KNOWLEDGE SUMMARY

Keywords: SEA TURTLE; BLOOD; HAEMATOLOGY; BIOCHEMISTRY; COLD-STUNNING; COLD-STUN; ACIDOSIS

Prognostic value of haematological indices in sea turtles presenting for cold-stunning (sustained hypothermia)

McCaide Wooten, DVM MPVM^1*

¹ UC Davis School of Veterinary Medicine, 1 Garrod Dr., Davis, CA 95616, USA
* Corresponding author email: caide.wooten.dvm@gmail.com

Vol 8, Issue 1 (2023)
Submitted 10 Dec 2021; Published: 01 Mar 2023; next review: 06 Dec 2024
DOI: https://doi.org/10.18849/ve.v8i1.561

PICO question

In sea turtles presenting for cold-stunning (sustained hypothermia), what blood analytes routinely evaluated at intake provide the most prognostic value?

Clinical bottom line

Category of research

Prognosis.

Number and type of study designs reviewed

Ten studies were included in this evaluation including the following study designs: eight retrospective case series, one cross-sectional, and one retrospective cohort.

Strength of evidence

Weak.

Outcomes reported

The most consistent finding across all included studies in cold-stunned sea turtles was acidosis (suspected both respiratory or metabolic components) characterised by reduced blood pH, elevated partial pressure of carbon dioxide (pCO₂), and reduced partial pressure of oxygen (pO₂). However, this finding was not necessarily linked with failure of rehabilitation. Rather, sea turtles presenting for cold-stunning that did not survive rehabilitative therapy were typically in extreme states of homeostatic derangement involving acidosis, but often in conjunction with additional abnormalities (e.g. anaemia, sepsis, organ failure or dysfunction, pneumonia, etc.).

Conclusion

As might be expected, the evaluated literature did not reveal a single or series of blood analytes that were definitively linked with the success or failure of rehabilitation in sea turtles presenting for cold-stunning. However, they did identify analytes that may provide the most clinical value in this clinical situation including packed cell volume (PCV), estimated white blood cell count (WBC), total and / or ionised calcium, pH, potassium (K), and lactate. Review of the available studies on the topic provides insightful information that can aid clinicians addressing this syndrome to triage and treat affected individuals most effectively. It also elucidated areas of opportunity for further research.

How to apply this evidence in practice

The application of evidence into practice should take into account multiple factors, not limited to: individual clinical expertise, patient’s circumstances and owners’ values, country, location or clinic where you work, the individual case in front of you, the availability of therapies and resources.

Knowledge Summaries are a resource to help reinforce or inform decision making. They do not override the responsibility or judgement of the practitioner to do what is best for the animal in their care.

The evidence

Based on the included studies, the most consistent biochemical finding in the blood of marine turtles presenting to rescue and rehabilitation studies for cold-stunning is acidosis, generally characterised by reduced blood pH, elevated partial pressure of carbon dioxide (pCO₂), and reduced partial pressure of oxygen (pO₂). At present, a respiratory or metabolic aetiology of this finding is undetermined, but it was suggested in several studies there are likely elements of both components contributing to the common finding given the nature of the syndrome.

It should be taken into consideration that nine of the ten studies (Innis et al., 2009; Powell et al., 2021; Stacy et al., 2013; Innis et al., 2007; Innis et al., 2019; Hunt et al., 2012; McNally & Innis, 2020; Keller et al., 2012; and Rockwell et al., 2017) evaluated here emanated from the same institution (The New England Aquarium, Boston, MA, USA), and as such are more subject to internal biases. However, this institution is also one of the most likely to be encountering cold-stunned sea turtles given its location along the northwest Atlantic, where juvenile and subadult animals with lower surface area-to-body mass ratios are found to frequent shallower coastal waters that are subject to the effects of rapid oceanic, meteorological, and atmospheric changes.

