The adaptive immune response to Trichuris in wild versus laboratory mice: An established model system in context.

Laboratory model organisms have provided a window into how the immune system functions. An increasing body of evidence, however, suggests that the immune responses of naive laboratory animals may differ substantially to those of their wild counterparts. Past exposure, environmental challenges and physiological condition may all impact on immune responsiveness. Chronic infections of soil-transmitted helminths, which we define as establishment of adult, fecund worms, impose significant health burdens on humans, livestock and wildlife, with limited treatment success. In laboratory mice, Th1 versus Th2 immune polarisation is the major determinant of helminth infection outcome. Here we compared antigen-specific immune responses to the soil-transmitted whipworm Trichuris muris between controlled laboratory and wild free-ranging populations of house mice (Mus musculus domesticus). Wild mice harbouring chronic, low-level infections produced lower levels of cytokines in response to Trichuris antigen than laboratory-housed C57BL/6 mice. Wild mouse effector/memory CD4+ T cell phenotype reflected the antigen-specific cytokine response across the Th1/Th2 spectrum. Increasing egg shedding was associated with body condition loss. However, local Trichuris-specific Th1/Th2 balance was positively associated with worm burden only in older wild mice. Thus, although the fundamental relationships between the CD4+ T helper cell response and resistance to T. muris infection are similar in both laboratory and wild M. m. domesticus, there are quantitative differences and age-specific effects that are analogous to human immune responses. These context-dependent immune responses demonstrate the fundamental importance of understanding the differences between model and natural systems for translating mechanistic models to 'real world' immune function.

Salivary testosterone changes during oral glucose tolerance tests in overweight and obese men - postprandial or circadian variation?

BACKGROUND: Serum total testosterone (T) decreases postprandially. Postprandial salivary testosterone (SalT) responses, however, have not been studied. We report on the effect of glucose ingestion on fasting SalT concentrations. OBJECTIVE: To investigate the effect of oral glucose ingestion on fasting SalT. METHODS: Salivary and blood samples were collected between 09.00 and 09.30 and two hours after a 75g oral glucose load in 32 men with mean (standard deviation) age of 52 (5.7) years and body mass index of 32.6 (5.56) kg/m2. Free T and bioavailable testosterone (BAT) were calculated using the Vermeulen equation. RESULTS: Two hours following oral glucose, there was a decrease in fasting mean (standard deviation) SalT [178.2 (56.6) vs 146.0 (42.2) pmol/L; p = 0.0003], serum cortisol [332 (105.0) vs 239 (75.3) nmol/L; p = <0.0001], prolactin [193 (75.0) vs 127 (55.9) mIU/L; p = <0.0001] and TSH [1.60 (0.801) vs 1.16 (0.584) mIU/L; p = <0.0001]. Plasma glucose increased [6.2 (0.72) vs 8.1 (3.71) mmol/L; p = 0.0029]. Serum total T, SHBG, albumin, Free T, BAT, gonadotrophins and FT4 remained unchanged. CONCLUSIONS: SalT decreased postprandially. A concomitant decrease in serum cortisol, prolactin and TSH reflecting diurnal variation offers an alternative explanation for the decrease in SalT independent of food consumption. Further studies are required to determine whether morning temporal changes in SalT are related to food consumption or circadian rhythm or both.

Sex drives colonic mucin sialylation in wild mice.

Mucin protein glycosylation is important in determining biological properties of mucus gels, which form protective barriers at mucosal surfaces of the body such as the intestine. Ecological factors including: age, sex, and diet can change mucus barrier properties by modulating mucin glycosylation. However, as our understanding stems from controlled laboratory studies in house mice, the combined influence of ecological factors on mucin glycosylation in real-world contexts remains limited. In this study, we used histological staining with 'Alcian Blue, Periodic Acid, Schiff's' and 'High-Iron diamine' to assess the acidic nature of mucins stored within goblet cells of the intestine, in a wild mouse population (Mus musculus). Using statistical models, we identified sex as among the most influential ecological factors determining the acidity of intestinal mucin glycans in wild mice. Our data from wild mice and experiments using laboratory mice suggest estrogen signalling associates with an increase in the relative abundance of sialylated mucins. Thus, estrogen signalling may underpin sex differences observed in the colonic mucus of wild and laboratory mice. These findings highlight the significant influence of ecological parameters on mucosal barrier sites and the complementary role of wild populations in augmenting standard laboratory studies in the advancement of mucus biology.

