The haemostatic system consists of 4 areas that may be impaired alone or in combination:
- vessels,
- platelets,
- plasmatic coagulation,
- fibrinolysis.
These 4 areas vary with regards to their evaluability using laboratory methods (see Table 5.1 below).
Vessels
Vessel wall damage is the most difficult to assess. However, it plays an important role in clinical toxicology because many venoms contain haemorrhagins (= venom components that damage vessel walls). While bleeding time does reflect endothelial damage, it is also influenced by platelet count and function. Many haemorrhagin-containing venoms also have components that either directly or indirectly damage, destroy or consume platelets. Moreover, bleeding time cannot be standardised.
The difficulty of assessing vessel condition (in clinical toxicology this means haemorrhagin activity) with laboratory methods is a significant reason for the uncertainty in assessing the bleeding risk. Even without combined impairment of plasmatic coagulation and platelets the integrity of the endothelium of small vessels can be compromised by haemorrhagins to such an extent that overt bleeding may occur.
The risk of haemorrhagin-induced bleeding must be assessed primarily on the basis of clinical experience with the venomous or poisonous animal that caused the accident (see Biomedical database).
However, vessel integrity is not only at risk from haemorrhagins following an accident with a venomous or poisonous animal. Secondary effects due to venom activity that results in the release of fibrin(ogen) split products and complement factors (C3a, C5a) can also cause vessel damage.
Platelets
The contribution of platelets to haemostasis is much more readily ascertainable using laboratory methods than that of the vessels.
Because of the possibility of combined haemorrhagic and platelet-damaging effects of venoms, bleeding time is also not conclusive for platelet evaluation. Only if the vessels, in particular the small vessels, are intact does bleeding time provide an overview of whether the platelet count and function are sufficient for haemostasis. Standardisation has its pitfalls, and the linear correlation between bleeding time and platelet count is only valid with certain limitations. In haemostatic defects with increased peripheral consumption, i.e. increased platelet turnover, such as occur in clinical toxicology, there is a predominance of young platelets, which are haemostatically more efficient than old platelets. The relationship between the platelet count and bleeding time is not linear in these cases.
Although platelet counting represents a quantitative method of platelet evaluation, it is not sufficient by itself to assess the contribution of platelets to haemostasis. Based on experience, a platelet count of 20,000–50,000 platelets/µl is sufficient to prevent spontaneous bleeding in the presence of intact vessels and efficient plasmatic coagulation, but this is only valid if platelet function is unimpaired.
However, specific evaluation of platelet function requires more complex laboratory methods that can only be carried out in special laboratories and are not inexpensive.
Plasmatic coagulation and fibrinolysis
Plasmatic coagulation and fibrinolysis can be assessed in great detail using laboratory methods. However, difficulties are posed by the choice of tests that are suitable for investigating a particular problem, as well as their interpretation, particularly in the case of complex haemostatic defects.
As with all laboratory investigations, sensitivity and specificity need to be taken into account when interpreting the results. Table 5.3 (see below) shows the sensitivity ranges for commonly used tests. The sensitivities of the two global tests prothrombin time (PT) and activated partial thromboplastin time (aPTT) clearly demonstrate how advanced clotting factor consumption may already be before this is reflected in abnormal test results.
Determination of the combination of tests used to assess haemostasis is influenced by whether the venomous or poisonous animal that caused the accident could be identified. If the animal can be identified, the sections titled "Laboratory and physical investigations" in the Biomedical database entries provide decision aids, provided that there is well-documented clinical experience with the relevant animal. Otherwise, haemostasis screening that is certain to identify those patients at risk for bleeding is used (Fig. 5.7, see below).
Fig. 5.7 Haemostasis screening: basic programme and interpretation of the test results.
n = normal; ↑ = increased; ↓ = decreased.
PT prothrombin time,
aPTT activated partial thromboplastin time,
TT thrombin time,
Fbg fibrinogen,
FSPtot fibrinogen split products (total),
D-Dimers fibrin-specific D split products that originate from the degradation of factor XIIIa-cross-linked fibrin chains,
FV, FVII, FVIII, FIX, FX, FXI, FXII
clotting factors,
CoF clotting co-factors,
PK prekallikrein,
HMWK high-molecular-weight kininogen.
