Development and evaluation of indirect enzyme-linked immunosorbent assays for the determination of immune response to multiple clostridial antigens in vaccinated captive bred southern white rhinoceros (Ceratotherium simum simum)

Background An overall increase in poaching of white rhinoceros results in captive breeding becoming a significant component of white rhinoceros conservation. However, this type of conservation comes with its own difficulties. When wildlife is captured, transported and/or confined to a boma environment, they are more predisposed to diseases caused by bacterial organisms such as spore forming Clostridium spp. A southern white rhinoceros (Ceratotherium simum simum) population on a captive bred farm was suspected to be affected by Clostridium infections. These endangered animals were apparently exposed to Clostridium spp., in the conservation area previously used for cattle farming. The rhinoceros population on the breeding operation property was vaccinated with a multi-component clostridial vaccine registered for use in cattle. Multiple indirect enzyme-linked immunosorbent assays (iELISAs) were developed in order to evaluate the serum antibody titres of these vaccinated animals. In evaluating vaccine efficacy, the gold standard mouse neutralization test (MNT) was not available and therefore iELISAs were developed for the detection of serum antibodies to C. perfringens type A (alpha toxin), C. chauvoei (whole cell), C. novyi (alpha toxin), C. septicum (alpha toxin) and C. sordellii (lethal toxin) in the white rhinoceros population using international reference sera of equine origin. Antibody titres against each clostridial antigen was evaluated in the vaccinated white rhinoceros population (n = 75). Analytical specificity showed slight cross-reactions for C. chauvoei and C. perfringens type A with the other antigens. Individual assay cut-off values were calculated with 95% confidence. Coefficient of variance (CV) values for both the international reference sera and in-house control sera across all the antigens were well below 16%, indicating good assay repeatability. This convenient and fast assay is suitable for monitoring humoral immune responses to clostridial antigens in vaccinated white rhinoceroses. Results Checkerboard titrations indicated optimal antigen and antibody concentrations to be used for each respective iELISA developed. Each titration set of the respective international reference and in-house control sera showed good repeatability with low standard deviations and coefficient of variance values calculated between repeats for each antigen. Individual assays proved repeatable and showed good analytical sensitivity and specificity. Conclusions The developed iELISAs are able to evaluate antibody profiles of phospholipase C, C. chauvoei whole cells, TcnA, ATX, TcsL in white rhinoceros serum using international reference sera.

Proximity-based vocal networks reveal social relationships in the Southern white rhinoceros

Vocal communication networks can be linked to social behaviour, allowing a deeper understanding of social relationships among individuals. For this purpose, the description of vocal dyads is fundamental. In group-living species, this identification is based on behavioural indicators which require a high level of reactivity during social interactions. In the present study, we alternatively established a proximity-based approach to investigate whether sex-specific differences in vocal communication reflect social behaviour in a species with rather loose social associations and low levels of reactivity: the Southern white rhinoceros (Ceratotherium simum simum). We performed audio- and video recordings of 30 captive animals from seven groups. Vocal networks for the four most common call types were constructed by considering conspecifics at close distance (≤ 1 body length) to the sender as potential receivers. The analysis of the resulting unidirectional structures showed that not only the sex of the sender but also the sex of the potential receiver, the quality of social interactions (affiliative or agonistic) as well as association strength predict the intensity of vocal interactions between group members. Thus, a proximity-based approach can be used to construct vocal networks providing information about the social relationships of conspecifics—even in species with loose social associations where behavioural indicators are limited.