enzymes do not reach the intestine), or when blood sampling for cTLI is not performed under fasting conditions or occurs during an episode of pancreatitis (residual enzyme still present is released into the blood).

 

6.    (Total) bile acids

Bile acids are synthesised in the liver and secreted into the small intestine during digestion.
Approximately 95% are reabsorbed in the ileum. Disruption of this reabsorption – for example, due to chronic inflammation – leads to increased amounts of primary bile acids in the colon, which can trigger secretory diarrhoea. Similarly, dysbiosis with reduced conversion into secondary bile acids by Clostridium hiranonis (renamed Peptacetobacter hiranonis) can affect these processes.

 

7.   Dysbiosis investigations

The intestinal microbiota plays a key role in the pathogenesis of chronic enteropathies. Dysbiosis tests quantify relevant bacterial marker species (e.g. Faecalibacterium, Turicibacter, Clostridium hiranonis) using PCR-based methods and assess their deviation from the physiological state. An altered score indicates disrupted microbial homeostasis and may have prognostic relevance.

 

Conclusion

Over recent years, the evaluation of faecal biomarkers has become increasingly integrated into the diagnosis and therapeutic monitoring of chronic enteropathies. While α1-antitrypsin and calprotectin can provide direct indications of protein loss and inflammation,other may offer additional information on the functional and microbial integrity of the gut. Their targeted use can provide a basis for therapeutic decisions, follow-up assessments, and prognosis. It remains essential to interpret results in the clinical context and, where appropriate, to combine several parameters to enhance diagnostic value.

 

Dr. Jennifer von Luckner

 

Our services for enteropathies

• Intestinal profile (serum)
• Diarrhoea profiles (faeces)
• Parasite profiles (faeces)
• Dysbiosis analysis (faeces)
• Dysbiosis profile (faeces)
  …and many more.