Also of note is the variability in study designs, outcomes of interest, and population sizes. These factors varied widely between the evaluated manuscripts and the strict evaluation of blood parameters as they pertained to the survival or death of cold-stunned turtles was infrequently the primary objective. Only three of seven known species of extant marine turtles were evaluated here, and of those, the Kemp’s ridley sea turtle (Lepidochelys kempii) is overrepresented, so blind generalisation of the findings herein is not advised; again, this is likely secondary to these reports primarily emanating from one institution and the nature of the Kemp’s ridley sea turtle being the most frequently encountered species succumbing to cold-stunning along the New England coastline. Further characterisation of blood parameters in cold-stunned sea turtles is needed in additional geographic regions and among additional species, where applicable, for better understanding of the physiological impacts of this syndrome. Given projections for increased cold-stunning events secondary to anthropogenically-influenced warming ocean surface temperatures (Griffin et al., 2019), opportunities for study and refinement of clinical methods for rehabilitation will only increase.

The overall presented evidence is deemed to be of ‘weak’ strength. However, the author finds the included materials to be of utmost academic, and particularly clinical, relevance.

Summary of the evidence

Innis et al. (2007)

 

Innis et al. (2009)

 

Anderson et al. (2011)

 

Hunt et al. (2012)

 

Keller et al. (2012)

 

Stacy et al. (2013)

 

Rockwell et al. (2017)

 

Innis et al. (2019)

 

McNally & Innis (2020)

 

Powell et al. (2021)

 

Appraisal, application and reflection

For clarity, the most relevant haematological and biochemical parameters evaluated in the included studies are categorised, analysed, and summarised below:

Haematological indices

Elevated packed cell volume* (PCV) values at first blood evaluation of affected turtles were observed in multiple studies. In general, this finding was attributed to haemoconcentration given the tendency for convalescent PCV values to decrease in conjunction with other biochemical indicators of dehydration (Innis et al., 2009; Keller et al., 2012; and Rockwell et al., 2017). Keller et al. (2012) and Innis et al. (2009) also indicate a wide range of presenting PCVs, with several individuals presenting with anaemia. This finding tended to be associated with individuals that presented with one or more chronic comorbidities and was not necessarily found to be directly associated with cold-stunning.¹

Regarding the leukogram, a common finding was for cold-stunned animals to present with elevated total white blood cell counts (WBC) (Innis et al., 2009; Powell et al., 2021; Hunt et al., 2012; and McNally & Innis, 2020). McNally and Innis (2020) postulated the elevated WBC observed in their study to most likely be linked with stress given the concurrent findings of heterophilia, lymphopenia, eosinopenia, and increased heterophil-to-lymphocyte ratio. As Hunt et al. (2012) showed, a component of corticosterone influence appears likely in cold-stunning cases. Interestingly, that study also revealed that non-surviving turtles had significantly lower WBCs at admission compared to survivors; although the finding of total leukopenia was likely multi-factorial in origin, this finding may suggest that leukopenia in the context of cold-stunning could be a negative prognostic indicator.

*It should also be acknowledged that the evaluated manuscripts use both ‘packed cell volume’ and ‘haematocrit’ to describe the metric derived by determining the percent volumetric contribution of red blood cells to a whole blood sample following centrifugation. By definition, this is a ‘packed cell volume’. ‘Haematocrit’ is the terminology given to the metric generated by multiplying mean cell volume and red blood cell count that have been directly measured by an automated haematology analyser. However, since there are currently no haematology analysers calibrated to evaluate avian or herptile blood samples due to the nucleation of red blood cells in these species, it is not necessarily incorrect to use these terms interchangeably in this context.

Potassium

Significantly higher potassium (K) concentrations were reported in non-surviving turtles vs surviving turtles on initial evaluation in three different studies (Innis et al., 2009; Innis et al., 2019; and Keller et al., 2012). K values were also included in all mortality prediction indices (MPIs) developed by Stacy et al. (2013) with higher values associated with higher index values. All of these studies evaluated potassium in Kemp’s ridley turtles, and the corresponding findings were consistent with previously reported potassium findings in cold-stunned sea turtles (Carminati et al., 1994; and Turnbull et al., 2000). Conversely, several studies found initially decreased K levels relative to a control population (Anderson et al., 2011) and convalescent values in recovered turtles (Powell et al., 2021; and McNally & Innis, 2020). These three studies evaluated green (Anderson et al., 2011) and loggerhead (Powell et al., 2021; and McNally & Innis, 2020) turtles.

