Experimental evidence for opposing effects of high deer density on tick-borne pathogen prevalence and hazard

BACKGROUND: Identifying the mechanisms driving disease risk is challenging for multi-host pathogens, such as Borrelia burgdorferi sensu lato (s.l.), the tick-borne bacteria causing Lyme disease. Deer are tick reproduction hosts but do not transmit B. burgdorferi s.l., whereas rodents and birds are competent transmission hosts. Here, we use a long-term deer exclosure experiment to test three mechanisms for how high deer density might shape B. burgdorferi s.l. prevalence in ticks: increased prevalence due to higher larval tick densities facilitating high transmission on rodents (M1); alternatively, reduced B. burgdorferi s.l. prevalence because more larval ticks feed on deer rather than transmission-competent rodents (dilution effect) (M2), potentially due to ecological cascades, whereby higher deer grazing pressure shortens vegetation which decreases rodent abundance thus reducing transmission (M3). METHODS: In a large enclosure where red deer stags were kept at high density (35.5 deer km(−2)), we used an experimental design consisting of eight plots of 0.23 ha, four of which were fenced to simulate the absence of deer and four that were accessible to deer. In each plot we measured the density of questing nymphs and nymphal infection prevalence in spring, summer and autumn, and quantified vegetation height and density, and small mammal abundance. RESULTS: Prevalence tended to be lower, though not conclusively so, in high deer density plots compared to exclosures (predicted prevalence of 1.0% vs 2.2%), suggesting that the dilution and cascade mechanisms might outweigh the increased opportunities for transmission mechanism. Presence of deer at high density led to shorter vegetation and fewer rodents, consistent with an ecological cascade. However, Lyme disease hazard (density of infected I. ricinus nymphs) was five times higher in high deer density plots due to tick density being 18 times higher. CONCLUSIONS: High densities of tick reproduction hosts such as deer can drive up vector-borne disease hazard, despite the potential to simultaneously reduce pathogen prevalence. This has implications for environmental pathogen management and for deer management, although the impact of intermediate deer densities now needs testing. GRAPHICAL ABSTRACT: [Image: see text] SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s13071-021-05000-0

Anaplasma phagocytophilum ecotype analysis in cattle from Great Britain

Anaplasma phagocytophilum (A. phagocytophilum) is the aetiological agent of tick-borne fever in cattle and sheep, and granulocytic anaplasmosis in human and dogs. Livestock, companion animal and human infections with A. phagocytophilum have been reported globally. Across England and Wales, two isolates (called ecotypes) have been reported in ticks. This study examined A. phagocytophilum isolates present in livestock and wildlife in Great Britain (GB), with a particular focus on cattle. Clinical submissions (EDTA blood) from cattle (n = 21) and sheep (n = 3) were received by APHA for tick-borne disease testing and the animals were confirmed to be infected with A. phagocytophilum using a PCR targeting the Msp2 gene. Further submissions from roe deer (n = 2), red deer (n = 2) and Ixodes ricinus ticks (n = 22) were also shown to be infected with A. phagocytophilum. Subsequent analysis using a nested PCR targeting the groEL gene and sequencing confirmed the presence of ecotype I in cattle, sheep, red deer and Ixodes ricinus, and ecotype II in roe deer and I. ricinus removed from deer carcasses. Despite the presence of two ecotypes, widely distributed in ticks from England and Wales, only ecotype I was detected in cattle in this study

Sequences related to Chimay rhabdovirus are widely distributed in Ixodes ricinus ticks across England and Wales

Ticks are the main arthropod vector of pathogens to humans and livestock in the British Isles. Despite their role as a vector of disease, many aspects of tick biology, ecology, and microbial association are poorly understood. To address this, we investigated the composition of the microbiome of adult and nymphal Ixodes ricinus ticks. The ticks were collected on a dairy farm in Southwest England and RNA extracted for whole genome sequencing. Sequences were detected from a range of microorganisms, particularly tick-associated viruses, bacteria, and nematodes. A majority of the viruses were attributed to phlebo-like and nairo-like virus groups, demonstrating a high degree of homology with the sequences present in I. ricinus from mainland Europe. A virus sharing a high sequence identity with Chimay rhabdovirus, previously identified in ticks from Belgium, was detected. Further investigations of I. ricinus ticks collected from additional sites in England and Wales also identified Chimay rhabdovirus viral RNA with varying prevalence in all tick populations. This suggests that Chimay rhabdovirus has a wide distribution and highlights the need for an extended exploration of the tick microbiome in the United Kingdom (UK)

