How to Draw Blood on a Leopard Gecko

The use of venepuncture in reptiles is an important diagnostic step in determining illness and disease. There are several venepuncture sites that vary between species and knowledge of these will ensure a diagnostic sample. Knowing how to correctly handle the reptile will prevent any unnecessary injury to the handler or the animal. The correct gauge needle and smallest syringe need to be used so as not to damage the red blood cells, and the site needs to be cleaned appropriately. This article will look at the venepuncture sites and techniques used in the Chelonian and Squamata species.

There are approximately 9000 known species of reptile (Rendle and Crack-nell, 2012). These are subdivided into four categories:

• Chelonia: turtles, tortoises and terrapins

• Squamata:

  • ◦ Sauria — lizards

  • ◦ Serpentes — snakes

• Crocodilia: alligators and crocodiles

• Rhynchocephalia: tuatara.

Reptiles can be difficult to clinically assess and a blood sample can be an important diagnostic step. A blood profile may include:

• Renal parameters: high levels of uric acid may indicate renal disease

• Glucose levels: may indicate hypoglycaemia in a malnourished animal. Hyperglycaemia in bearded dragons can be indicative of neoplasia

• Calcium and phosphorous levels: can indicate if a female is gravid or if the animal is suffering from nutritional secondary hyperparathyroidism (NSHP)

• Haematology: high or low white-cell count can indicate a bacterial or viral disease

• Hydration levels: high urea may indicate dehydration (e.g. in tortoises)

• Total protein levels: indicate liver function and nutritional stasis.

This article will focus on blood-sampling techniques in Chelonia and Squamata.

Reptiles have a different cardiovascular system to that of mammals as their heart has three chambers, two atria and one ventricle. It functions predominantly in the same way as a four-chambered heart. The ventricle has three sub-chambers and receives blood from the right and left atrium (Aspinall and Cappello, 2009). Reptiles have a renal portal system which directs blood from the tail and hind limbs to the kidneys. Therefore, drugs that have been injected into the caudal aspect of the body may be directed and excreted through the kidneys before reaching the systemic circulation (Aspinall and Cappello, 2009). However, recent evidence suggests that this may not be of clinical importance and therefore needs further evaluation.

Blood collection

There are several aspects to consider when taking a blood sample from a reptile:

• A safe sample is 10% of total blood volume in the healthy reptile. Reptile blood volume equates to around 5–8% of body-weight; therefore, 0.5–0.8% of bodyweight (g) can be taken (Heard et al, 2004)

• The venepuncture site should be thoroughly cleaned. An appropriate disinfectant is povidone-iodine

• Gloves should be worn and hands must be washed after handling reptiles as they are naturally predisposed to Salmonella, an important reptile zoonosis (Raftery, 2004)

• Some reptiles may be severely dehydrated, restricting the ability to take a blood sample. Hospitalising is recommended for bathing and fluid therapy, which supports the cardiovascular system and increases the chance of taking a successful sample

• Care should be taken with the handling of reptiles as increased pressure may cause severe contusions. This is potentially fatal in reptiles as a result of the toxicity from the breakdown products of the muscle (Maclean and Raiti, 2004)

• Lithium heparinised tubes are preferred for haematology. Ethylenediaminetetraacetic acid (EDTA) samples have been shown to cause haemolysis and are therefore not advised (Wilkinson, 2004).

Chelonians

Sites which are commonly used for venepuncture in Chelonia are:

• Jugular vein

• Supravertebral (subcarapacial) vein

• Dorsal coccygeal vein.

Other sites include:

• Occipital sinus

• Brachial vein

• Femoral vein

• Heart (see Table 1).

Table 1.

Blood profile reference range — Chelonia: Spur Thigh Tortoise (Testudo graeca). References provided by PALs Laboratory.

Biochemistry
Albumin	g/litre	13.0–26.0
Total protein	g/litre	23.0–44.0
Uric acid	Umol/litre	124–446
AST*	Iu/litre	0.00–210
Creatine kinase	Iu/litre	0.00–2620
Urea	Mmol/litre	0.00–2.10
Calcium	Mmol/litre	1.65–3.30
Phosphorous	Mmol/litre	0.48–1.81
Haematology
White cell count	10^9/litre	1.0–4.5
Haemoglobin	g/dl	6.4–15.7
PCV**	%	17–42
Heterophils	10^9/litre	0.50–5.00
Lymphocytes	10^9/litre	0.27-2.50
Monocytes	10^9/litre	0.00–0.20

*

AST: aspartate aminotransferase

**

PCV: packed cell volume

Jugular vein

The jugular vein (Figure 1) lies laterally and superficially on the neck. Jugular venepuncture should be attempted first as this sample is least likely to be diluted with lymph (Barrows et al, 2004). A 5/8-inch 23-25 gauge needle and the smallest syringe possible should be used.

Figure 1. A Hermann's tortoise (Testudo hermanni) jugular vein blood sample.