The observed discord between studies regarding potassium is not fully explained. The observation by Anderson et al. (2011) that differences in potassium concentrations ‘may reflect differences in species, environmental conditions, sampling times, and methodology among studies’ (p. 252) remains poignant given the currently available evidence; animal age (juvenile vs subadult), chronicity of hypothermia, renal functionality, and blood pH status (Lutz et al., 1989) are other suggested factors at play. However, what does appear to be clear is that there is a negative prognostic relationship with severe hyperkalaemia in the Kemp’s ridley.

Acid-base indices

Contrary to what is reported in experimental studies of blood pH and hypothermia in sea turtles (Kraus et al., 1980; Lutz et al., 1989; and Moon et al., 1997), the presence of relative physiological acidosis in cold-stunned turtles was noted across multiple papers (with temperature correction) (Anderson et al., 2011; Keller et al., 2012; Innis et al., 2007; Innis et al., 2019; Rockwell et al., 2017; and Stacy et al., 2013).Metabolic and respiratory origins for this finding are both supported due to hypoventilation (with secondary hypercarbia and anaerobic metabolic processes), reduced perfusion at lower body temperatures, reduced / exhausted bicarbonate buffering capacity, and reduced renal clearance. Keller et al. (2012) reported significantly increased pCO₂ and significantly reduced pO₂, pH, and HCO₃ in non-surviving turtles compared to those that survived cold-stunning. Mortalities in the Innis et al. (2019) report were also linked with significantly elevated pCO₂ and significantly reduced pH and pO₂ relative to surviving loggerheads. pH, pCO₂, and pO₂ were also included across all MPIs developed by Stacy et al. (2013) where decreased pH, elevated pCO₂, and decreased pO₂ increased likelihood of mortality. This evidence supports the use of marked blood gas derangements, when corrected for temperature, for inclusion as a negative prognostic indicator in cold-stunned turtles.

Calcium

Relative to convalescent values, initial ionised calcium (Innis et al., 2007; Innis et al., 2019; and Keller et al., 2012) or total calcium (Innis et al., 2009; and McNally & Innis, 2020) values of surviving turtles were reduced at presentation across multiple studies. Anderson et al. (2011) also noted hypocalcaemia (total and ionised) in cold-stunned individuals relative to controls. Interestingly, for turtles that did not survive rehabilitation, Keller et al. (2012) reported significant increases in ionised calcium over the first few days of hospitalisation relative to survivors; Innis et al. (2007) also showed significantly increased total calcium among non-survivors at presentation relative to survivors. The mechanism for this disparity has not been fully described, but for the turtles that died, the authors suggested that compromised renal function, loss of cation homeostatic mechanisms, compensatory homeostatic responses to acidosis, and / or iatrogenic causes could be factoring into the observation. Given these observations, the prognostic value of ionised or total calcium are still undetermined. Hypocalcaemia appears that it should be an expected finding in cold-stunned turtles, but further investigation into calcium status as it pertains to this syndrome is needed to describe how changes in calcium over time might be associated with mortality.

Lactate

The use of point- and serial-lactate evaluation as a means of identifying the magnitude of anaerobic metabolism occurring in domestic dogs and cats presenting to emergency facilities has become a classic example of a prognostic test in the veterinary literature in recent years (Kohen et al., 2018; Saint-Pierre et al., 2021; and Zacher et al., 2010). In the evaluated studies, lactate values in cold-stunned turtles did not exceed levels reported in healthy sea turtles exposed to forced submergence, extended voluntary dives, or trawl net capture (Berkson, 1966; Lutz et al., 1985; Stabenau et al., 1991; and Wood et al., 1984). Innis et al. (2007) noted that initial lactate concentrations were significantly elevated relative to convalescent values. A later study showed significantly greater lactate levels in turtles that died vs those that survived (Innis et al., 2019). Keller et al. (2012) demonstrated evidence of significantly different net positive change of lactate values (despite medical intervention) over the first 2 to 3 days of hospitalisation in cold-stunned turtles that died relative to net negative change in turtles that survived over the same period. This latter study showed that there may be a negative prognostic relationship with serially increasing lactate concentrations following hospitalisation for cold-stunning. More recent evidence has shown that serum lactate levels in clinically healthy loggerhead turtles restrained for health exams increase significantly over 15 minutes of restraint (Mones et al., 2021). In this context, a clinician must keep in mind the time at which a lactate sample, or series of samples, was taken in order to appropriately interpret the findings.