Understanding conceptus-maternal interactions: what tools do we need to develop?

Communication between the maternal endometrium and developing embryo/conceptus is critical to support successful pregnancy to term. Studying the peri-implantation period of pregnancy is critical as this is when most pregnancy loss occurs in cattle. Our current understanding of these interactions is limited, due to the lack of appropriate in vitro models to assess these interactions. The endometrium is a complex and heterogeneous tissue that is regulated in a transcriptional and translational manner throughout the oestrous cycle. While there are in vitro models to study endometrial function, they are static and 2D in nature or explant models and are limited in how well they recapitulate the in vivo endometrium. Recent developments in organoid systems, microfluidic approaches, extracellular matrix biology, and in silico approaches provide a new opportunity to develop in vitro systems that better model the in vivo scenario. This will allow us to investigate in a more high-throughput manner the fundamental molecular interactions that are required for successful pregnancy in cattle.

Discrete patterns of microbiome variability across timescales in a wild rodent population.

Mammalian gastrointestinal microbiomes are highly variable, both within individuals and across populations, with changes linked to time and ageing being widely reported. Discerning patterns of change in wild mammal populations can therefore prove challenging. We used high-throughput community sequencing methods to characterise the microbiome of wild field voles (Microtus agrestis) from faecal samples collected across 12 live-trapping field sessions, and then at cull. Changes in α- and ß-diversity were modelled over three timescales. Short-term differences (following 1-2 days captivity) were analysed between capture and cull, to ascertain the degree to which the microbiome can change following a rapid change in environment. Medium-term changes were measured between successive trapping sessions (12-16 days apart), and long-term changes between the first and final capture of an individual (from 24 to 129 days). The short period between capture and cull was characterised by a marked loss of species richness, while over medium and long-term in the field, richness slightly increased. Changes across both short and long timescales indicated shifts from a Firmicutes-dominant to a Bacteroidetes-dominant microbiome. Dramatic changes following captivity indicate that changes in microbiome diversity can be rapid, following a change of environment (food sources, temperature, lighting etc.). Medium- and long-term patterns of change indicate an accrual of gut bacteria associated with ageing, with these new bacteria being predominately represented by Bacteroidetes. While the patterns of change observed are unlikely to be universal to wild mammal populations, the potential for analogous shifts across timescales should be considered whenever studying wild animal microbiomes. This is especially true if studies involve animal captivity, as there are potential ramifications both for animal health, and the validity of the data itself as a reflection of a 'natural' state of an animal.

The wild mouse bone marrow has a unique myeloid and lymphoid composition and phenotype.

The murine bone marrow has a central role in immune function and health as the primary source of leukocytes in adult mice. Laboratory mice provide a human-homologous, genetically manipulable and reproducible model that has enabled an immeasurable volume of high-quality immunological research. However, recent research has questioned the translatability of laboratory mouse research into humans and proposed that the exposure of mice to their wild and natural environment may hold the key to further immunological breakthroughs. To date, there have been no studies providing an in-depth cellular analysis of the wild mouse bone marrow. This study utilized wild mice from an isolated island population (Isle of May, Scotland, UK) and performed flow cytometric and histological analysis to characterize the myeloid, lymphoid, hematopoietic progenitor, and adipocyte compartments within the wild mouse bone marrow. We find that, compared to laboratory mouse bone marrow, the wild mouse bone marrow differs in every cell type assessed. Some of the major distinctions include; a smaller B cell compartment with an enriched presence of plasma cells, increased proportions of KLRG1+ CD8+ T cells, diminished CD11b expression in the myeloid lineage and a five-fold enlargement of the eosinophil compartment. We conclude that the wild mouse bone marrow is dramatically distinct from its laboratory counterparts, with multiple phenotypes that to our knowledge have never been observed in laboratory models. Further research into these unique features may uncover novel immunological mechanisms and grant a greater understanding of the role of the immune system in a natural setting.

The Diagnosis of Normocalcaemic Hyperparathyroidism is Strikingly Dissimilar Using Different Commercial Laboratory Assays.

