Fish Shellfish Immunol. 2017 Apr;63:424-437. doi: 10.1016/j.fsi.2017.02.039.

Who needs the hotspot? The effect of temperature on the fish host immune response to Tetracapsuloides bryosalmonae the causative agent of proliferative kidney disease.

Bailey C1, Segner H1, Casanova-Nakayama A1, Wahli T1,2.

1 University of Berne, Vetsuisse Faculty, Centre for Fish and Wildlife Health, Länggassstrasse 122, CH-3012 Berne, Switzerland.
2 Electronic address: thomas.wahli@vetsuisse.unibe.ch.

Abstract

Proliferative kidney disease (PKD) of salmonids, caused by Tetracapsuloides bryosalmonae may lead to high mortalities at elevated water temperatures. However, it has not yet been investigated how temperature affects the fish host immune response to T. bryosalmonae. We exposed YOY (young of the year) rainbow trout (Oncorhynchus mykiss) to T. bryosalmonae at two temperatures (12 °C and 15 °C) that reflect a realistic environmental scenario and could occur in the natural habitat of salmonids. We followed the development of the parasite, host pathology and immune response over seven weeks. We evaluated the composition and kinetics of the leukocytes and their major subgroups in the anterior and posterior kidney. We measured immune gene expression profiles associated with cell lineages and functional pathways in the anterior and posterior kidney. At 12 °C, both infection prevalence and pathogen load were markedly lower. While the immune response was characterized by subtle changes, mainly an increased amount of lymphocytes present in the kidney, elevated expression of Th1-like signature cytokines and strong upregulation of the natural killer cell enhancement factor, NKEF at week 6 P.E. At 15 °C the infection prevalence and pathogen burden were ominously greater. While the immune response as the disease progressed was associated with a Th2-like switch at week 6 P.E and a prominent B cell response, evidenced at the tissue, cell and transcript level. Our results highlight how a subtle, environmentally relevant difference in temperature resulted in diverse outcomes in terms of the immune response strategy, altering the type of interaction between a host and a parasite.

KEYWORDS:

Immune response; PKD; Proliferative kidney disease; Rainbow trout; Resistance; Temperature; Tetracapsuloides bryosalmonae; Th1; Th2; Tolerance

PMID: 28238860

 

Supplement

Background

Infectious diseases can cause rapid population declines or even species extinctions. Pathogens can be sensitive to environmental change, creating synergisms that could affect biodiversity [1]. Environmental change can directly via physiological and immunological processes or indirectly via ecological mechanisms modulate host-parasite interactions and disease outcomes. Pertinent to this is PKD (Proliferative Kidney Disease) of salmonids in which induced mortality has been associated with elevated temperatures [2].  However, it is unknown how environmental variability may influence host immune processes during infection.

PKD studies have demonstrated that at 12 °C, the host develops a version of the disease with fewer clinical signs and lower mortality [3]. This may represent a stable co-evolved host parasite system. However, as hosts and their parasites have an intimate and antagonistic relationship, the effect of temperature may influence the host-pathogen relationship. The outcome of this relationship may derive from two noted strategies: tolerance which is defined as the ability to limit the health impact of a given pathogen burden, or resistance which is defined as the ability to limit pathogen burden [4]. The immune strategy used by the host will drive the impact of the disease on the fitness of the individual. Whether the immune responses of salmonids could be assigned to one of the two strategies, and whether this changes with temperature, has not yet been elucidated.

The following research questions were addressed in this study 1) What is the host immune response in the coevolved model system and how does this affect the outcome of the disease? 2) How does increased temperature modulate the host immune response in comparison to the coevolved model system and how does this affect the outcome of the disease? , and 3) What are the drivers for the difference in the immune response at each temperature? While we pay a great deal of attention to the cellular and molecular immune repertoire of the fish host, we will also take an evolutionary perspective to identify if the host at either temperature uses a noted strategy to increase its fitness. Granting this, it cannot be ruled out that the immune response is dynamic and not static and may be temporally dependent. Hence, the immune response at the warmer temperature may be faster than the cooler temperature, and a comparison of the immune response at a sequential time point may trigger false conclusions. In addition, the host immune response may be influenced via an indirect effect of temperature on parasite intensity. Therefore, to bring a sense of clarity to the study, we attempted to elucidate potential drivers of the immune response.