The dynamics of haemostasis
It is essential to be aware that the results of laboratory investigations only ever provide a static picture at a specific time point in what is a dynamic process. The production of clotting factors and platelets on the one hand, and their inactivation and consumption on the other, determine the haemostatic potential demonstrated by the laboratory investigations for a defined point in time. The way in which the venom was applied, usually subcutaneously, as well as the transport of haemotoxins, usually lymphatic, can also cause delays in the onset of haemostatic defects. The intervals at which tests are repeated and the length of time for which a patient is observed must be precisely defined in each individual case in order not to prematurely exclude a haemostatic defect and to record trends in changes.
Laboratory methods
The laboratory programme that can be used to detect haemostatic defects depends on the aim and the available resources.
Under the most basic conditions only clotting time and possibly also a test kit for determination of fibrinogen split products are available to support the clinical assessment. Desirable would be a basic programme such as that suggested below ("Basic programme"). For research purposes, a more extended programme would be required, in order to assess haemostatic defects in detail and to clarify pathophysiological relationships. This requires specialised laboratories and is expensive.
'Bedside' tests
If a patient is being treated under very basic conditions, usually only the clotting time (CT) or the whole-blood clotting test are available for evaluation of blood coagulation (Fig. 5.8 and 5.9 below). A test kit for the determination of fibrinogen split products may possibly also be available. This test increases the sensitivity of the detection of coagulation disorders associated with the formation of such split products compared to the clotting time alone. Both clotting time and the whole-blood clotting test are suitable for detecting systemic activity of the venom and for establishing an indication for antivenom administration in cases of envenoming caused by coagulation-promoting venoms (Reid et al. 1963b, Warrell et al. 1977, Than-Than et al. 1987, 1988, Kamiguti et al. 1991).
Fig. 5.8 Clotting time.
Fig. 5.9 Whole-blood clotting test.
In the simplified whole-blood clotting test proposed by Warrell et al., several millilitres of venous blood are placed in a clean, dry test tube and left for 20 min. The test tube is then tilted in order to determine if a clot has formed (Warrell et al. 1977).
Although the low sensitivity of clotting time presents a problem for the early determination of a plasmatic haemostatic defect, an abnormal result on this test does clearly indicate that a major defect is already present.
It is important to be aware that although incoagulability of the blood on the whole-blood clotting test does generally indicate a marked fibrinogen deficiency, this result can also be caused by strong anti-platelet or anti-coagulative activity of the venom (Fig. 5.9 above) (Sano-Martins et al. 1994).
The reliability of the whole-blood clotting test as an indicator of decreased plasma fibrinogen concentrations was investigated in patients bitten by Bothrops species, in most cases by Bothrops jararaca. Blood samples that did not coagulate within 20 min generally had decreased fibrinogen concentrations. There was found to be a close relationship between the results of the whole-blood clotting test and the corresponding plasma fibrinogen levels. This applied to the initial investigations at the time of hospitalisation as well as investigations performed after antivenom administration. Proof of clotting ability 6–12 h after antivenom administration in patients who had incoagulable blood at the time of hospitalisation indicated the neutralisation of circulating venom components (Sano-Martins et al. 1994). These results are confirmed by the low serum venom concentrations measured at these time points using the ELISA method (Theakston et al. 1992).
In cases of envenoming accompanied by defibrinogenation, the whole-blood clotting test is superior to fibrinogen determination, as the former test is simpler, faster and more reliable. Moreover, it can also be used to assess the efficacy of antivenom. The advantages of this test in those rural regions of the world in which most such accidents occur and in which no laboratory facilities are available are obvious (Sano-Martins et al. 1994).
Basic programme
The choice of laboratory tests depends on the conditions under which the accident occurred and the purpose of the investigations.
If it is known which venomous or poisonous animal caused the accident, the Biomedical database entries provide information on the type of haemostatic defect that can be expected as well as the relevant tests for the purposes of diagnosis and follow-up.
If it is not known which animal caused the accident, the following haemostasis screening should be carried out (see Fig. 5.7 above):
- prothrombin time (PT) or activated partial thromboplastin time (aPTT),
- fibrinogen or thrombin time (TT),
- fibrinogen split products (FSPtot) and, if these are increased, also D-dimers,
- platelets.
Table 5.2 (see below) gives an overview of the most important venom-induced haemostatic defects and the possibilities of narrowing down the diagnosis with the aid of the basic programme.
If the results of the haemostasis screening are within the normal ranges of the tests, then either there was no coagulation disorder present at the time of blood collection or the disorder is of an extent that is below the sensitivity of the utilised test (see Table 5.3 below).
