Mitogen-induced lymphocyte proliferation in loggerhead sea turtles: comparison of methods and effects of gender, plasma testosterone concentration, and body condition on immunity.” Vet

Abstract A fully functioning immune system is vital to the survival of threatened and endangered sea turtles. Immunological protection against diseases in any organism can be reduced by a number of natural and anthropogenic factors, such as seasonal changes, malnutrition, disease states, and contaminant exposure. These factors are even more critical when they occur in endangered species or populations. To identify alterations in the immunological health of loggerhead sea turtles (Caretta caretta), the mitogen-induced lymphocyte proliferation (LP) assay was developed using peripheral blood leukocytes (PBLs). Collection and culture conditions were optimized for this assay using non-lethal blood samples collected from free-ranging turtles along the southeastern US coast. During the collection, two anticoagulants (sodium heparin and lithium heparin) were compared to determine effects of different ions on assay results. Optimal culture conditions were established for loggerhead PBLs while two different methods of measuring LP were compared: (1) the traditional radioactive heparin type did not influence the results of the LP assay. Lastly, using these optimized methods, we investigated the effect of gender, plasma testosterone concentration, and body condition on LP in loggerhead turtles and found that none of the parameters largely influenced LP.

Comparison of blood values in foraging, nesting, and stranded loggerhead turtles (Caretta caretta) along the coast of Georgia, USA

The health status of 83 loggerhead sea turtles (Caretta caretta; 39 foraging, 31 nesting, and 13 stranded turtles) was analyzed using physical examinations, hematology, plasma biochemistry, plasma protein electrophoresis, and toxicologic parameters. Significant differences were noted in a number of health parameters between turtles exhibiting each of these behaviors. On physical examinations, stranded turtles had the highest prevalence of heavy carapace epibiont loads, miscellaneous abnormalities, emaciation, and weakness. Differences in hematologic values included a lower packed cell volume, higher number of lymphocytes, and lower number of monocytes in stranded turtles; lower white blood cell counts in foraging turtles; and significant differences in total solid values among turtles exhibiting all behaviors with the lowest values in stranded turtles and the highest values in nesting turtles. Differences in plasma biochemistry values included the highest uric acid, creatine kinase, and CO(2) values in stranded turtles; the highest glucose and potassium values in foraging turtles; and the highest cholesterol and triglyceride values, and lowest alanine aminotransferase, in nesting turtles. Differences in total protein, albumin, and globulin were found using plasma biochemistry values, with lowest values in stranded turtles and highest values in nesting females, whereas differences in blood urea nitrogen between turtles included the lowest values in nesting turtles and the highest in foraging turtles. Plasma organochlorine and polychlorinated biphenyl levels were below their limits of quantification in the 39 foraging, 11 nesting, and three stranded turtles tested. A statistically significant difference was noted in the level of whole blood mercury between the 23 foraging and 12 nesting turtles tested. There was no difference in arsenic or lead levels between turtles exhibiting any of the three behaviors. Although a few limitations exist with the present study and include unknown ambient temperatures, turtle handling times that varied from 15 min to 53 min per turtle, and the use of a different laboratory for processing complete blood counts and plasma biochemistries in stranded versus foraging and nesting turtles, we provide baseline blood values for two cohorts (foraging and nesting) of loggerhead sea turtles on the coast of Georgia. Additionally, we demonstrate significant differences in clinical findings and blood parameters between foraging, nesting, and stranded loggerhead turtles in the region

Chronic debilitation in stranded loggerhead sea turtles (<i>Caretta caretta</i>) in the southeastern United States: Morphometrics and clinicopathological findings

<div><p>Chronically debilitated loggerhead sea turtles (<i>Caretta caretta</i>) (DT) are characterized by emaciation, lethargy, and heavy barnacle coverage. Although histopathological findings associated with this condition have been reported, only limited data is available on health variables with clinical application. The objectives of this study were to 1) to compare morphometrics, clinicopathological variables, and immune functions of DTs to a group of apparently healthy loggerhead turtles to better understand the pathophysiology of the condition and 2) to assess health parameters in live debilitated turtles as they recovered during rehabilitation in order to identify potential prognostic indicators. We examined and sampled 43 DTs stranded from North Carolina to Florida for 47 health variables using standardized protocols to further characterize the condition. DTs were grouped into categories of severity of the condition, and those that survived were sampled at four time points through rehabilitation. All groups and time points were compared among DTs and to clinically healthy loggerhead turtles. Compared to healthy turtles, DTs had significantly lower body condition index, packed cell volume (PCV), total white blood cell (WBC) count, lymphocytes, glucose (Glc), total protein, all protein fractions as determined by electrophoresis, calcium (Ca), phosphorus (P), Ca:P ratio, potassium (K), lymphocyte proliferation, and greater heterophil toxicity and left-shifting, uric acid (UA), aspartate aminotransferase, creatine kinase, lysozyme, and respiratory burst. From admission to recovery, hematology and plasma chemistry data improved as expected. The most informative prognostic indicators, as determined by correlations with a novel severity indicator (based on survival times), were plastron concavity, P, albumin, total solids, UA, lymphocyte proliferation, WBC, K, Glc, Ca:P, and PCV. The results of this study document the wide range and extent of morphometric and metabolic derangements in chronically debilitated turtles. Monitoring morphometrics and clinicopathological variables of these animals is essential for diagnosis, treatment, and prognosis during rehabilitation.</p></div

Sample sizes and characteristics of debilitated (DT) and healthy loggerhead turtles sampled in the current study.

<p>Sample sizes and characteristics of debilitated (DT) and healthy loggerhead turtles sampled in the current study.</p

Diagram of severity indicator with five case examples of debilitated loggerhead sea turtles.

<p>Images show turtles at time of admission.</p

Comparison of immune function variables measured in debilitated loggerhead sea turtles (DTs) before and during rehabilitation and compared to healthy loggerhead turtles (means and one standard deviation).

<p>Each turtle group on the x-axis was significantly different (p<0.05 or p<0.00883 for repeated measures) from groups represented by letters above data. (A) lymphocyte proliferation with ConA; (B) lymphocyte proliferation with PHA; (C) respiratory burst with Ca ionophore; and (D) lysozyme activity. Lines represent an individual turtle through rehabilitation. For samples sizes in each diagram, please refer to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0200355#pone.0200355.s007" target="_blank">S4 Table</a>.</p

Comparison of plastron concavity measured in debilitated loggerhead sea turtles (DTs) before and during rehabilitation and compared to healthy loggerhead turtles (means and one standard deviation).

<p>Lines represent an individual turtle through rehabilitation. Each turtle group on the x-axis was significantly different (p<0.05 or p<0.00883 for repeated measures) from groups represented by letters above data. Sample size for each group: 15 [X], 9 [Y], 6 [A]–[D], 1[H].</p

Health variables that significantly correlated with the severity indicator in debilitated loggerhead sea turtles (DTs).

<p>Spearman rank correlation coefficients and p-values are shown. (A) plastron concavity; (B) packed cell volume (PCV); (C) glucose; (D) phosphorus; (E) albumin by bromocresol green method; (F) albumin by plasma electrophoresis; (G) B-lymphocyte proliferation.</p