Free thyroxine

Though only evaluated in one of the included manuscripts (Hunt et al., 2012) and not necessarily identified as a prognostic indicator for release, free thyroxine (fT4) is identified here given its potential usefulness in connection to return to normal function in sea turtles being rehabilitated for cold-stunning. Hunt et al. (2012) showed that in the week prior to sampling, hospitalised turtles that were documented to be eating every day and scored as ‘active’ (vs ‘quiet’) had significantly increased fT4 concentrations over those that were not. This finding shows value for further investigation into thyroid hormone activity in sea turtles and the potential application of T4 supplementation in sea turtles being rehabilitated for cold-stunning.

Conclusions

The summative findings of the reviewed articles most strongly indicate the primary pathological effect of cold-stunning in the studied species of sea turtles, particularly the Kemp’s ridley, to be a physiological acidosis (likely both metabolic and respiratory in origin). Secondary imbalances in electrolyte status, immune status, and cation balance appear to ensue in a compensatory fashion and anaerobic metabolic processes generally coincide. Frequently, sea turtles presenting for cold-stunning may also present with other comorbidities (sepsis, pneumonia, anaemia, etc.). Prognostically, total leukopenia, extreme derangements of blood acid-base parameters (increased pCO₂, decreased pO₂, reduced blood pH), severe hyperkalaemia, serially increasing blood lactate concentrations, and possibly rapid increases in total or ionised calcium have been identified in association with increased likelihood of death.

Areas identified for potential research include, but are not limited to, further characterisation of the metabolic processes affected by the speed of rewarming and optimisation to prevent exacerbation of the acidotic state in cold-stunned turtles, description of baseline thyroid activity in healthy sea turtles, implementation of thyroid supplementation in turtles showing reduced activity and / or hyporexia while in rehabilitation, evaluation of serial lactate values across sea turtle species presenting for rehabilitation, and comparative study of blood, gross pathological, and histopathological findings in sea turtles that succumb to cold-stunning.

The current recommendations for rehabilitation of cold-stunned sea turtles (Innis & Staggs, 2017; and Wyneken et al., 2006) generally result in favourable outcomes with 50–80% of animals being successfully rehabilitated and released (Innis & Staggs, 2017). When turtles that succumb to the syndrome within the first 3 days of treatment are excluded, that statistic approaches 90% (Innis & Staggs, 2017; and Wyneken et al., 2006). With continued improvement of the existent protocols designed to remedy this syndrome for the vulnerable species experiencing it, these statistics can only improve.

Methodology

Search Strategy

 

Exclusion / Inclusion Criteria

 

Search Outcome

 

ORCID

McCaide Wooten: https://orcid.org/0000-0001-8121-0824

Conflict of Interest

The author declares no conflicts of interest.

