The application of next-generation sequence-based DNA barcoding for bloodmeal detection in host-seeking wild-caught Ixodes scapularis nymphs.

OBJECTIVE: Our objective was to apply next-generation sequence-based DNA barcoding to identify the remnant larval bloodmeals in wild-caught host-seeking (unengorged) Ixodes scapularis nymphs (n = 216). To infer host species identification, vertebrate DNA was amplified using universal primers for cytochrome c oxidase subunit I (COI) and sequenced using next-generation sequencing (NGS) for comparison against known barcode references. RESULTS: Bloodmeal identification was unsuccessful in most samples (99% of 216 specimens) demonstrating a very low detection rate of this assay. Sequences that surpassed quality thresholds were obtained for 41.7% of nymphs (n = 90) and of those, confident species identification was obtained for 15.6% of nymphs (n = 14). Wild host identifications were only obtained from 2 specimens, where DNA from the eastern grey squirrel (Sciurus carolinensis) was identified. Human and bovine DNA was identified in remaining nymphs and considered to be contaminants. Further optimization of the technique is required to improve detection of remnant bloodmeals in host-seeking nymphs.

N Increased risk of tick-borne diseases with climate and environmental changes.

Climate warming and other environmental changes have contributed to the expansion of the range of several tick species into higher latitudes in North America. As temperatures increase in Canada, the environment becomes more suitable for ticks and the season suitable for tick activity lengthens, so tick-borne diseases are likely to become more common in Canada. In addition to Lyme disease, four other tick-borne diseases (TBDs) have started to emerge and are likely to increase: Anaplasmosis; Babesiosis; Powassan virus; and Borrelia miyamotoi disease. Increased temperature increases the survival and activity period of ticks, increases the range of both reservoir and tick hosts (e.g. mice and deer) and increases the duration of the season when people may be exposed to ticks. Other ticks and TBDs may spread into Canada as the climate changes. The public health strategies to mitigate the impact of all TBDs include surveillance to detect current and emerging TBDs, and public health actions to prevent infections by modifying environmental and social-behavioral risk factors through increasing public awareness. Clinical care strategies include patient education, early detection, laboratory testing, and treatment.

Increased risk of endemic mosquito-borne diseases in Canada due to climate change.

There are currently over 80 species of mosquito endemic in Canada-although only a few of these carry pathogens that can cause disease in humans. West Nile virus, Eastern equine encephalitis virus and the California serogroup viruses (including the Jamestown Canyon and snowshoe hare viruses) are mosquito-borne viruses that have been found to cause human infections in North America, including in Canada. Over the last 20 years, the incidence of most of these endemic mosquito-borne diseases (MBD) has increased approximately 10% in Canada, due in large part to climate change. It is anticipated that both the mosquito lifecycle and virus transmission patterns will be affected by climate change, resulting in an increase in both the range and local abundance of several important mosquito species. Laboratory studies and mathematical modelling suggest that increased ambient temperatures, changes in precipitation and extreme weather events associated with climate change will likely continue to drive mosquito vector and MBD range expansion, increasing the duration of transmission seasons and leading to MBD-related epidemics. Furthermore, Canada's endemic MBDs have complex transmission cycles, involving multiple reservoir hosts (birds and mammals), multiple pathogens and multiple mosquito species-all of which may be sensitive to climate and other environmental changes, and making forecasting of potential emerging trends difficult. These expected climate-induced changes in mosquitoes and MBDs underline the need for continued (and expanded) surveillance and research to ensure timely and accurate evaluation of the risks to the public health of Canadians.

Could exotic mosquito-borne diseases emerge in Canada with climate change?