Growth and retreat of the last British–Irish Ice Sheet, 31 000 to 15 000 years ago: the BRITICE-CHRONO reconstruction

The BRITICE-CHRONO consortium of researchers undertook a dating programme to constrain the timing of advance, maximum extent and retreat of the British–Irish Ice Sheet between 31 000 and 15 000 years before present. The dating campaign across Ireland and Britain and their continental shelves, and across the North Sea included 1500 days of field investigation yielding 18 000 km of marine geophysical data, 377 cores of sea floor sediments, and geomorphological and stratigraphical information at 121 sites on land; generating 690 new geochronometric ages. These findings are reported in 28 publications including synthesis into eight transect reconstructions. Here we build ice sheet-wide reconstructions consistent with these findings and using retreat patterns and dates for the inter-transect areas. Two reconstructions are presented, a wholly empirical version and a version that combines modelling with the new empirical evidence. Palaeoglaciological maps of ice extent, thickness, velocity, and flow geometry at thousand-year timesteps are presented. The maximum ice volume of 1.8 m sea level equivalent occurred at 23 ka. A larger extent than previously defined is found and widespread advance of ice to the continental shelf break is confirmed during the last glacial. Asynchrony occurred in the timing of maximum extent and onset of retreat, ranging from 30 to 22 ka. The tipping point of deglaciation at 22 ka was triggered by ice stream retreat and saddle collapses. Analysis of retreat rates leads us to accept our hypothesis that the marine-influenced sectors collapsed rapidly. First order controls on ice-sheet demise were glacio-isostatic loading triggering retreat of marine sectors, aided by glaciological instabilities and then climate warming finished off the smaller, terrestrial ice sheet. Overprinted on this signal were second order controls arising from variations in trough topographies and with sector-scale ice geometric readjustments arising from dispositions in the geography of the landscape. These second order controls produced a stepped deglaciation. The retreat of the British–Irish Ice Sheet is now the world’s most well-constrained and a valuable data-rich environment for improving ice-sheet modelling.publishedVersio

Albiglutide and cardiovascular outcomes in patients with type 2 diabetes and cardiovascular disease (Harmony Outcomes): a double-blind, randomised placebo-controlled trial

Background: Glucagon-like peptide 1 receptor agonists differ in chemical structure, duration of action, and in their effects on clinical outcomes. The cardiovascular effects of once-weekly albiglutide in type 2 diabetes are unknown. We aimed to determine the safety and efficacy of albiglutide in preventing cardiovascular death, myocardial infarction, or stroke. Methods: We did a double-blind, randomised, placebo-controlled trial in 610 sites across 28 countries. We randomly assigned patients aged 40 years and older with type 2 diabetes and cardiovascular disease (at a 1:1 ratio) to groups that either received a subcutaneous injection of albiglutide (30–50 mg, based on glycaemic response and tolerability) or of a matched volume of placebo once a week, in addition to their standard care. Investigators used an interactive voice or web response system to obtain treatment assignment, and patients and all study investigators were masked to their treatment allocation. We hypothesised that albiglutide would be non-inferior to placebo for the primary outcome of the first occurrence of cardiovascular death, myocardial infarction, or stroke, which was assessed in the intention-to-treat population. If non-inferiority was confirmed by an upper limit of the 95% CI for a hazard ratio of less than 1·30, closed testing for superiority was prespecified. This study is registered with ClinicalTrials.gov, number NCT02465515. Findings: Patients were screened between July 1, 2015, and Nov 24, 2016. 10 793 patients were screened and 9463 participants were enrolled and randomly assigned to groups: 4731 patients were assigned to receive albiglutide and 4732 patients to receive placebo. On Nov 8, 2017, it was determined that 611 primary endpoints and a median follow-up of at least 1·5 years had accrued, and participants returned for a final visit and discontinuation from study treatment; the last patient visit was on March 12, 2018. These 9463 patients, the intention-to-treat population, were evaluated for a median duration of 1·6 years and were assessed for the primary outcome. The primary composite outcome occurred in 338 (7%) of 4731 patients at an incidence rate of 4·6 events per 100 person-years in the albiglutide group and in 428 (9%) of 4732 patients at an incidence rate of 5·9 events per 100 person-years in the placebo group (hazard ratio 0·78, 95% CI 0·68–0·90), which indicated that albiglutide was superior to placebo (p<0·0001 for non-inferiority; p=0·0006 for superiority). The incidence of acute pancreatitis (ten patients in the albiglutide group and seven patients in the placebo group), pancreatic cancer (six patients in the albiglutide group and five patients in the placebo group), medullary thyroid carcinoma (zero patients in both groups), and other serious adverse events did not differ between the two groups. There were three (<1%) deaths in the placebo group that were assessed by investigators, who were masked to study drug assignment, to be treatment-related and two (<1%) deaths in the albiglutide group. Interpretation: In patients with type 2 diabetes and cardiovascular disease, albiglutide was superior to placebo with respect to major adverse cardiovascular events. Evidence-based glucagon-like peptide 1 receptor agonists should therefore be considered as part of a comprehensive strategy to reduce the risk of cardiovascular events in patients with type 2 diabetes. Funding: GlaxoSmithKline

Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition)

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. For example, a key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process versus those that measure fl ux through the autophagy pathway (i.e., the complete process including the amount and rate of cargo sequestered and degraded). In particular, a block in macroautophagy that results in autophagosome accumulation must be differentiated from stimuli that increase autophagic activity, defi ned as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (inmost higher eukaryotes and some protists such as Dictyostelium ) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the fi eld understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. It is worth emphasizing here that lysosomal digestion is a stage of autophagy and evaluating its competence is a crucial part of the evaluation of autophagic flux, or complete autophagy. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. Along these lines, because of the potential for pleiotropic effects due to blocking autophagy through genetic manipulation it is imperative to delete or knock down more than one autophagy-related gene. In addition, some individual Atg proteins, or groups of proteins, are involved in other cellular pathways so not all Atg proteins can be used as a specific marker for an autophagic process. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field

The impacts of host community composition on Lyme disease risk in Scotland

Emergence of zoonoses are driven by multiple factors, ranging from climate change to urbanization and human behaviours. Because many zoonotic pathogens are maintained in wild reservoir hosts, these factors of emergence may affect disease risk by changing host community parameters. Thus, it is important to understand the effect of host community composition on disease risk. This is particularly relevant for vector-borne zoonoses as host community composition might affect both reservoir host and vector populations. In the northern hemisphere, Lyme disease, a zoonosis caused by the bacterial complex Borrelia burgdorferi sensu lato, is the most prevalent vector-borne disease affecting humans. Transmitted by Ixodid ticks, its epidemiological cycle is complex and depends on environmental factors and host community composition, which together, influence both tick survival and the prevalence of B. burgdorferi s.l. Small mammals are competent reservoir hosts for B. afzelii, a genospecies belonging to the B. burgdorferi s.l. complex, while birds can transmit B. valaisiana and B. garinii, two other genospecies in the complex. Thus, pathogen prevalence will depend on the abundance of competent reservoir hosts in an environment. Deer species, on the other hand, are non-competent hosts and could, theoretically, lower B. burgdorferi s.l. prevalence by diverting immature ticks from feeding on competent reservoir hosts, a concept called the dilution effect. However, deer also act as the main tick reproduction hosts, feeding adult female ticks, and can therefore maintain high tick populations. In this thesis, I used a range of approaches (large-scale cross-sectional survey, deer exclosure and wood mouse supplementary feeding experiment) to test hypotheses and understand how host community composition drives Lyme disease hazard (defined here as the density of infected nymphal ticks) in Scotland and to investigate the drivers of host communities. I investigated how the ratio of small mammals to deer affected Lyme disease hazard by conducting a large cross-sectional survey across sites selected to specifically cover the full range of deer densities (Chapter 2) and collecting data on host abundance (deer and small mammals), tick density and pathogen prevalence. There was a positive association between the density of infected nymphs (DIN) for B. afzelii and deer density, regardless of small mammal abundance. I also observed a positive association between deer density and quest in deer density was also negatively associated with human activity, which therefore has the potential to impact Lyme disease hazard. Having shown how the full range of deer densities shapes Lyme disease hazard, which also suggested that deer may affect small mammal densities (Chapter 2), I then wanted to investigate further the possibility that high deer densities may affect Lyme disease by causing ecological cascades, through an impact on vegetation and small mammals (Chapter 3). I used an experimental design consisting of replicated fenced deer exclosures to investigate the effects of high deer density versus deer absence on Lyme disease hazard through ecological cascades. Consistent with my predictions, high deer density plots had 18 times more questing nymphs compared to plots where deer were absent. High deer density plots also had 13 times fewer small mammals and were associated with shorter and sparser vegetation and shorter trees, highlighting the impacts of browsing pressure by deer on small mammal communities. I found that