 

Methodology  

Experimentally the study aims were achieved by exposing YOY rainbow trout to an equal amount of T. bryosalmonae at two temperatures, 12 °C and 15 °C that reflect a realistic environmental scenario and could occur in the natural habitat of salmonids. The 12 °C infection group represented the coevolved model system, whereas 15 °C represented the increased temperature infection group. We followed the development of the parasite, host pathology and immune response over seven weeks. We evaluated the composition and kinetics of the leukocytes and their major subgroups in the anterior and posterior kidney. In addition, we measured immune gene expression profiles associated with cellular lineages and functional pathways in the anterior and posterior kidney.

 

Outcome

What is the host immune response in the coevolved model system and how does this affect the outcome of the disease? 

At 12 °C the fish host immune response strategy per se is to tolerate the parasite and limit the harm caused by a given burden. This is evidenced here by the subtle fine-tuning of the immune response, such as the influx of lymphocytes, and expression of Th1 signature cytokines and NKEF and IL-10 and their ensuing effect on a stable parasite load (Fig. 1 and Fig. 2). Granting this, as the parasite could not achieve 100% prevalence; equal consideration must be paid to the effect of temperature on virulence and on pathogenicity as well as the capabilities of the immune response which equally contribute to the stable, yet tolerable, system we have observed at the cooler temperature.

 

How does increased temperature modulate the host immune response in comparison to the coevolved model system and how does this affect the outcome of the disease?

At the increased temperature of 15 °C infection prevalence and pathogen burden were ominously greater resulting in clinical PKD. The immune response as the disease progressed was associated with a Th2 like switch and a prominent B cell response, evidenced at the tissue, cell and transcript level (Fig. 1 and Fig. 2) . Our results highlight how a small change in temperature resulted in diverse outcomes in terms of the immune response strategy, altering the type of interaction between a host and a parasite. Hence, at 15 °C it appears that the fish immune response strategy is to limit the parasite burden; evolutionary theory deems this strategy resistance, evidenced by the over active immune response trying to combat the growing pathogen burden. But as parasite intensity increases the immune response must as well, which may provoke more pathogenic traits from the parasite ending in a severe infection. The high expression levels of genes in this system seem to suggest some sort of immunological molecular warfare between the host and parasite. Furthermore, the grossly inflamed tissue at 15 °C experienced by hosts may not only result from the direct effects of parasite development but also from such an overzealous immune response inducing a chronic PKD immunopathology.

 

 

Figure. 1.  Comparative summary of mean relative fold change at week 6 P.E (post exposure) of Th1 and Th2-like cytokines measured in the AK (anterior kidney) and PK (posterior kidney) of infected fish at different temperatures.  Relative fold change was initially normalized to rainbow trout EF-1α and subsequently expressed as fold change relative to expression levels of control fish.

Figure. 2.  Comparative summary of mean relative fold change at week 6 P.E (post exposure) of genes associated with different cellular lineages measured in the AK (anterior kidney) and PK (posterior kidney) of infected fish at different temperatures.  Relative fold change was initially normalized to rainbow trout EF-1α and subsequently expressed as fold change relative to expression levels of control fish.

 

What are the drivers for the difference in the immune response at each temperature?