Of the 3,500 species of mosquitoes worldwide, only a small portion carry and transmit the mosquito-borne diseases (MBDs) that cause approximately half a million deaths annually worldwide. The most common exotic MBDs, such as malaria and dengue, are not currently established in Canada, in part because of our relatively harsh climate; however, this situation could evolve with climate change. Mosquitoes native to Canada may become infected with new pathogens and move into new regions within Canada. In addition, new mosquito species may move into Canada from other countries, and these exotic species may bring exotic MBDs as well. With high levels of international travel, including to locations with exotic MBDs, there will be more travel-acquired cases of MBDs. With climate change, there is the potential for exotic mosquito populations to become established in Canada. There is already a small area of Canada where exotic Aedes mosquitoes have become established although, to date, there is no evidence that these carry any exotic (or already endemic) MBDs. The increased risks of spreading MBDs, or introducing exotic MBDs, will need a careful clinical and public health response. Clinicians will need to maintain a high level of awareness of current trends, to promote mosquito bite prevention strategies, and to know the laboratory tests needed for early detection and when to report laboratory results to public health. Public health efforts will need to focus on ongoing active surveillance, public and professional awareness and mosquito control. Canadians need to be aware of the risks of acquiring exotic MBDs while travelling abroad as well as the risk that they could serve as a potential route of introduction for exotic MBDs into Canada when they return home.

Weather-based forecasting of mosquito-borne disease outbreaks in Canada.

Early warning systems to predict infectious disease outbreaks have been identified as a key adaptive response to climate change. Warming, climate variability and extreme weather events associated with climate change are expected to drive an increase in frequency and intensity of mosquito-borne disease (MBD) outbreaks globally. In Canada, this will mean an increased risk of endemic and emerging MBD outbreaks such as West Nile virus and other MBDs. The availability of timely information on the risk of impending MBD outbreaks has important public health implications, by allowing implementation of mosquito control measures and targeted communications regarding the need for increased personal protective measures-before an outbreak occurs. In Canada, both mechanistic and statistical weather-based models have been developed to predict West Nile virus outbreaks. These include models for different species of mosquitoes that transmit West Nile virus in different geographical areas of Canada. Although initial results have been promising, further validation and assessment of forecasting skill are needed before wide scale implementation. Weather-based forecasting for other emerging MBDs in Canada, such as Eastern equine encephalitis, may also be feasible.

What is the real number of Lyme disease cases in Canada?

BACKGROUND: Lyme disease is emerging in Canada due to expansion of the range of the tick vector Ixodes scapularis from the United States. National surveillance for human Lyme disease cases began in Canada in 2009. Reported numbers of cases increased from 144 cases in 2009 to 2025 in 2017. It has been claimed that few (< 10%) Lyme disease cases are reported associated with i) supposed under-diagnosis resulting from perceived inadequacies of serological testing for Lyme disease, ii) expectation that incidence in Canadian provinces and neighbouring US states should be similar, and iii) analysis of serological responses of dogs to the agent of Lyme disease, Borrelia burgdorferi. We argue that performance of serological testing for Lyme disease is well studied, and variations in test performance at different disease stages are accounted for in clinical diagnosis of Lyme disease, and in surveillance case definitions. Extensive surveillance for tick vectors has taken place in Canada providing a clear picture of the emergence of risk in the Canadian environment. This surveillance shows that the geographic scope of I. scapularis populations and Lyme disease risk is limited but increasing in Canada. The reported incidence of Lyme disease in Canada is consistent with this pattern of environmental risk, and the differences in Lyme disease incidence between US states and neighbouring Canadian provinces are consistent with geographic differences in environmental risk. Data on serological responses in dogs from Canada and the US are consistent with known differences in environmental risk, and in numbers of reported Lyme disease cases, between the US and Canada. CONCLUSION: The high level of consistency in data from human case and tick surveillance, and data on serological responses in dogs, suggests that a high degree of under-reporting in Canada is unlikely. We speculate that approximately one third of cases are reported in regions of emergence of Lyme disease, although prospective studies are needed to fully quantify under-reporting. In the meantime, surveillance continues to identify and track the ongoing emergence of Lyme disease, and the risk to the public, in Canada.

Risk factors associated with seropositivity to California serogroup viruses in humans and pet dogs, Quebec, Canada.

