Progestogen metabolites for use in pregnancy monitoring of 13-lined ground squirrels (Ictidomys tridecemlineatus)

Graphical abstract 13-lined ground squirrels (TLGS; Ictidomys tridecemlineatus) are small, omnivorous, fossorial, hibernating sciurids. TLGS are seasonal induced ovulators, with a ~28-day gestation period. The main goal of this study was to ascertain whether enzyme-linked immunosorbent assay (ELISA) of TLGS fecal samples can be used to non-invasively detect pregnancy. Competitive ELISAs for progestogen metabolites were conducted on feces collected from a group of (n =13) females. Feces were collected thrice weekly during the breeding season and frozen for subsequent analysis. Competitive ELISAs were run using progesterone kits ), setting data against seven different time-points between hibernation, emergence, and litter birthdate. Eleven females produced litters. ELISA data from the (n = 2) non-pregnant females demonstrated no rise in progestogen metabolites at any point over 28 days. In contrast, data from the (n = 11) pregnant females all demonstrated a pronounced rise in progestogen metabolites, with most animals displaying progesterone withdrawal in the final week of gestation. A >20-fold rise in progestogen metabolite was observed halfway through gestation (P < 005). Analysis on litter size and progestogen metabolite concentration showed no significant correlation (r2 = −0.615). Initial correlation analysis done on sex ratio of litters vs progestogen metabolites showed no significant effect of progesterone on sex ratios (males: r2 = −0.772, females: r2 = 0.375). This work demonstrated that TLGS also undergo progesterone withdrawal about a week before parturition. We have ascertained that a commercially available progesterone assay kit can detect a significant elevation in progestogen metabolites in this species about halfway through gestation. Lay summary This research was conducted to discover whether pregnancy prediction is possible in female 13-lined ground squirrels (TLGS; a small hibernating ground squirrel named for their number of stripes). Pregnancy status in this species, we postulated, could be anticipated by generating profiles for individuals via a non-invasive technique known as fecal endocrine hormone profiling. Fecal samples were collected from 13 females thrice weekly for 4 weeks post-hibernation in the breeding season of 2016. Fecal samples were then processed and run through an assay known as an ELISA giving concentrations of hormone metabolites excreted through feces. We then set these samples against time points to develop a profile for each female. We have ascertained that elevated progesterone (potential pregnancy) can be detected by a commercially available assay kit. Understanding hormone patterns in animals gives researchers a better idea of best husbandry practices, including breeding in managed care.

Differential responses of prairie rodents to edge effects from recreational trails

Edge effects are a common phenomenon in which an ecological variable changes with respect to distance from a habitat edge. Recreational trails may constitute a habitat edge for prairie rodents because of high human presence, high predator presence, or limited shelter compared to the prairie core. Despite the prevalence of trails in conservation parcels, their effect on wildlife distribution remains largely unstudied. We examined the impacts of recreational trails on small mammal activity in the restored prairies of the Cowling Arboretum at Carleton College. The prairies were restored from 1995 to 2008 and now comprise a contiguous prairie block of approximately 155 ha. Over 2 consecutive summers, we used infrared motion-sensing cameras to record the relative amount of time rodents spend at baited stations placed at different distances from the trail. The results varied between taxa: voles (Microtus spp.) avoided trail edges whereas mouse (Cricetidae and Dipodidae) and thirteen-lined ground squirrel (Ictidomys tridecemlineatus) activity was unaffected by trail proximity. Trails may therefore have species-specific effects on small mammals, with potential consequences for the connectivity and distribution of populations.

CNGA3 acts as a cold sensor in hypothalamic neurons

Most mammals maintain their body temperature around 37°C, whereas in hibernators it can approach 0°C without triggering a thermogenic response. The remarkable plasticity of the thermoregulatory system allowed mammals to thrive in variable environmental conditions and occupy a wide range of geographical habitats, but the molecular basis of thermoregulation remains poorly understood. Here we leverage the thermoregulatory differences between mice and hibernating thirteen-lined ground squirrels (Ictidomys tridecemlineatus) to investigate the mechanism of cold sensitivity in the preoptic area (POA) of the hypothalamus, a critical thermoregulatory region. We report that, in comparison to squirrels, mice have a larger proportion of cold-sensitive neurons in the POA. We further show that mouse cold-sensitive neurons express the cyclic nucleotide-gated ion channel CNGA3, and that mouse, but not squirrel, CNGA3 is potentiated by cold. Our data reveal CNGA3 as a hypothalamic cold sensor and a molecular marker to interrogate the neuronal circuitry underlying thermoregulation.