References

1. Anderson, E. T., Harms, C. A., Stringer, E. M. & Cluse, W. M. (2011). Evaluation of hematology and serum biochemistry of cold-stunned green sea turtles (Chelonia mydas) in North Carolina, USA. Journal of Zoo and Wildlife Medicine. 42(2), 247–255. DOI: https://doi.org/10.1638/2010-0217.1
2. Berkson, H. (1966). Physiological adjustments to prolonged diving in the Pacific green turtle (Chelonia mydas agassizii). Comparative Biochemistry and Physiology. 18(1), 101–119. DOI: https://doi.org/10.1016/0010-406X(66)90335-5
3. Carminati, C., Gerle E, Kiehn LL. & Pisciotta RP. (1994). Blood chemistry comparison of healthy vs. hypothermic juvenile Kemp’s ridley sea turtles (Lepidochelys kempii) in the New York Bight. Proceedings of the 14th Annual Workshop on Sea Turtle Conservation and Biology. 203–207.
4. Griffin, L. P., Griffin, C. R., Finn, J. T., Prescott, R. L., Faherty, M., Still, B. M., & Danylchuk, A. J. (2019). Warming seas increase cold-stunning events for Kemp’s ridley sea turtles in the northwest Atlantic. PLoS ONE. 14(1). DOI: https://doi.org/10.1371/journal.pone.0211503
5. Hunt, K. E., Innis, C. & Rolland, R. M. (2012). Corticosterone and thyroxine in cold-stunned Kemp’s ridley sea turtles (Lepidochelys kempii). Journal of Zoo and Wildlife Medicine. 43(3), 479–493. DOI: https://doi.org/10.1638/2011-0149R1.1
6. Innis, C. J., Tlusty, M., Merigo, C. & Weber, E. S. (2007). Metabolic and respiratory status of cold-stunned Kemp’s ridley sea turtles (Lepidochelys kempii). Journal of Comparative Physiology. B, Biochemical, Systemic, and Environmental Physiology. 177(6), 623–630. DOI: https://doi.org/10.1007/S00360-007-0160-9
7. Innis, C. J., Ravich, J. B., Tlusty, M. F., Hoge, M. S., Wunn, D. S., Boerner-Neville, L. B., Merigo, C. & Weber, E. S. (2009). Hematologic and plasma biochemical findings in cold-stunned Kemp’s ridley turtles: 176 cases (2001–2005). Journal of the American Veterinary Medical Association. 235(4), 426–432. DOI: https://doi.org/10.2460/javma.235.4.426
8. Innis, C. & Staggs, L. (2017). Cold-Stunning. In: C. Manire, T. Norton, B. Stacy, C. Innis, & C. Harms, eds., Sea Turtle Health & Rehabilitation. Plantation, FL: J. Ross, 675–687.
9. Innis, C. J. (2019). Medical management and rehabilitation of sea turtles. In: S. Divers & S. Stahl, eds., Mader’s Reptile and Amphibian Medicine and Surgery, 3rd ed. St. Louis, MO: Elsevier, 1382–1388.
10. Innis, C. J., McGowan, J. P. & Burgess, E. A. (2019). Cold-stunned loggerhead sea turtles (Caretta caretta): initial vs. convalescent physiologic status and physiologic findings sssociated with death. Journal of Herpetological Medicine and Surgery. 29(3–4), 105–112. DOI: https://doi.org/10.5818/19-06-204.1
11. Keller, K. A., Innis, C. J., Tlusty, M. F., Kennedy, A. E., Bean, S. B., Cavin, J. M. & Merigo, C. (2012). Metabolic and respiratory derangements associated with death in cold-stunned Kemp’s ridley turtles (Lepidochelys kempii): 32 cases (2005–2009). Journal of the American Veterinary Medical Association. 240(3), 317–323. DOI: https://doi.org/10.2460/JAVMA.240.3.317
12. Kohen, C. J., Hopper, K., Kass, P. H. & Epstein, S. E. (2018). Retrospective evaluation of the prognostic utility of plasma lactate concentration, base deficit, pH, and anion gap in canine and feline emergency patients. Journal of Veterinary Emergency and Critical Care. 28(1), 54–61. DOI: https://doi.org/10.1111/vec.12676
13. Kraus, D. R. & Jackson, D. C. (1980). Temperature effects on ventilation and acid-base balance of the green turtle. American Journal of Physiology-Regulatory, Integrative and Comparative Physiology. 239(3), R254–R258. DOI: https://doi.org/10.1152/ajpregu.1980.239.3.R254
14. Lutz, P. L. & Bentley, T. B. (1985). Respiratory Physiology of Diving in the Sea Turtle. Copeia. 1985(3), 671. DOI: https://doi.org/10.2307/1444761