We assessed the impact of intact parathyroid hormone (iPTH) and adjusted calcium analyses on Abbott, Roche and Siemens analytical platforms in the diagnosis of normocalcaemic primary hyperparathyroidism (NCPHPT). These assays are used by over 85% of clinical laboratories in the UK. Over five months, consecutive serum samples from outpatients with NCPHPT in the laboratory with Abbott assays were identified, aliquoted and stored at -80°C. Frozen aliquots were transported monthly to the other two laboratories. After thawing, samples were mixed and analysed immediately for calcium, albumin and iPTH in the laboratories with Abbott, Roche and Siemens analytical platforms. Adjusted calcium was calculated using the equation used in the respective laboratory. Diagnostic concordance of iPTH and adjusted calcium were assessed using manufacturer-provided assay-specific reference intervals and the pathology harmony reference interval respectively. Fifty-five patients with NCPHPT were identified using Abbott assays. Of these, 16 (29.1%) and 11 (20.0%) had NCPHPT, 9 (16.4%) and 13 (23.6%) had hypercalcaemic primary hyperparathyroidism, and 30 (54.6%) and 31 (56.4%) patients had normal results when analysed in laboratories with Roche and Siemens assays, respectively. The diagnosis of NCPHPT was strikingly different depending on the commercial assay used. There is a pressing need for iPTH assay harmonisation and robust reference intervals. Reference intervals may become invalid if an assay drifts, as exemplified by adjusted calcium in this study.

Utility of plasma NGAL for the diagnosis of AKI following cardiac surgery requiring cardiopulmonary bypass: a systematic review and meta-analysis.

The objective of this study was to assess the diagnostic value of plasma neutrophil gelatinase-associated lipocalin (pNGAL) for the early diagnosis of acute kidney injury (AKI) in adult patients following cardiac surgery requiring cardiopulmonary bypass (CPB). Electronic databases and other resources were systematically searched for relevant studies. Risk of bias was assessed using the Quality Assessment for Diagnostic Accuracy Studies 2 (QUADAS-2) tool. Studies were assigned to a sub-group based on the timing of the pNGAL sample in relation to the cessation of CPB. These were < 4 h, 4-8 h, 12 h or 24 h post-cessation of CPB. Summary values for sensitivity and specificity were estimated using the hierarchical summary receiver operator characteristic (ROC) curve model. A random-effects meta-analysis of each pair of sensitivity and specificity estimates from each included study was performed. In total, 3131 patients from 16 studies were included. When taken at 4-8 h following CPB, pNGAL had superior performance for the diagnosis of AKI in the defined population when compared to earlier and later time points. Prediction regions and confidence intervals, however, demonstrated significant variability in pooled estimates of sensitivity and specificity. This is likely due to population and study design heterogeneity, lack of standardisation of assays and thresholds, and inability to distinguish the different molecular forms of NGAL. In conclusion, the diagnostic utility of pNGAL in this clinical setting is inconclusive and large individual studies of representative populations of cardiac surgery patients using assays that specifically detect NGAL in its monomeric form are required.

Clinical concordance assessment should be an integral component of laboratory method comparison studies: A regression transference of routine clinical data approach.

Manufacturer-provided assay-specific reference intervals may not compensate for between assay differences leading to method related clinical discordance. Not all laboratories, however, assess clinical concordance as part of method comparison studies. We assessed the utility of method comparison data combined with routine clinical data to assess method related clinical concordance in the diagnosis and management of subclinical hypothyroidism (SCH). Passing-Bablok method comparison regression analysis was performed for both thyroid stimulating hormone (TSH) and free thyroxine (fT4) from 100 samples analysed by Abbott and Roche methods. Primary care samples indicative of SCH were identified from the laboratory information system (LIMS) of two laboratories (one with Roche methods and one with Abbott) over four months. For Roche and Abbott TSH and fT4 results, the Passing-Bablok regression equations were used to predict Abbott and Roche results respectively. The predicted results were interpreted using manufacturer-provided assay-specific reference intervals and compared to those of the previous SCH sample exchange study. On laboratory method comparison, Roche TSH and fT4 results were 30 ± 13% and 16 ± 7% higher compared to Abbott assays respectively. Of those with results indicative of SCH by Roche assays, 76.8% would have had normal thyroid function using predicted Abbott assay results and 46.9% of those with results indicative of SCH by Abbott assays would have had a biochemical indication for levothyroxine replacement using predicted Roche assay results. The results from the regression-based approach were comparable to the previous SCH sample exchange study. A regression-based approach using routine method comparison data and real-world clinical data identifies potential clinical discordance. We suggest that clinical concordance assessment be an integral component of laboratory method comparison studies.

Acid freezes uricolysis in addition to glycolysis: Utility of fluoride-EDTA-citrate tube in the measurement of uric acid for patients administered rasburicase.