Restraint of the Chelonian may require sedation or anaesthesia. In the ill or easily restrained patient, this may be possible with the help of an assistant. The animal is held in lateral recumbency with the neck extended to visualise the vein. To extend the head, gentle pressure is applied behind the mandibles. The forelimbs should be restrained by retracting them caudally, taking care not to exert excessive force as this could result in iatrogenic fractures. The assistant should place pressure on the thoracic inlet to facilitate a better visualisation; in the smaller Chelonian, this can be achieved by using a cotton bud so as not to restrict the view of the vein (Heard et al, 2004).

Supravertebral (subcarapacial) vein

The supravertebral vein (Figure 2) lies midline on the ventral aspect of the carapace, dorsal to the cervical vertebrae. A 1-inch 23-25 gauge needle and the smallest syringe possible should be used.

Figure 2. A Hermann's tortoise (Testudo hermanni) supravertebral vein blood sample.

The needle is inserted where the skin meets the shell and advanced caudally at a 60° dorsal angle. Negative pressure is applied gently until blood has been aspirated. If the carapace is reached, the needle should be withdrawn slightly, then redirected and advanced while still applying negative pressure (Heard et al, 2004). Lymphatic vessels in this region increase the risk of lymph dilution.

Dorsal coccygeal vein

The dorsal coccygeal vein (Figure 3) is found on the midline aspect of the tail, dorsal to the vertebrae and is easier to locate in the larger Chelonian. A 5/8-inch 21-25 gauge needle and the smallest syringe possible should be used.

Figure 3. A Red-footed tortoise (Chelonoidis carbonaria) dorsal coccygeal vein blood sample.

The needle is inserted slowly at a 45° angle into the midline at the junction where the tail meets the carapace (Barrows et al, 2004). It is advanced until the vertebrae have been reached and then slowly withdrawn while applying gentle negative pressure until blood has been aspirated. This area must be cleaned thoroughly to prevent faecal contamination (Heard et al, 2004). Lymphatic vessels in this region increase the risk of lymph dilution.

Squamata — Sauria

Venepuncture sites which are commonly used in Sauria are:

• Ventral coccygeal vein

• Heart (cardiocentesis)

• Ventral abdominal vein (see Table 2).

Table 2

Blood profile reference range — Sauria: Bearded Dragon (Pogona vitticeps). References provided by PALs Laboratory.

Biochemistry
Albumin	g/litre	25–35
Total protein	g/litre	52–72
Uric acid	Umol/litre	0–350
AST	Iu/litre	0–80
Creatine Kinase	Iu/litre	0–550
Urea	Mmol/litre	0.0–1.4
Sodium	Mmol/litre	140–160
Calcium	Mmol/litre	2.2–3.4
Phosphorous	Mmol/litre	1.1–2.3
Haematology
White cell count	10^9/litre	1.5–8.5
Haemoglobin	g/dl	7.7–12.0
PCV	%	17–35
Heterophils	10^9/litre	1.6–5.0
Lymphocytes	10^9/litre	0.2–4.0
Monocytes	10^9/litre	0.042–2.204

Poor handling of certain Sauria species can result in autotomy, a defence mechanism whereby the tail is shed at natural fracture planes (Maclean and Raiti, 2004) (Figure 4). The tail may take many months to regrow and will never have the same scale or colour formation as the original (Raftery, 2004). In some cases, the tail may never regrow; therefore, gentle handling with little pressure placed on the tail is advised.

Figure 4. A leopard gecko (Eublepharis macularius) exhibiting autotomy.

Ventral coccygeal vein

The ventral coccygeal vein (Figure 5) lies on the midline ventral aspect of the vertebrae and is the preferred and most effective site for blood sampling.

Figure 5. A ventral coccygeal vein blood sample in the bearded dragon (Pogona vitticeps).

In small lizards, a 5/8-inch 23-35 gauge needle and the smallest syringe possible should be used. In medium-to-large lizards, a 1-inch 25-gauge needle should be used (Heard et al, 2004). The site should be cleaned thoroughly before venepuncture takes place.

The lizard should be restrained in dorsal recumbency and the tail supported by an assistant. A towel used to restrain the lizard is advised to protect the handler from injury. The needle is introduced into the proximal third of the midline tail and advanced perpendicular to the tail towards the vertebrae until they come into contact. The needle is retracted slowly while applying negative pressure until blood has been drawn. It must be taken slowly owing to the low volume of blood within this vein (Jenkins, 1996). Larger lizards can be placed in sternal recumbency with the tail placed over the edge of the table and the sample taken from below.

Heart (cardiocentesis)

There is a great level of risk involved with obtaining a blood sample via cardiac puncture. A 23-25 gauge needle is needed and the smallest syringe possible should be used.

The cardiac position in different species can vary. In some, for example, the heart is cranial between the shoulders; in others, the heart is positioned mid-abdomen. It is good practice to use a Doppler flow-detection monitor to locate the exact position of the heart. The cranial heart is approached from the thoracic inlet or from between the ribs. Slight negative pressure should be applied as the needle is directed caudally towards the heart, parallel to the base of the sternum. Often this sample is taken from lizards that are too small to have a sample obtained from the ventral coccygeal vein and are therefore at great risk of cardiac damage. Careful restraint of the lizard is essential and an anaesthetic may be advised for this sample. Sudden movements from the lizard may result in laceration of the cardiac tissue which can result in severe haemorrhage and even death (Jenkins, 1996).