15. Lutz, P. L., Bergey, A. & Bergey, M. (1989). Effects of temperature on gas exchange and acid-base balance in the sea turtle Caretta caretta at rest and during routine activity. Journal of Experimental Biology. 144(1), 155–169. DOI: https://doi.org/10.1242/jeb.144.1.155
16. McNally, K. L. & Innis, C. J. (2020). Plasma biochemistry and hematologic values of cold-stunned loggerhead sea turtles (Caretta caretta). Journal of Herpetological Medicine and Surgery. 30(2), 88. DOI: https://doi.org/10.5818/19-08-209.1
17. Mones, A. B., Gruber, E. J., Harms, C. A., Lohmann, C. M. F., Lohmann, K. J. & Lewbart, G. A. (2021). Lactic acidosis induced by manual restraint for health evaluation and comparison of two point-of-care analyzers in healthy loggerhead sea turtles (Caretta caretta). Journal of Zoo and Wildlife Medicine. 52(4), 1195–1204. DOI: https://doi.org/10.1638/2021-0029
18. Moon, D. Y., MacKenzie, D. S. & Owens, D. W. (1997). Simulated hibernation of sea turtles in the laboratory: I. feeding, breathing frequency, blood pH, and blood gases. The Journal of Experimental Zoology. 278(6), 372–380.
19. Powell, A. L., Tuxbury, K. A., Cavin, J. M., Stacy, B. A., Frasca, S., Stacy, N. I., Brisson, J. O., Solano, M., Williams, S. R., McCarthy, R. J. & Innis, C. J. (2021). Osteomyelitis in cold-stunned Kemp’s ridley sea turtles (Lepidochelys kempii) hospitalized for rehabilitation: 25 cases (2008–2018). Journal of the American Veterinary Medical Association. 259(10), 1206–1216. DOI: https://doi.org/10.2460/JAVMA.20.08.0443
20. Rockwell, K. E., Innis, C. J., Merigo, C. & Prescott, R. (2017). The effect of delayed hospitalization in cold-stunned Kemp’s ridley turtles (Lepidochelys kempii). Journal of Herpetological Medicine and Surgery. 27(3), 93–96. DOI: https://doi.org/10.5818/17-05-114.1
21. Saint-Pierre, L. M., Hopper, K. & Epstein, S. E. (2021). Retrospective evaluation of the prognostic utility of plasma lactate concentration and serial lactate measurements in dogs and cats presented to the emergency room (January 2012 – December 2016): 4863 cases. Journal of Veterinary Emergency and Critical Care. 32(1), 1–151. DOI: https://doi.org/10.1111/vec.13106
22. Stabenau, E. K., Heming, T. A. & Mitchell, J. F. (1991). Respiratory, acid-base and ionic status of Kemp’s ridley sea turtles (Lepidochelys kempi) subjected to trawling. Comparative Biochemistry and Physiology Part A: Physiology. 99(1–2), 107–111. DOI: https://doi.org/10.1016/0300-9629(91)90243-6
23. Stacy, N. I., Innis, C. J. & Hernandez, J. A. (2013). Development and evaluation of three mortality prediction indices for cold-stunned Kemp’s ridley sea turtles (Lepidochelys kempii). Conservation Physiology. 1(1). DOI: https://doi.org/10.1093/CONPHYS/COT003
24. Turnbull, B. S., Smith, C. R. & Stamper, A. M. (2000). Medical implications of hypothermia in threatened loggerhead (Caretta caretta) and endangered Kemp’s ridley (Lepidochelys kempi) and green (Chelonia mydas) sea turtles. Joint Conference of the American Association of Zoo Veterinarians and the International Association of Aquatic Animal Medicine. 31–35. [online]. Available from: https://www.vin.com/doc/?id=3864468 [Accessed 29 Nov 2021].
25. Wood, S. C., Gatz, R. N. & Glass, M. L. (1984). Oxygen transport in the green sea turtle. Journal of Comparative Physiology B. 154(3), 275–280. DOI: https://doi.org/10.1007/BF02464407
26. Wyneken, J., Mader, D. R., Weber, E. S. & Merigo, C. (2006). Medical care of sea turtles. In: D. Mader, ed., Reptile Medicine and Surgery, 2^nd ed. St. Louis, MO: Elsevier, 972–1007.
27. Zacher, L. A., Berg, J., Shaw, S. P. & Kudej, R. K. (2010). Association between outcome and changes in plasma lactate concentration during presurgical treatment in dogs with gastric dilatation-volvulus: 64 cases (2002–2008). Journal of the American Veterinary Medical Association. 236(8), 892–897. DOI: https://doi.org/10.2460/javma.236.8.892

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