Jamestown Canyon and snowshoe hare viruses are two emerging human pathogens associated with cases of neuroinvasive disease in North America. This study aimed to identify environmental and individual risk factors for seropositivity to these arboviruses in humans and pet dogs from Québec, Canada, 2012-2014. In humans, areas with moderate densities of white-tailed deer (Odocoileus virginianus) were associated with higher odds of seropositivity compared with areas with low densities of white-tailed deer (OR 2.50, P = 0.009) and odds of seropositivity were higher in males than in females (OR 2.03, P = 0.016). Among humans reporting more than 10 mosquito bites weekly, the odds of being seropositive were 4.44 times higher (P = 0.004) for people living in hardwood forested areas. Exposure to areas with coniferous forests was identified as the main environmental risk factor for seroconversion in dogs (OR 2.39, P = 0.04). These findings may help target further public health research, diagnostic and surveillance efforts in Canada.

Emerging arboviruses in Quebec, Canada: assessing public health risk by serology in humans, horses and pet dogs.

Periodic outbreaks of West Nile virus (WNV), Eastern equine encephalitis virus (EEEV) and to a lesser extent, California serogroup viruses (CSGV), have been reported in parts of Canada in the last decade. This study was designed to provide a broad assessment of arboviral activity in Quebec, Canada, by conducting serological surveys for these arboviruses in 196 horses, 1442 dogs and 485 humans. Sera were screened by a competitive enzyme linked immunosorbent assay and positive samples confirmed by plaque reduction neutralisation tests. The percentage of seropositive samples was 83·7%, 16·5%, 7·1% in horses, 18·8%, 0·6%, 0% in humans, 11·7%, 3·1%, 0% in adult dogs and 2·9%, 0·3%, 0% in juvenile dogs for CSGV, WNV and EEEV, respectively. Serological results in horses and dogs appeared to provide a meaningful assessment of risk to public health posed by multiple arboviruses.

Characterizing environmental risk factors for West Nile virus in Quebec, Canada, using clinical data in humans and serology in pet dogs.

The identification of specific environments sustaining emerging arbovirus amplification and transmission to humans is a key component of public health intervention planning. This study aimed at identifying environmental factors associated with West Nile virus (WNV) infections in southern Quebec, Canada, by modelling and jointly interpreting aggregated clinical data in humans and serological data in pet dogs. Environmental risk factors were estimated in humans by negative binomial regression based on a dataset of 191 human WNV clinical cases reported in the study area between 2011 and 2014. Risk factors for infection in dogs were evaluated by logistic and negative binomial models based on a dataset including WNV serological results from 1442 dogs sampled from the same geographical area in 2013. Forested lands were identified as low-risk environments in humans. Agricultural lands represented higher risk environments for dogs. Environments identified as impacting risk in the current study were somewhat different from those identified in other studies conducted in north-eastern USA, which reported higher risk in suburban environments. In the context of the current study, combining human and animal data allowed a more comprehensive and possibly a more accurate view of environmental WNV risk factors to be obtained than by studying aggregated human data alone.

Evaluating the submission of digital images as a method of surveillance for Ixodes scapularis ticks.

Widespread access to the internet is offering new possibilities for data collection in surveillance. We explore, in this study, the possibility of using an electronic tool to monitor occurrence of the tick vector of Lyme disease, Ixodes scapularis. The study aimed to compare the capacity for ticks to be identified in web-based submissions of digital images/photographs, to the traditional specimen-based identification method used by the provincial public health laboratory in Quebec, Canada. Forty-one veterinary clinics participated in the study by submitting digital images of ticks collected from pets via a website for image-based identification by an entomologist. The tick specimens were then sent to the provincial public health laboratory to be identified by the 'gold standard' method using a microscope. Of the images submitted online, 74·3% (284/382) were considered of high-enough quality to allow identification. The laboratory identified 382 tick specimens from seven different species, with I. scapularis representing 76% of the total submissions. Of the 284 ticks suitable for image-based species identification, 276 (97·2%) were correctly identified (Kappa statistic of 0·92, Z = 15·46, P < 0·001). This study demonstrates that image-based tick identification may be an accurate and useful method of detecting ticks for surveillance when images are of suitable quality.

Characterizing areas of potential human exposure to eastern equine encephalitis virus using serological and clinical data from horses.