Ventral abdominal vein

The ventral abdominal vein (Figure 6) lies superficially on the ventral midline. A 5/8-inch 25-gauge needle and the smallest syringe possible should be used.

Figure 6. A crested gecko (Rhacodactylus ciliatus) ventral abdominal vein blood sample.

Lacerations of this vein prove a high risk and only a well restrained or anaesthetised patient should have a sample taken from here. The needle should be inserted in the proximal two thirds of the abdominal midline superficially under the skin. This site is not commonly used except in cases where the tail is absent. Particular care should be taken when using this technique.

Squamata — Serpentes

Venepuncture sites commonly used in the Serpente species are:

• Heart (cardiocentesis)

• Ventral coccygeal vein

• Palatine vein (see Table 3).

Table 3

Blood profile reference range — Serpentes: Corn Snake (Pantherophis guttatus). References provided by PALs Laboratory

Biochemistry
Albumin	g/litre	18–34
Total protein	g/litre	46–84
Uric acid	Umol/litre	77–500
AST	Iu/litre	9–224
Creatine kinase	Iu/litre	77–2460
Urea	Mmol/litre	0–2.14
Sodium	Mmol/litre	147–183
Calcium	Mmol/litre	3.00–4.90
Phosphorous	Mmol/litre	0.61–1.94
Haematology
White cell count	10^9/litre	1.02–12.0
Haemo-globin	g/dl	9.7–12.0
PCV	%	21–52
MCHV	g/dl	32–40
Heterophils	10^9/litre	0.135–8.35
Lymphocytes	10^9/litre	0.406–22.9
Monocytes	10^9/litre	0.042–2.204
Azurophils	10^9/litre	0.075–5.338

Heart (cardiocentesis)

The heart (Figure 7) lies roughly in the proximal one third of the body; however, this varies depending on the species. A 23-25 gauge needle and the smallest syringe possible should be used.

Figure 7. Cardiocentesis in a Madagascan tree boa (Sanzinia madagas-cariensis).

The snake is restrained in dorsal recumbency with gentle pressure placed behind the head, taking care to avoid iatrogenic trauma caused by excessive pressure. The body must be supported and in the case of larger snakes, this may require more than one assistant.

The heart is located by manual palpation, using a Doppler flow-detection monitor or simply by observing the scutes for signs of the heart beating. Once located, the heart should be stabilised with a thumb and finger placed cranially and/or caudally around it (Jenkins, 1996). The needle should be inserted at a 45° angle between the ventral scales and advanced cranially towards the heart. The blood flow will correspond to the movement of the heart. Care should be taken with this sample as there is a high risk of cardiac damage; an anaesthetic may be advised. Sudden movements from the snake may result in laceration of the cardiac tissue which can result in severe haemorrhage and even death.

Ventral coccygeal vein

The ventral coccygeal vein (Figure 8) lies along the ventral midline of the coccygeal vertebrae. A 5/8-inch or 1-inch 23-25 gauge needle (depending on the size of the snake) and the smallest syringe possible should be used. The site should be cleaned appropriately to reduce the risk of faecal contamination.

Figure 8. A Madagascan tree boa (Sanzinia madagascariensis) ventral coccygeal vein blood sample.

The snake should be held by an assistant in dorsal recumbency, taking care to stabilise the caudal tail. The needle is introduced into the midline tail, caudal to the vent; it is advanced towards the vertebrae perpendicular to the tail until they come into contact. The needle is retracted slowly while applying negative pressure until blood has been drawn.

Palatine vein

The palatine vein lies on the medial surface of the palatine. The risk of injury from snake bite is great. A 25–27 gauge needle and the smallest syringe possible should be used.

This technique should be used mainly in larger snakes and it is advisable to only obtain this sample under anaesthetic. It is important to prevent contamination from oral fluids. This site is not often used for venepuncture.

Conclusion

Illness and disease are commonly seen in reptiles resulting in the need for further diagnostic steps including venepuncture. There are several venepuncture sites that differ between the Chelonia, Sauria and Serpentes species, and knowledge of these is vital to obtain a diagnostic sample.

Thorough cleaning of the site is important before obtaining a sample and selecting the correct gauge needle and smallest syringe possible is important so as not to lacerate the vein or damage the red blood cells. Care must be taken with the handling of the reptile as autotomy can occur; severe contusions could also result in fatalities. Knowledge of the species and how to correctly handle the reptile is important to obtain a sample of diagnostic quality and to prevent any injury to both the handler and the animal.

Key Points

• There are several venepuncture sites and these vary across reptile species.

• Safe and careful handling is important as some Sauria species can exhibit autotomy; contusions can also be fatal to reptiles.

• Reptiles have a different cardiovascular system to that of mammals.

• Some techniques require an anaesthetic to prevent laceration and damage to the vein.

• Heparinised tubes should be used for the collection of blood in the majority of cases.

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How to Draw Blood on a Leopard Gecko

Source: https://www.theveterinarynurse.com/review/article/how-to-obtain-a-blood-sample-in-reptiles-via-venepuncture