The Influence of Environmental Conditions and Co-Occurring Parasites on Blood Physiology in Eastern Hellbenders (Cryptobranchus alleganiensis)

Climate change, habitat degradation, and infectious disease are major drivers of global amphibian declines. Amphibians are particularly susceptible to these factors due to their unique physiology and habitat requirements. Thus, investigating components of amphibian physiology and evaluating the influence of environmental conditions, perceived threats, and encounters with infectious agents is essential to conserving imperiled amphibian species. Here, I measured hematocrit, hemoglobin, relative proportions of polychromatic red blood cells, and white blood cell differentials in a wild population of male Eastern Hellbender salamanders (Cryptobranchus alleganiensis) which remain with their nest for up to ~8 months and have frequent encounters with parasitic leeches (Placobdella appalachiensis) that transmit hematophagous endoparasites (Trypanosoma sp.), often resulting in coinfection. Results indicate that hematocrit and hemoglobin increase in response to acute stress and with temperature. Additionally, the magnitude of the stress-induced hemoconcentration response was greater at lower temperatures. Hellbenders exhibited an increase in the proportion of neutrophils and eosinophils in circulation as temperatures decreased while the proportions of lymphocytes and basophils had an inverse effect. Furthermore, the proportion of neutrophil precursors also increased as temperature decreased, which signifies recruitment of innate immune cells during seasonally cold periods. Coinfection of the leech and trypanosome parasites resulted in decreased hematocrit and hemoglobin and a marked increase in polychromatic red blood cells which is indicative of regenerative anemia in the hellbender host. However, these effects were not present in individuals only infected with trypanosomes, implicating the leech vector as the key contributor to anemia in hellbenders. Moreover, the proportion of neutrophils and eosinophils increased, while lymphocytes decreased, in response to leech attachment. However, as parasitemia of leeches and trypanosomes increased concurrently, the proportion of lymphocytes increase in circulation while neutrophils and eosinophils decrease, underscoring the complexities associated with coinfection and multi-parasite interactions. Together, this research provides novel insights into the blood physiology of an imperiled salamander by establishing reference values essential for population surveillance while also describing how these values fluctuate across season and in response to extrinsic factors, with an emphasis on co-occurring hematophagous parasites.