Prevalence of zoonotic Bartonella among prairie rodents in Illinois

Bartonella is a genus of gram-negative bacteria that includes a variety of human and veterinary pathogens. These pathogens are transmitted from reservoirs to secondary hosts through the bite of arthropod vectors including lice and fleas. Once in the secondary host, the bacteria cause a variety of pathologies including cat-scratch disease, endocarditis, and myocarditis. Reservoirs of these bacteria are numerous and include several species of large mammals, mesocarnivores, and small mammals. Research on reservoirs of these bacteria has focused on western North America, Europe, and Asia, with little focus on the eastern and central United States. We assessed the prevalence of zoonotic Bartonella species among prairie-dwelling rodent species in the midwestern United States. Tissue samples (n = 700) were collected between 2015 and 2017 from five rodent species and screened for the presence of Bartonella DNA via PCR and sequencing of two loci using Bartonella-specific primers. Bartonella were prevalent among all five species, with 13-lined ground squirrels (Ictidomys tridecemlineatus) serving as a likely reservoir of the pathogen B. washoensis, and other rodents serving as reservoirs of the pathogens B. grahamii and B. vinsonii subsp. arupensis. These results demonstrate the value of studies of disease ecology in grassland systems, particularly in the context of habitat restoration and human–vector interactions.

The heart of a hibernator: EGFR and MAPK signaling in cardiac muscle during the hibernation of thirteen-lined ground squirrels, Ictidomys tridecemlineatus

Background Thirteen-lined ground squirrels (Ictidomys tridecemlineatus) experience dramatic changes in physiological and molecular parameters during winter hibernation. Notably, these animals experience reduced blood circulation during torpor, which can put numerous stresses on their hearts. The present study evaluates the role played by the epidermal growth factor receptor (EGFR) in signal transduction during hibernation at low body temperature to evaluate signaling mechanisms. By investigating the regulation of intracellular mitogen activated protein kinase (MAPK) pathway responses, anti-apoptosis signals, downstream transcription factors, and heat shock proteins in cardiac muscle we aim to determine the correlation between upstream tyrosine phosphorylation events and downstream outcomes. Methods Protein abundance of phosphorylated EGFR, MAPKs and downstream effector proteins were quantified using immunoblotting and Luminex® multiplex assays. Results Monitoring five time points over the torpor/arousal cycle, EGFR phosphorylation on T654, Y1068, Y1086 was found to increase significantly compared with euthermic control values particularly during the arousal process from torpor, whereas phosphorylation at Y1045 was reduced during torpor. Phosphorylation of intracellular MAPK targets (p-ERK 1/2, p-JNK, p-p38) also increased strongly during the early arousal stage with p-p38 levels also rising during prolonged torpor. However, of downstream MAPK effector kinases that were measured, only p-Elk-1 levels changed showing a decrease during interbout arousal (IA). Apoptosis markers revealed a strong reduction of the pro-apoptotic p-BAD protein during entrance into torpor that remained suppressed through torpor and IA. However, active caspase-9 protein rose strongly during IA. Levels of p-AKT were suppressed during the transition phases into and out of torpor. Of four heat shock proteins assessed, only HSP27 protein levels changed significantly (a 40% decrease) during torpor. Conclusion We show evidence of EGFR phosphorylation correlating to activation of MAPK signaling and downstream p-ELK1 suppression during hibernation. We also demonstrate a reduction in p-BAD mediated pro-apoptotic signaling during hibernation with active caspase-9 protein levels increasing only during IA. I. tridecemlineatus has natural mechanisms of tissue protection during hibernation that is largely due to cellular regulation through phosphorylation-mediated signaling cascade. We identify a possible link between EGFR and MAPK signaling via p-ERK, p-p38, and p-JNK in the cardiac muscle of these hibernating mammals that correlates with an apparent reduction in caspase-9 apoptotic signaling. This reveals a piece of the mechanism behind how these mammals are resilient to cardiac stresses during hibernation that would otherwise be damaging.