BACKGROUND: In samples from patients administered rasburicase, ex vivo uricolysis leads to spuriously low uric acid results. The manufacturer's recommendation of storing the sample in ice-water until analysis, however, does not fully arrest uricolysis. Since uricase activity is affected by pH and metal chelators, we assessed uricolysis inhibition in sodium fluoride-ethylenediaminetetraacetic acid (EDTA)-citrate sample tube (FC Mix tube, Greiner) used primarily for plasma glucose. METHOD: A serum pool was spiked with rasburicase and uric acid measured at 15, 45, 90, 150, 240 and 1080 min in a lithium heparin tube in ice-water, plain tube at room temperature (RT), EDTA tube at RT, FC Mix tube in ice-water, FC Mix tube at RT and FC Mix tube at RT prepared by dissolving FC Mix in serum. RESULTS: The rate of urate decay was lowest in the FC Mix tube independent of temperature, then lithium heparin tube in ice-water, then EDTA tube at RT and highest in the plain tube at RT. Uric acid concentrations in the prepared FC Mix tube at RT and heparin tube in ice-water were, respectively, 98.2% and 93.8% of control values at 90 min, 97.1% and 89.3% of control values at 4 h, and remained higher in the prepared FC Mix tube at all time points. CONCLUSION: NaF-EDTA-citrate mixture largely arrested rasburicase mediated ex vivo uricolysis without the need for sample cooling. We propose that sample tubes containing NaF-EDTA-citrate be used for the measurement of uric acid in patients administered rasburicase.

Parasitic nematodes of the genus Syphacia Seurat, 1916 infecting Cricetidae in the British Isles: the enigmatic status of Syphacia nigeriana.

Oxyurid nematodes (Syphacia spp.) from bank (Myodes glareolus) and field/common (Microtus spp.) voles, from disparate geographical sites in the British Isles, were examined morphologically and genetically. The genetic signatures of 118 new isolates are provided, based primarily on the rDNA internal transcribed spacers (ITS1-5.8S-ITS2) region and for representative isolates also on the small subunit 18S rDNA region and cytochrome c oxidase subunit 1 (cox-1) gene locus. Genetic data on worms recovered from Microtus spp. from the European mainland and from other rodent genera from the Palaearctic, North America and West Africa are also included. We test historical hypotheses indicating that S. nigeriana is a generalist species, infecting a range of different rodent genera. Our results establish that S. nigeriana is a parasite of both bank and field voles in the British Isles. An identical genotype was also recorded from Hubert's multimammate mouse (Mastomys huberti) from Senegal, but Mastomys spp. from West Africa were additionally parasitized by a related, although genetically distinct Syphacia species. We found no evidence for S. petrusewiczi in voles from the British Isles but isolates from Russia and North America were genetically distinct and formed their own separate deep branch in maximum likelihood molecular phylogenetic trees.

Biochemical assessment of adequate levothyroxine replacement in primary hypothyroidism differs with different TSH assays: potential clinical implications.

AIM: Thyroid stimulating hormone (TSH) assays provided by Abbott Laboratories and Roche Diagnostics are used by approximately 75% of laboratories in the UK. We assessed the potential impact of Abbott and Roche TSH assay differences on the biochemical assessment of levothyroxine replacement in primary hypothyroidism. METHOD: Samples from 100 consecutive primary care patients (83 women, median age 64 years, IQR 51-73 years) with primary hypothyroidism on adequate levothyroxine based on an Abbott Architect TSH in the reference range were analysed for TSH on Roche cobas within 24 hours. The Abbott and Roche TSH results were compared. Over 1 year, TSH results from patients in primary care from the laboratories with Abbott and Roche methods were compared. RESULTS: The median (IQR) Roche TSH (2.5 (1.3-3.6) mIU/L) was 30%±10% higher (p<0.001) than Abbott TSH (1.9 (1.1-2.6) mIU/L). Although all Abbott TSH results were in the Abbott specific reference range, 14 patients (14%) had Roche TSH results above the Roche specific reference range. In the 1 year gather, Roche TSH (1.9 (1.3-2.9) mIU/L, n=103 932) results were higher (p<0.001) than Abbott TSH (1.5 (1.0-2.2) mIU/L, n=1 10 544) results. The TSH results were above their assay-specific upper reference limit in 10.7% of Roche results and 4.2% of Abbott results. CONCLUSION: Biochemical assessment of levothyroxine replacement may be dependent on the type of TSH assay. Laboratorians and clinicians should be aware that the lack of harmonisation between TSH methods and their assay-specific reference ranges may potentially lead to different patient management decisions. We suggest lot verification in laboratories should include processes to identify cumulative drift in assay performance.