the reduction in competent host abundance had repercussions on B. burgdorferi s.l. prevalence in questing ticks, which was twice as high in deer exclusion plots (2.2% in deer exclusion plots vs 1.0% in high deer density plots). Despite the negative effect of high deer density on B. burgdorferi s.l. prevalence, DIN was five times higher in high deer density plots. These results demonstrate that the positive effect of deer on tick density can outweigh their negative effect on B. burgdorferi s.l. prevalence caused by a dilution effect and through their negative effects on small mammals. In chapters 2 and 3, I found that deer appeared to be more important than small mammals in driving DIN, through their strong role as tick reproduction hosts, irrespective of any dilution effects on prevalence. However, few of my sites had high small mammal densities while many had high deer densities. I therefore wanted to test the effect of a variation in small mammal abundance on ticks, B. burgdorferi s.l. prevalence and DIN using a supplementary feeding experiment to increase small mammal densities. More specifically, the objective for this study (Chapter 4) was to understand the effects of a change in food resources on small mammal populations and the repercussions on Lyme disease hazard. Fluctuations in resources availability are common occurrences in nature so it is important to understand the consequences this could have for disease hazard. This involved an experimental design where two trapping grids out of four were supplemented with food for two consecutive years. Density of live-trapped wood mice (Apodemus sylvaticus) was twice as high in food supplemented grids during the year of treatment. Food supplementation per se, which reflects what would happen after a mast seeding event, did not affect the density of nymphs, B. burgdorferi s.l. prevalence or DIN. However, there was a positive correlation between wood mouse abundance and questing nymph abundance the following spring. I hypothesised a negative association between wood mouse abundance and the prevalence of the bird associated B. garinii and B. valaisiana (dilution effect where more ticks would be feeding on mice instead of birds) but I did not find evidence for that. Unexpectedly, there was a positive association between wood mouse abundance and DIN for B. garinii and B.valaisiana the following spring. This could occur either if mice acted like deer did in Chapters 2 and 3, as a tick amplification host, since they increased nymph density but not B. garinii and B. valaisiana prevalence and/or if the food supplementation that increased mice also increased birds, which I did not measure. Even though food supplementation per se did not have a direct impact on DIN, wood mouse abundance had a positive effect on tick density and DIN for B. garinii and B. valaisiana and these results highlight how variations in food resources can affect small mammals and Lyme disease hazard. The cross-sectional study (Chapter 2) is the first, to my knowledge, to test how the ratio of competent reservoirs hosts (small mammals) to non-competent hosts (deer) affects Lyme disease hazard, whilst also specifically selecting sites with the full range of deer densities to robustly test the dilution effect. While I predicted that DIN should be modulated by variations in the density of both hosts, my results really pointed to deer density being the key component driving Lyme disease hazard. This result, which was further confirmed in Chapter 3, could be used for deer management decisions in Scottish woodlands. This thesis also demonstrates the complexity of Lyme disease ecology: not only due to the multiple host types and multiple pathogens in the burgdorferi s.l. complex, but also in terms of how the hosts themselves interact and affect each other’s distributions and densities. There is now clearly a need for further research to better understand the mechanisms driving Lyme disease hazard, including interspecific interactions and other drivers suggested in this thesis, such as human disturbance and food resources, that affect host abundance. Combining experimental designs and large-scale surveys allowed a better understanding of the effects of deer and small mammals simultaneously on Lyme disease. Indeed, the experimental designs (Chapters 3 and 4) offered the chance to vary the density of one host species and observe the cascading effects on other hosts, tick density and burgdorferi s.l. prevalence. Results from the cross sectional-survey (Chapter 2) not only corroborated the key associations observed but, importantly, allowed me to robustly test the dilution effect by examining the effects of a wide range of deer densities in the natural environment on DIN. The results obtained can be used to understand and predict how environmental changes (e.g. increase of resources, increase of human activity) might impact host species and Lyme disease hazard. Furthermore, they could be used to predict the effects that a change in deer management might have on tick density and DIN

