Brianna R. Beechler

Beyond phytohaemagglutinin: assessing vertebrate immune function across ecological contexts

1. Over the past decade, there has been a substantial increase in interest in the immune system and the role it plays in the regulation of disease susceptibility, giving rise to the field of eco-immunology. 2. Eco-immunology aims to understand changes in host immune responses in the broader framework of an organisms evolutionary, ecological and life-history contexts. 3. The immune system, however, is complex and multifaceted and can be intimidating for the nonimmunologist interested in incorporating immunological questions into their research. Which immune responses should one measure and what is the biological significance of these measures? 4. The focus of this review is to describe a wide range of eco-immunology techniques, from the simple to the sophisticated, with the goal of providing researchers with a range of options to consider incorporating in their own research programs. 5. These techniques were chosen because they provide relatively straightforward, biologically meaningful assessments of immune function, many of which can be performed across a range of ecological contexts (i.e. field vs. laboratory) and in a wide range of vertebrate animals without relying on species-specific reagents. 6. By incorporating assessments of immune function into their specific research questions, animal ecologists will gain a more comprehensive understanding of organism-environment interactions.

Inter-epidemic and between-season persistence of rift valley fever: vertical transmission or cryptic cycling?

Rift Valley fever (RVF) is an emerging zoonotic mosquito-borne infectious disease that has been identified as a risk for spread to other continents and can cause mass livestock mortality. In equatorial Africa, outbreaks of RVF are associated with high rainfall, when vector populations are at their highest. It is, however, unclear how RVF virus persists during the inter-epidemic periods and between seasons. Understanding inter-epidemic persistence as well as the role of vectors and hosts is paramount to creating effective management programmes for RVF control. We created a mathematical model for the spread of RVF and used the model to explore different scenarios of persistence including vertical transmission and alternate wildlife hosts, with a case study on buffalo in Kruger National Park, South Africa. Our results suggest that RVF persistence is a delicate balance between numerous species of susceptible hosts, mosquito species, vertical transmission and environmental stochasticity. Further investigations should not focus on a single species, but should instead consider a myriad of susceptible host species when seeking to understand disease dynamics.

Innate immunity in free-ranging African buffalo (Syncerus caffer): associations with parasite infection and white blood cell counts.

Mammalian immunology has been studied in great detail in laboratory animals, but few of the tools and less of the insight derived from these studies have been placed in the context of natural, outbred wildlife populations subject to variable environments. We investigated patterns of innate immunity in free-ranging African buffalo in relation to host traits (age, reproductive status, body condition, white blood cell counts) and disease status (bovine tuberculosis [BTB], gastrointestinal nematodes, coccidia, ticks). We evaluated and used an in vitro assay measuring bactericidal competence of blood to assess a component of innate immunity in 200 female buffalo captured at Kruger National Park, South Africa, in June/July and October 2008. Animals with BTB had higher bactericidal competence of blood. Animals with higher neutrophil counts had higher bactericidal competence, whereas animals with lower lymphocyte counts had higher bactericidal competence. This pattern was driven by animals captured at the end of the dry season (October) and may be evidence of immune polarization, whereby individuals are unable to upregulate multiple components of immunity simultaneously. Bactericidal competence did not vary with host pregnancy status, body condition, age, lactation, tick infestation, nematode egg count, or coccidia oocyst count. Overall, we demonstrate that the bactericidal competence assay is practical and informative for field-based studies in wild bovids. Our results also show a correlation between bactericidal competence and bovine tuberculosis infection and reveal possible functional polarizations between different types of immune response in a free-ranging mammal.

EVALUATION OF HEMATOLOGIC VALUES IN FREE-RANGING AFRICAN BUFFALO (SYNCERUS CAFFER)

As part of a large-scale disease screening program, blood samples were collected from 534 African buffalo (Syncerus caffer) in South Africas Hluhluwe-iMfolozi Park in October 2005 and May 2006 to establish age- and sex-specific reference intervals for erythrogram and leukogram values. Sixty-seven of the animals were positive for bovine tuberculosis (TB), allowing for comparisons between TB-positive and TB-negative groups. Positive animals had basopenia and slight lymphopenia compared to TB-negative animals. Blood values were compared to those reported for captive African buffalo, American bison (Bos bison), and cattle (Bos taurus). The free-ranging buffalo sampled in this study had higher white blood cell counts than captive buffalo, and this difference was driven by lymphocytes. Free-ranging buffalo also had higher red blood cell counts, mean corpuscular hemoglobin concentration (MCHC), white blood cell counts, neutrophils and lymphocytes, and lower mean corpuscular volume (MCV) than cattle. Demographic and environmental factors strongly affected hematologic values in the study population. Older animals had significantly higher hemoglobin, hematocrit, MCV, and mean corpuscular hemoglobin (MCH), while younger animals had a higher red blood cell count, red cell distribution width (RDW), and white blood cell count, which was due to lymphocytes and basophils. Females had a higher hemoglobin concentration, hematocrit, MCV, MCH, and basophils than males. At the end of the wet season, hemoglobin, red blood cell count, hematocrit, MCHC, RDW, white blood cell count, and neutrophils were all significantly higher, while basophils and MCV were lower, than at the end of the dry season. Our results emphasize the need to use species-specific data when interpreting hematologic values and point to important differences in hematology between captive and free-ranging animals of the same species. Strong variability in hematologic values with animal age and sex, season, and herd affiliation indicates that “normal” hematologic values in wild animals vary throughout their lives and subject to fluctuating environmental conditions.