A lesson from the wild: The natural state of eosinophils is Ly6Ghi.

With a long history of promoting pathological inflammation, eosinophils are now emerging as important regulatory cells. Yet, findings from controlled laboratory experiments so far lack translation to animals, including humans, in their natural environment. In order to appreciate the breadth of eosinophil phenotype under non-laboratory, uncontrolled conditions, we exploit a free-living population of the model organism Mus musculus domesticus. Eosinophils were present at significantly higher proportions in the spleen and bone marrow of wild mice compared with laboratory mice. Strikingly, the majority of eosinophils of wild mice exhibited a unique Ly6Ghi phenotype seldom described in laboratory literature. Ly6G expression correlated with activation status in spleen and bone marrow, but not peritoneal exudate cells, and is therefore likely not an activation marker per se. Intermediate Ly6G expression was transiently induced in a small proportion of eosinophils from C57BL/6 laboratory mice during acute infection with the whipworm Trichuris muris, but not during low-dose chronic infection, which better represents parasite exposure in the wild. We conclude that the natural state of the eosinophil is not adequately reflected in the standard laboratory mouse, which compromises our attempts to dissect their functional relevance. Our findings emphasize the importance of studying the immune system in its natural context - alongside more mechanistic laboratory experiments - in order to capture the entirety of immune phenotypes and functions.

Samples spiked with pituitary-derived thyroid-stimulating hormone may disguise the extent of differences between thyroid-stimulating hormone assays.

BACKGROUND: A large discordance in the diagnosis and potential management of hypothyroidism using Abbott and Roche thyroid assays has been reported recently. The difference in Abbott and Roche thyroid-stimulating hormone (TSH) results in these studies was larger than anticipated from the external quality assessment (EQA) reports. METHODS: Abbott and Roche TSH method means in UK NEQAS for thyroid hormones distributions 430 to 454 were compared against the amount of TSH spiked. A TSH deplete serum pool was spiked with various concentrations of pooled high TSH serum and 3rd WHO International Standard for TSH (WHO-IS). Four serum pools with TSH close to clinical decision limits were spiked with two concentrations of WHO-IS. RESULTS: On review of EQA data, median (IQR) Roche: Abbott TSH ratio was lower (p < 0.001) in 48 pools spiked with TSH (1.11 (1.07-1.16)) compared to 41 pools not spiked (1.29 (1.25-1.31)) and the decrease was proportionate to the contribution of spiked TSH to total TSH in the samples (ρ=-0.908, p < 0.001). In spiking experiments, the relationship of Roche and Abbott TSH was different in TSH deplete pool spiked with WHO-IS (RocheTSH=1.13*AbbottTSH-0.52) and high TSH serum (RocheTSH=1.43*AbbottTSH-0.50), respectively. The Roche: Abbott TSH ratio decreased and the method agreement improved on spiking serum pools with WHO-IS. CONCLUSION: Abbott and Roche TSH assays are not in harmony in human serum samples but the agreement was better in samples spiked with WHO-IS which contains pituitary-derived TSH. Use of pituitary-derived TSH spiked samples, such as provided by EQA schemes, may mask clinically significant between-assay differences.

Hypophosphatasia.

Hypophosphatasia (HPP) is a group of inherited disorders characterised by the impaired mineralisation of bones and/or teeth and low serum alkaline phosphatase (ALP) activity. It is caused by a mutation in the ALPL gene encoding the tissue-non-specific isoenzyme of ALP (TNSALP) resulting in a loss of function. The disease is highly heterogenous in its clinical expression ranging from stillbirth without mineralised bone to the mild form of late adult onset with symptoms and signs such as musculoskeletal pain, arthropathy, lower-extremity fractures, premature loss of teeth or an incidental finding of reduced serum ALP activity. A classification based on the age at diagnosis and the presence or absence of bone symptoms was historically used: perinatal, prenatal benign, infantile, childhood, adult and odontohypophosphatasia. These subtypes are known to have overlapping signs and complications. Three forms of HPP distinguishable by their genetic characteristics have been described: severe, moderate and mild. Severe forms of HPP (perinatal and infantile severe) are recessively inherited, whereas moderate HPP may be dominantly or recessively inherited. The biochemical hallmark of HPP is persistently low serum ALP for age and increase in natural substrates of TNSALP, pyridoxal 5'-phosphate and phosphoethanolamine supported by radiological findings. The diagnosis is confirmed by ALPL sequencing. A multidisciplinary team of experts is essential for the effective management. Calcium restriction is recommended in infants/children to manage hypercalcaemia. A targeted enzyme replacement therapy for HPP has become available and correct diagnosis is crucial to allow early treatment.