No net effect of host density on tick-borne disease hazard due to opposing roles of vector amplification and pathogen dilution

To better understand vector-borne disease dynamics, knowledge of the ecological interactions between animal hosts, vectors, and pathogens is needed. The effects of hosts on disease hazard depends on their role in driving vector abundance and their ability to transmit pathogens. Theoretically, a host that cannot transmit a pathogen could dilute pathogen prevalence but increase disease hazard if it increases vector population size. In the case of Lyme disease, caused by Borrelia burgdorferi s.l. and vectored by Ixodid ticks, deer may have dual opposing effects on vectors and pathogen: deer drive tick population densities but do not transmit B. burgdorferi s.l. and could thus decrease or increase disease hazard. We aimed to test for the role of deer in shaping Lyme disease hazard by using a wide range of deer densities while taking transmission host abundance into account. We predicted that deer increase nymphal tick abundance while reducing pathogen prevalence. The resulting impact of deer on disease hazard will depend on the relative strengths of these opposing effects. We conducted a cross-sectional survey across 24 woodlands in Scotland between 2017 and 2019, estimating host (deer, rodents) abundance, questing Ixodes ricinus nymph density, and B. burgdorferi s.l. prevalence at each site. As predicted, deer density was positively associated with nymph density and negatively with nymphal infection prevalence. Overall, these two opposite effects canceled each other out: Lyme disease hazard did not vary with increasing deer density. This demonstrates that, across a wide range of deer and rodent densities, the role of deer in amplifying tick densities cancels their effect of reducing pathogen prevalence. We demonstrate how noncompetent host density has little effect on disease hazard even though they reduce pathogen prevalence, because of their role in increasing vector populations. These results have implications for informing disease mitigation strategies, especially through host management

Experimental Evidence for Opposing Effects of High Deer Density on Tick-Borne Pathogen Prevalence and Hazard

Abstract Background: Identifying the mechanisms driving disease risk is challenging for multi-host pathogens, such as Borrelia burgdorferi s.l., the tick-borne bacteria causing Lyme disease. Deer are tick reproduction hosts but do not transmit B. burgdorferi s.l., whereas rodents and birds are competent transmission hosts. Here, we use a long-term deer exclosure experiment to test three mechanisms for how high deer density might shape B. burgdorferi s.l. prevalence in ticks: increased prevalence due to higher larval tick densities facilitating high transmission on rodents (M1); alternatively, reduced B. burgdorferi s.l. prevalence because more larval ticks feed on deer rather than transmission-competent rodents (dilution effect) (M2), potentially due to ecological cascades, whereby higher deer grazing pressure lowers vegetation which decreases rodent abundance thus reducing transmission (M3).Methods: In a large enclosure where red deer stags were kept at high density (32.5 deer/km²), we used an experimental design consisting of eight plots of 0.23ha, four being fenced to simulate the absence of deer and four that were accessible to deer. In each plot we measured the density of questing nymphs and nymphal infection prevalence in spring, summer and autumn and quantified vegetation height and density, and small mammal abundance Results: Prevalence tended to be lower, though not conclusively so, in high deer density plots compared to exclosures (predicted prevalence of 1.0% vs 2.2%), suggesting that the dilution (M2) and cascade (M3) mechanisms might outweigh the increased opportunities for transmission (M1). Presence of deer at high density led to lower vegetation and fewer rodents, consistent with an ecological cascade. However, Lyme disease hazard (density of infected I. ricinus nymphs) was five times higher in high deer density plots due to tick density being 18 times higher.Conclusion: High densities of tick reproduction hosts such as deer can drive up vector-borne disease hazard, despite the potential to simultaneously reduce pathogen prevalence. This has implications for environmental pathogen management and for deer management, although the impact of intermediate deer densities now needs testing.</jats:p