Karina Acevedo-Whitehouse

Activation of an inflammatory response is context-dependent during early development of the California sea lion

Variations in immune function can arise owing to trade-offs, that is, the allocation of limited resources among costly competing physiological functions. Nevertheless, there is little information regarding the ontogeny of the immune system within an ecological context, and it is still unknown whether development affects the way in which resources are allocated to different immune effectors. We investigated changes in the inflammatory response during early development of the California sea lion (Zalophus californianus) and examined its association with body condition, as a proxy for the availability of energetic resources. We found that the relationship between inflammation and body condition varied according to developmental stage and circulating levels of leucocyte populations, a proxy for current infection. Body condition was related to the magnitude of the inflammatory response during two of the three developmental periods assessed, allowing for the possibility that the availability of pup energetic reserves can limit immune function. For older pups, the ability to mount an inflammatory response was related to their circulating levels of neutrophils and the neutrophil to lymphocyte ratio, implying that the infection status of an individual will influence its ability to respond to a new challenge. Our results suggest that trade-offs may occur within the immune system and highlight the importance of taking into account ontogeny in ecoimmunological studies.

Melanin granules melanophages and a fully-melanized epidermis are common traits of odontocete and mysticete cetaceans

BACKGROUNDnThe cellular mechanisms used to counteract or limit damage caused by exposure of marine vertebrates to solar ultraviolet (UV) radiation are poorly understood. Cetaceans are vulnerable because they lack protective skin appendages and are obliged to surface continuously to breathe, thus being exposed repeatedly to UV light. Although molecular mechanisms of photoprotection of cetaceans have been studied, there is limited knowledge about their epidermal structure and photoprotective effectors.nnnOBJECTIVEnTo describe and compare the epidermis of mysticete and odontocete cetaceans and identify potentially photoprotective traits.nnnANIMALSnTwenty eight free-living individuals belonging to six cetacean species were sampled in the Mexican Central Pacific and Gulf of California. Species sampled were the bottlenose dolphin, pantropical spotted dolphin, spinner dolphin, Brydes whale, fin whale and humpback whale.nnnMETHODSnHistological and cytological evaluation of skin biopsy tissue collected in the field between 2014 and 2016.nnnRESULTSnAll cetaceans had only three epidermal layers, lacking both the stratum granulosum and stratum lucidum. A relatively thick stratum corneum with a parakeratosis-like morphology was noted. Melanin was observed within keratinocytes in all epidermal layers, including the stratum corneum and apical melanin granules obscured the keratinocyte nucleus. Keratinocytes had a perinuclear halo. Keratinocyte diameter differed between cetacean suborders and amongst species. Melanophage clusters were common in most cetacean species.nnnCONCLUSIONSnThe widespread presence of melanin and the unexpectedly high number of melanophages may constitute a unique photoprotective trait of cetaceans and could reflect primitive adaptations to their environment and to their obligate marine-bound life.

Climatic anomaly affects the immune competence of California sea lions

The past decades have been characterized by a growing number of climatic anomalies. As these anomalies tend to occur suddenly and unexpectedly, it is often difficult to procure empirical evidence of their effects on natural populations. We analysed how the recent sea surface temperature (SST) anomaly in the northeastern Pacific Ocean affects body condition, nutritional status, and immune competence of California sea lion pups. We found that pup body condition and blood glucose levels of the pups were lower during high SST events, although other biomarkers of malnutrition remained unchanged, suggesting that pups were experiencing early stages of starvation. Glucose-dependent immune responses were affected by the SST anomaly; specifically, pups born during high SST events had lower serum concentrations of IgG and IgA, and were unable to respond to an immune challenge. This means that not only were pups that were born during the SST anomaly less able to synthesize protective antibodies; they were also limited in their ability to respond rapidly to nonspecific immune challenges. Our study provides empirical evidence that atypical climatic conditions can limit energetic reserves and compromise physiological responses that are essential for the survival of a marine top predator.

Protein and RNA extraction from mucosal swabs: a minimally invasive source of ecological data for studies of natural populations

1.Mucosal swabs have long been used to study various physiological processes in humans. In contrast, systematic sampling of mucosae is rare for wild animals, and except for its use as a source of DNA for population genetics, its potential as a tool to study physiologically-relevant processes of natural populations has not been explored fully. n n2.We collected swabs from the rectal, genital, nasal and oral mucosae of California sea lion pups and investigated differences in RNA and protein yield, quality, and integrity between storage times and mucosal types. Downstream applications were tested on the extracted products to determine the potential value of mucosal sampling in free-ranging mammals. n n3.For most samples, RNA yield was stable regardless of storage time, and RNA quality and integrity was equal for all mucosal types. Ribosomal fragments of sizes expected for mammals, yeast and bacteria were observable in genital and rectal samples, but in nasal and oral samples only bacterial ribosomal fragments were observable. Amplification of selected transcripts was successful for all samples. The protein profile was distinct between mucosae, and samples with high protein yields were useful for antibody detection. n n4.Our study demonstrates that mucosal swabbing is a minimally-invasive tool that yields useful physiological data for free ranging wildlife. Minimum experience is needed to collect samples, processing is inexpensive and downstream applications for ecological studies are realistic. n nThis article is protected by copyright. All rights reserved.