The diagnosis and management of subclinical hypothyroidism is assay-dependent- Implications for clinical practice.

OBJECTIVE: We assessed the commutativity of Roche and Abbott thyroid assays in the diagnosis and management of subclinical hypothyroidism (SCH). The Roche and Abbott thyroid assays are used by approximately 75% of clinical laboratories in the UK. METHOD: Consecutive samples received from primary care on patients with SCH who had a raised thyroid-stimulating hormone (TSH) <10 mIU/L and a normal free thyroxine (fT4) from two laboratories using either Roche or Abbott thyroid assays were identified over 10 working days. Following identification, samples were analysed at the other site within 24 hours. Diagnostic and management discordance were studied using the relevant manufacturer-provided reference ranges. RESULTS: We identified 93 patients with SCH (53 using the Roche assay). Roche TSH and fT4 results were respectively 40% ± 15% and 16% ± 7% higher (P < .001) compared to Abbott results. Of the 93 patients, 41 (44%) were concordant for SCH on both methods. Of the 53 patients with SCH on the Roche assays, 40 (75.5%) had normal thyroid function and 13 (24.5%) had SCH when analysed using the Abbott assays. Of the 40 patients with SCH on the Abbott assays, 28 (70%) had SCH and 12 (30%) had results indicative for levothyroxine replacement when analysed on the Roche assays. Of these 12 patients, four had TSH > 10 mIU/L, five had low fT4 and three had both. CONCLUSION: The diagnosis and management of SCH is strikingly different when using TSH and fT4 assays provided by Abbott Laboratories and Roche Diagnostics. Clinicians and laboratorians should be aware that between-assay differences and variations in reference ranges will directly impact the diagnosis and management of subclinical hypothyroidism.

Relationships between immune gene expression and circulating cytokine levels in wild house mice.

Quantitative PCR (qPCR) has been commonly used to measure gene expression in a number of research contexts, but the measured RNA concentrations do not always represent the concentrations of active proteins which they encode. This can be due to transcriptional regulation or post-translational modifications, or localization of immune environments, as can occur during infection. However, in studies using free-living non-model species, such as in ecoimmunological research, qPCR may be the only available option to measure a parameter of interest, and so understanding the quantitative link between gene expression and associated effector protein levels is vital.Here, we use qPCR to measure concentrations of RNA from mesenteric lymph node (MLN) and spleen tissue, and multiplex ELISA of blood serum to measure circulating cytokine concentrations in a wild population of a model species, Mus musculus domesticus.Few significant correlations were found between gene expression levels and circulating cytokines of the same immune genes or proteins, or related functional groups. Where significant correlations were observed, these were most frequently within the measured tissue (i.e., the expression levels of genes measured from spleen tissue were more likely to correlate with each other rather than with genes measured from MLN tissue, or with cytokine concentrations measured from blood).Potential reasons for discrepancies between measures including differences in decay rates and transcriptional regulation networks are discussed. We highlight the relative usefulness of different measures under different research questions and consider what might be inferred from immune assays.

Immune state is associated with natural dietary variation in wild mice Mus musculus domesticus.

The ability, propensity and need to mount an immune response vary both among individuals and within a single individual over time.A wide array of parameters has been found to influence immune state in carefully controlled experiments, but we understand much less about which of these parameters are important in determining immune state in wild populations.Diet can influence immune responses, for example when nutrient availability is limited. We therefore predict that natural dietary variation will play a role in modulating immune state, but this has never been tested.We measured carbon and nitrogen stable isotope ratios in an island population of house mice Mus musculus domesticus as an indication of dietary variation, and the expression of a range of immune-related genes to represent immune state.After accounting for potential confounding influences such as age, sex and helminth load, we found a significant association between carbon isotope ratio and levels of immune activity in the mesenteric lymph nodes, particularly in relation to the inflammatory response.This association demonstrates the important interplay between diet and an animal's response to immune challenges, and therefore potentially its susceptibility to disease. A plain language summary is available for this article.