Enemies and Turncoats: Bovine tuberculosis exposes pathogenic potential of Rift Valley fever virus in a common host, African Buffalo (Syncerus caffer)

The ubiquity and importance of parasite co-infections in populations of free-living animals is beginning to be recognized, but few studies have demonstrated differential fitness effects of single infection versus co-infection in free-living populations. We investigated interactions between the emerging bacterial disease bovine tuberculosis (BTB) and the previously existing viral disease Rift Valley fever (RVF) in a competent reservoir host, African buffalo, combining data from a natural outbreak of RVF in captive buffalo at a buffalo breeding facility in 2008 with data collected from a neighbouring free-living herd of African buffalo in Kruger National Park. RVF infection was twice as likely in individual BTB+ buffalo as in BTB− buffalo, which, according to a mathematical model, may increase RVF outbreak size at the population level. In addition, co-infection was associated with a far higher rate of fetal abortion than other infection states. Immune interactions between BTB and RVF may underlie both of these interactions, since animals with BTB had decreased innate immunity and increased pro-inflammatory immune responses. This study is one of the first to demonstrate how the consequences of emerging infections extend beyond direct effects on host health, potentially altering the dynamics and fitness effects of infectious diseases that had previously existed in the ecosystem on free-ranging wildlife populations.

Molecular characterisation of Anaplasma species from African buffalo (Syncerus caffer) in Kruger National Park, South Africa

Bovine anaplasmosis is a tick-borne disease, mainly caused by Anaplasma marginale and A. centrale and is distributed in tropical and sub-tropical areas. This study aimed to characterise A. marginale and A. centrale from African buffaloes in Kruger National Park (KNP), South Africa, using the DNA sequences of the genes coding for major surface protein (msp1β) and heat shock protein (groEL), respectively. A total of 747 blood samples were collected from February 2014 to August 2016 from African buffaloes kept in KNP, and DNAs were tested using a molecular-phylogenetic approach. Out of 747 samples tested, 129 (17.3%) and 98 (13.1%) were positive for single infection with A. marginale and A. centrale, respectively; whereas 113 (15.1%) were positive for both Anaplasma spp. Pairwise difference of 1.6-8.5% was observed in msp1β sequences of A. marginale whereas that was only 0.3-2.4% for groEL sequences of A. centrale. Separate phylogenetic analyses of msp1β and groEL sequences of A. marginale and A. centrale, respectively, revealed that sequences of Anaplasma spp. from African buffaloes were unique and they grouped separately when compared with previously published sequences of both species. This is the first study to characterise A. marginale and A. centrale from African buffalo using species specific molecular markers. This study will pave the way for future studies to assess genetic variation among Anaplasma spp. from wild ruminants using molecular markers that are better at differentiating between species and strains than the more commonly used 16S rRNA gene, and help to undertake health and fitness studies and host-parasite dynamics using quantitative molecular tools.

Within guild co-infections influence parasite community membership : a longitudinal study in African Buffalo

Experimental studies in laboratory settings have demonstrated a critical role of parasite interactions in shaping parasite communities. The sum of these interactions can produce diverse effects on individual hosts as well as influence disease emergence and persistence at the population level. A predictive framework for the effects of parasite interactions in the wild remains elusive, largely because of limited longitudinal or experimental data on parasite communities of free-ranging hosts. This 4-year study followed a community of haemoparasites in free-ranging African buffalo (Syncerus caffer). We detected infection by 11 haemoparasite species using PCR-based diagnostic techniques, and analyzed drivers of infection patterns using generalized linear mixed models to understand the role of host characteristics and season on infection likelihood. We tested for (i) effects of co-infection by other haemoparasites (within guild) and (ii) effects of parasites infecting different tissue types (across guild). We found that within guild co-infections were the strongest predictors of haemoparasite infections in the buffalo; but that seasonal and host characteristics also had important effects. In contrast, the evidence for across-guild effects of parasites utilizing different tissue on haemoparasite infection was weak. These results provide a nuanced view of the role of co-infections in determining haemoparasite infection patterns in free living mammalian hosts. Our findings suggest a role for interactions among parasites infecting a single tissue type in determining infection patterns.