The immune response of the fish host to the parasite may be directly driven by the temperature difference between 12 °C and 15 °C, but also indirectly driven through the temperature effect on the parasite intensity. Coincidentally, our results show a correlation between severity of disease and host immune response at the two temperatures. However, we also clearly showed the influence of temperature on parasite intensity. This greater parasite intensity at 15 °C could be a factor contributing to the different immune response at 15 °C, in particular since the parasite intensity at 15 °C compared to that at 12 °C was clearly higher than to be expected from the Q10 coefficient values. Still, different parasite intensities and accelerated metabolic rates at the 15 °C compared to 12 °C are not sufficient as explanation, mainly because the differences of the immune responses between the two temperature groups is not simply a quantitative one but a qualitative one. The fish changes its immune defense strategy to deal with the parasite: while at 12 °C the fish immune  strategy appears to tolerate the parasite and limit the harm caused by a given burden, at 15° C it seems to mount a resistance response to limit the parasite burden as explained above.  The differences we observed in parasite intensity did not clearly explain the differences we observed in the immune response. To substantiate this, we compared the immune response at time points when the parasite intensity was similar in the infection groups, such as at 12 ° C weeks 7 P.E (35442) and at 15 °C week 4 P.E (48285). From this analysis, we can clearly show differences in the immune response when the parasite intensity is similar. For example, at 15 °C there was a more pronounced pathological reaction in the tissue, increased amount of lymphocytes in the PK and increased mRNA levels of IgM sec (110.4 vs 2.8) and Pax5 (13.8 vs 3.9). This data demonstrates that the immune response is not just an indirect effect of parasite intensity but also temperature. Furthermore, the differences we observed were not temporally dependent, as there was not a faster immune response at the warmer temperature or a slower response at the cooler temperature. To illustrate this, we adhere to the immune response of 12 °C infection fish at week 7 P.E against the immune response of 15 °C infection fish at week 2 P.E. While there were parallels such as the upregulation of Th1-like cytokines, there were still major divergences. These included elevated expression patterns of Th2-like cytokines and of B cell markers, which were all expressed at 15 °C but not at 12 °C. strategy appears to tolerate the parasite and limit the harm caused by a given burden, at 15° C it seems to mount a resistance response to limit the parasite burden as explained above (Subsection General Discussion: 2.2 and 2.3).  The differences we observed in parasite intensity did not clearly explain the differences we observed in the immune response. To substantiate this, we compared the immune response at time points when the parasite intensity was similar in the infection groups, such as at 12 ° C weeks 7 P.E (35442) and at 15 °C week 4 P.E (48285). From this analysis, we can clearly show differences in the immune response when the parasite intensity is similar. For example, at 15 °C there was a more pronounced pathological reaction in the tissue, increased amount of lymphocytes in the PK and increased mRNA levels of IgM sec (110.4 vs 2.8) and Pax5 (13.8 vs 3.9). This data demonstrates that the immune response is not just an indirect effect of parasite intensity but also temperature. Furthermore, the differences we observed were not temporally dependant, as there was not a faster immune response at the warmer temperature or a slower response at the cooler temperature. To illustrate this, we adhere to the immune response of 12 °C infection fish at week 7 P.E against the immune response of 15 °C infection fish at week 2 P.E. While there were parallels such as the upregulation of Th1-like cytokines, there were still major divergences. These included elevated expression patterns of Th2-like cytokines and of B cell markers, which were all expressed at 15 °C but not at 12 °C.

 

Take home message

We detailed how an increase of only 3 °C had an impact on parasite proliferation and on the biological and biochemical processes of the host, upsetting the delicate co-evolved host-pathogen system. Whereas at 12 °C the hosts immunocompetence appears not to be negatively impacted and the host even tolerates a limited pathogen burden. What is clear from the work presented here is that environmental change acts at the individual, host–parasite interaction and ecological level. Changing environmental conditions may eventually alter the approach to optimal host response if defense mechanisms are regulated to maximize net fitness in the context of costs, ecological influences, and constraints [5]. Although we are only at an early stage in the projected trends of global warming, such ecological responses and their effects on not just the host-pathogen system, but on the entire ecosystem are already visible and of great conservation significance.

 

References

[1] C.D. Harvell, C.E. Mitchell, J.R. Ward, S. Altizer, A.P. Dobson, R.S. Ostfeld, M.D. Samuel, Climate warming and disease risks for terrestrial and marine biota, Science 296(5576) (2002) 2158-2162.

[2] B. Okamura, H. Harikainen, H. Schmidt-Posthaus, T. Wahli, Life cycle complexity, environmental change and the emerging status of salmonid proliferative kidney disease, Freshwater Biology 56(4) (2011) 735-753.

[3] B. K., S. H., B. R., S.-P. H., W. T., Proliferative kidney disease (PKD) of rainbow trout: temperature- and time- related changes of Tetracapsuloides bryosalmonae DNA in kidney, J. of Parasitology 136 (2009) 615-625.

[4] D.S. Schneider, J.S. Ayres, Two ways to survive infection: what resistance and tolerance can teach us about treating infectious diseases, Nature Reviews Immunology 8(11) (2008) 889-895.

[5] M. van Boven, F.J. Weissing, The evolutionary economics of immunity, The American Naturalist 163(2) (2004) 277-294.