It would be problematic to indicate a time period beyond which a haemostatic defect can be definitively excluded. There have been reports of patients in whom haemostatic defects became evident on laboratory tests after a delay of up to 3 days.
In the first hours after the bite, haemostasis screening should be carried out hourly. If the results indicate a haemostatic defect, then depending on the situation the indication for antivenom administration is either established immediately or made dependent on the further course of the haemostatic defect. Haemostasis is also monitored after antivenom administration in order to determine whether the treatment is successful and to decide if further antivenom doses are necessary. An important cause of recurrence of haemostatic defects is continued mobilisation of venom from a depot in the region of the bite.
Extended basic programme
As mentioned above, the extended basic programme serves primarily to evaluate haemostatic venom activity in detail. Among other parameters, clotting factors, plasminogen and inhibitors are determined.
Table 5.1 Impairments within the 4 areas of the haemostatic system and their effect on important haemostatic tests and determination of individual factors
| Haemostatic test | Impairment of |
|||
| vessels | platelets |
plasmatic coagulation |
fibrinolysis | |
| Bleeding time (BT) |
+ | ++ | ||
| Clotting time (CT) |
++ |
+ | ||
| Clot observation test (COT) | ++ | ++ | ++ | |
| Prothrombin time (PT) |
++ | + | ||
| Activated partial thromboplastin time (aPTT) |
++ | + | ||
| Thrombin time (TT) |
++ | + | ||
| Platelet count |
++ | + | ||
| Platelet function |
++ | |||
| Fibrinogen | ++ | + | ||
| Factor II | ++ | |||
| Factor V |
++ |
+ | ||
| Factor VII |
++ | |||
| Factor VIII | ++ | + | ||
| Factor X | ++ | |||
| Antithrombin III | ++ | |||
| Protein C | ++ | |||
| α₂-Antiplasmin | ++ | |||
| Plasmin | ++ | |||
| Fibrinogen split products (FSP) | ++ | |||
++ has a primary effect on the test
+ also has an effect on the test
Table 5.2 Laboratory findings ("bedside" tests, basic programme) and classification of the most important venom-induced haemostatic defects
| Haemostatic tests | Clotting time (CT) ↑ | |||||||
| Prothrombin time (PT) ↑ | ||||||||
| Activated partial thromboplastin time (aPTT) ↑ | ||||||||
| Fibrinogen split products (FSP total) ↑ | ||||||||
| D-dimers ↑ | ||||||||
| Platelets ↓ |
||||||||
| Haemostatic activity of the venom |
Intrinsic and/or extrinsic prothrombin activation
-------------------------- |
fibrinogen-coagulating activity via "thrombin-like" proteinases |
primary fibrin(ogen)olytic activity: fibrin(ogen)ases |
|||||
| Classification |
Disseminated intravascular coagulation (DIC)
plus
|
Fibrinogen turnover
plus
reactive (secondary) fibrinolysis |
Primary hyperfibrin(ogen)olysis |
|||||
Table 5.3 Sensitivities of prothrombin time (PT), activated partial thromboplastin time (aPTT) and thrombin time (TT)
| Haemostatic parameter | Coagulation test |
||||
| prothrombin time (PT) | activated partial thromboplastin time (aPTT) | thrombin time (TT) | desired haemostatic potential in surgical patients who are bleeding⁴ |
||
| Factor IX | not determined | <25–40%¹ | not determined |
minor bleeding: 30–40%, major bleeding: 50–60% |
|
| Factor VIII | not determined | <25–40%¹ | not determined |
minor bleeding: 30%, major bleeding: 80–100% for surgery; 20–40% postoperatively |
|
| Factor VII | <30%¹ | not determined | not determined | >10% | |
| Factor X |
|
<25–40%¹ | not determined | 10–40% | |
| Factor V |
|
<25–40%¹ | not determined | 10–30% | |
| Factor II | <30%¹ | <25–40%¹,³ | not determined | 10–25% | |
| Fibrinogen |
<0.6–0.8 g/l¹ <0.6 g/l² |
<0.6–0.8 g/l¹ <0.2 g/l² |
<0.6 g/l² | 0.5–1.0 g/l | |
| Factor XIII | not determined | not determined | not determined | ||
| Platelets | not determined | not determined | not determined | ||
| FSP total | >50 mg/l² | >50 mg/l² | >50 mg/l² | ||
¹ Santoro 1991
² Barthels and Poliwoda 1987
³ Furie and Furie 1992
4 Menitove 1990