Beyond mice and men: environmental change, immunity and infections in wild ungulates

In the face of rapid environmental change, anticipating shifts in microparasite and macroparasite dynamics, including emergence events, is an enormous challenge. We argue that immunological studies in natural populations are pivotal to meeting this challenge: many components of environmental change – shifts in biotic assemblages, altered climate patterns and reduced environmental predictability – may affect host immunity. We suggest that wild ungulates can serve as model systems aiding the discovery of immunological mechanisms that link environmental change with parasite transmission dynamics. Our review of eco‐immunological studies in wild ungulates reveals progress in understanding how co‐infections affect immunity and parasite transmission and how environmental and genetic factors interact to shape immunity. Changes in bioavailability of micronutrients have been linked to immunity and health in wild ungulates. Although physiological stress in response to environmental change has been assessed, downstream effects on immunity have not been studied. Moreover, the taxonomic range of ungulates studied is limited to bovids (bighorn sheep, Soay sheep, chamois, musk oxen, bison, African buffalo) and a few cervids (red deer, black‐tailed deer). We discuss areas where future studies in ungulates could lead to significant contributions in understanding the patterns of immunity and infection in natural populations and across species.

Serum biochemistry panels in African buffalo: Defining reference intervals and assessing variability across season, age and sex

Serum biochemical parameters can be utilized to evaluate the physiological status of an animal, and relate it to the animal’s health. In order to accurately interpret individual animal biochemical results, species-specific reference intervals (RI) must be established. Reference intervals for biochemical parameters differ between species, and physiological differences including reproductive status, nutritional resource availability, disease status, and age affect parameters within the same species. The objectives of this study were to (1) establish RI for biochemical parameters in managed African buffalo (Syncerus caffer), (2) assess the effects of age, sex, pregnancy, and season on serum biochemistry values, and (3) compare serum biochemistry values from a managed herd to a free-ranging buffalo herd and to values previously published for captive (zoo) buffalo. Season profoundly affected all biochemistry parameters, possibly due to changes in nutrition and disease exposure. Age also affected all biochemical parameters except gamma glutamyl transferase and magnesium, consistent with patterns seen in cattle. Sex and reproductive status had no detectable effects on the parameters that were measured. The biochemical profiles of managed buffalo were distinct from those observed in the free-ranging herd and captive buffalo. Biochemical differences between buffalo from captive, managed, and free-ranging populations may be related to nutritional restriction or lack of predation in the context of management or captivity. The reference intervals provided in this study, in addition to the seasonal and age-related patterns observed, provide a foundation for health investigations that may inform management strategies in this ecologically and economically important species.

Host immunity, nutrition and coinfection alter longitudinal infection patterns of schistosomes in a free ranging African buffalo population

Schistosomes are trematode parasites of global importance, causing infections in millions of people, livestock, and wildlife. Most studies on schistosomiasis, involve human subjects; as such, there is a paucity of longitudinal studies investigating parasite dynamics in the absence of intervention. As a consequence, despite decades of research on schistosomiasis, our understanding of its ecology in natural host populations is centered around how environmental exposure and acquired immunity influence acquisition of parasites, while very little is known about the influence of host physiology, coinfection and clearance in the absence of drug treatment. We used a 4-year study in free-ranging African buffalo to investigate natural schistosome dynamics. We asked (i) what are the spatial and temporal patterns of schistosome infections; (ii) how do parasite burdens vary over time within individual hosts; and (iii) what host factors (immunological, physiological, co-infection) and environmental factors (season, location) explain patterns of schistosome acquisition and loss in buffalo? Schistosome infections were common among buffalo. Microgeographic structure explained some variation in parasite burdens among hosts, indicating transmission hotspots. Overall, parasite burdens ratcheted up over time; however, gains in schistosome abundance in the dry season were partially offset by losses in the wet season, with some hosts demonstrating complete clearance of infection. Variation among buffalo in schistosome loss was associated with immunologic and nutritional factors, as well as co-infection by the gastrointestinal helminth Cooperia fuelleborni. Our results demonstrate that schistosome infections are surprisingly dynamic in a free-living mammalian host population, and point to a role for host factors in driving variation in parasite clearance, but not parasite acquisition which is driven by seasonal changes and spatial habitat utilization. Our study illustrates the power of longitudinal studies for discovering mechanisms underlying parasite dynamics in individual animals and populations.