MHC class II DRB diversity predicts antigen recognition and is associated with disease severity in California sea lions naturally infected with Leptospira interrogans

We examined the associations between California sea lion MHC class II DRB (Zaca-DRB) configuration and diversity, and leptospirosis. As Zaca-DRB gene sequences are involved with antigen presentation of bacteria and other extracellular pathogens, we predicted that they would play a role in determining responses to these pathogenic spirochaetes. Specifically, we investigated whether Zaca-DRB diversity (number of genes) and configuration (presence of specific genes) explained differences in disease severity, and whether higher levels of Zaca-DRB diversity predicted the number of specific Leptospira interrogans serovars that a sea lions serum would react against. We found that serum from diseased sea lions with more Zaca-DRB loci reacted against a wider array of serovars. Specific Zaca-DRB loci were linked to reactions with particular serovars. Interestingly, sea lions with clinical manifestation of leptospirosis that had higher numbers of Zaca-DRB loci were less likely to recover from disease than those with lower diversity, and those that harboured Zaca-DRB.C or -G were 4.5 to 5.3 times more likely to die from leptospirosis, regardless of the infective serovars. We propose that for leptospirosis, a disadvantage of having a wider range of antigen presentation might be increased disease severity due to immunopathology. Ours is the first study to examine the importance of Zaca-DRB diversity for antigen detection and disease severity following natural exposure to infective leptospires.

Atypical Red Blood Cells Are Prevalent in California Sea Lion Pups Born during Anomalous Sea Surface Temperature Events

To date, there is limited knowledge of the effects that abnormal sea surface temperature (SST) can have on the physiology of neonate pinnipeds. However, maternal nutritional deficiencies driven by alimentary restrictions would expectedly impact pinniped development and fitness, as an adequate supply of nutrients is essential for growth and proper functioning of all body systems, including red blood cell synthesis and clearance. Here, we investigated red blood cell morphology of California sea lion (CSL) pups from the San Benito Archipelago born during the 2014 and 2015 anomalously high SST events recorded in the northeastern Pacific Ocean. We examined whether atypical erythrocyte morphologies were more common in 2015, when the high SST event was more pronounced, and whether the stable isotope signature of pup fur, as an indicator of maternal feeding strategies, accounted for the number of atypical cells. Various atypical erythrocyte morphologies were more prevalent and more abundant than reference values. Evidence of iron deficiency was found in both years, and only pups born in 2014 showed evidence of active erythropoiesis. Microcytes and reticulocytes were more common in pups with higher isotopic δ13C and lower δ15N values, suggesting a probable relationship between maternal feeding strategies and the effect of climatic anomalies on red blood cell physiology of their pups. As developing pinnipeds require increased oxygen storage capacity for diving and foraging, the presence of atypical erythrocytes could be relevant to CSL pup fitness if the underlying cause is not reverted. This study is a first step to explore the effects that climatic alterations in the marine environment can have on the blood physiology of developing individuals.

Changes in mucosal and serum immunoglobulin levels of California sea lions during early development

To date, most studies on pinniped immunoglobulins have focused on circulating antibodies. However, systemic and local immune activities differ in terms of maturation, intensity, and types of effectors that participate. Here, we examined levels of three immunoglobulin isotypes, IgG, IgM and IgA, in the blood and mucosal membranes of free-living California sea lion pups. We investigated whether age, body condition and sex influenced their concentration. Isotype levels varied among tissues, with age-related patterns that could be indicative of differential regulation along development. Serum IgG and IgA increased linearly with age, reaching adult levels at five months of age, while IgM remained unchanged. Mucosal isotypes tended to be low in newborns and remained so until five months of age. Regardless of age, pups with better condition tended to have higher anal IgG levels and higher genital IgA levels, suggesting that their synthesis and transport to the mucosal membranes is costly. Intersex differences were only observed in the genital mucosa, where all isotypes differed between male and female pups, regardless of age, presumably due to histological and anatomical differences.

MHC class II transcription is associated with inflammatory responses in a wild marine mammal.

Inflammation is one of the most important non-specific and rapid responses that a vertebrate can elicit in response to damage or a foreign insult. To date, despite increasing evidence that the innate and adaptive branches of immunity are more intricately related than previously thought, few have examined interactions between the Major Histocompatibility Complex (MHC, a polymorphic region of the vertebrate genome that is involved with antigen presentation) and inflammation, and even less is known about these interactions in an eco-immunological context. Here, we examined the effect of MHC class II DRB gene multiplicity and transcription on phytohemagglutinin (PHA)-induced inflammation during the early stages of development of California sea lions. Neither constitutive nor expressed ZacaDRB diversity was found to be associated with pup responses to PHA at any of the stages of pup development. However, for two-month-old pups, those with a specific MHC-DRB locus (ZacaDRB-A) tended to have less efficient responsive inflammation. Transcription of distinct MHC-DRB loci was also linked to PHA-induced inflammation, with patterns that varied markedly between ages, and that suggested that ongoing infectious processes could limit the capacity to respond to a secondary challenge. Life history constraints and physiological processes associated with development of California sea lions, in conjunction with their changing pathogenic environment could explain the observed effects of MHC class II transcription on PHA-induced inflammation. To our knowledge, ours is the first study to examine the importance of expressed vs. constitutive MHC loci on inflammation in a natural population.