Eastern equine encephalitis (EEE) is a rare but severe emerging vector-borne disease affecting human and animal populations in the northeastern United States where it is endemic. Key knowledge gaps remain about the epidemiology of EEE virus (EEEV) in areas where its emergence has more recently been reported. In Eastern Canada, viral activity has been recorded in mosquitoes and horses throughout the 2000s but cases of EEEV in humans have not been reported so far. This study was designed to provide an assessment of possible EEEV human exposure by modelling environmental risk factors for EEEV in horses, identifying high-risk environments and mapping risk in the province of Quebec, Canada. According to logistic models, being located near wooded swamps was a risk factor for seropositivity or disease in horses [odds ratio (OR) 4·15, 95% confidence interval (CI) 1·16-14·8) whereas being located on agricultural lands was identified as protective (OR 0·75, 95% CI 0·62-0·92). A better understanding of the environmental risk of exposure to EEEV in Canada provides veterinary and public health officials with enhanced means to more effectively monitor the emergence of this public health risk and design targeted surveillance and preventive measures.

Surveillance for Lyme disease in Canada: 2009-2015.

OBJECTIVE: To summarize seven years of surveillance data for Lyme disease cases reported in Canada from 2009 to 2015. METHODS: We describe the incidence over time, seasonal and geographic distribution, demographic and clinical characteristics of reported Lyme disease cases. Logistic regression was used to explore differences between age groups, sex and year to better understand potential demographic risk factors for the occurrence of Lyme disease. RESULTS: The number of reported Lyme disease cases increased more than six-fold, from 144 in 2009 to 917 in 2015, mainly due to an increase in infections acquired in Canada. Most locally acquired cases were reported between May and November. An increase in incidence of Lyme disease was observed in provinces from Manitoba eastwards. This is consistent with our knowledge of range expansion of the tick vectors in this region. In the western provinces the incidence has remained low and stable. All cases reported by Alberta, Saskatchewan and Newfoundland and Labrador were acquired outside of the province, either elsewhere in Canada or abroad. There was a bimodal distribution for Lyme disease by age with peaks at 5-9 and 45-74 years of age. The most common presenting symptom was a single erythema migrans rash (74.2%) and arthritis (35.7%). Variations in the frequency of reported clinical manifestations were observed among age groups and years of study. CONCLUSION: Lyme disease incidence continues to increase in Canada as does the geographic range of ticks that carry the Lyme disease bacteria. Ongoing surveillance, preventive strategies as well as early disease recognition and treatment will continue to minimize the impact of Lyme disease in Canada.

Response to letter regarding article "How far north are migrant birds transporting the tick Ixodes scapularis in Canada? Insights from stable hydrogen isotope analyses of feathers".

In this letter we respond to correspondence and clarify the intent and importance of our article.

Evolutionary aspects of emerging Lyme disease in Canada.

In North America, Lyme disease (LD) is a tick-borne zoonosis caused by the spirochete bacterium Borrelia burgdorferi sensu stricto, which is maintained by wildlife. Tick vectors and bacteria are currently spreading into Canada and causing increasing numbers of cases of LD in humans and raising a pressing need for public health responses. There is no vaccine, and LD prevention depends on knowing who is at risk and informing them how to protect themselves from infection. Recently, it was found in the United States that some strains of B. burgdorferi sensu stricto cause severe disease, whereas others cause mild, self-limiting disease. While many strains occurring in the United States also occur in Canada, strains in some parts of Canada are different from those in the United States. We therefore recognize a need to identify which strains specific to Canada can cause severe disease and to characterize their geographic distribution to determine which Canadians are particularly at risk. In this review, we summarize the history of emergence of LD in North America, our current knowledge of B. burgdorferi sensu stricto diversity, its intriguing origins in the ecology and evolution of the bacterium, and its importance for the epidemiology and clinical and laboratory diagnosis of LD. We propose methods for investigating associations between B. burgdorferi sensu stricto diversity, ecology, and pathogenicity and for developing predictive tools to guide public health interventions. We also highlight the emergence of B. burgdorferi sensu stricto in Canada as a unique opportunity for exploring the evolutionary aspects of tick-borne pathogen emergence.

How far north are migrant birds transporting the tick Ixodes scapularis in Canada? Insights from stable hydrogen isotope analyses of feathers.

Lyme disease is emerging in Canada because of northward range expansion of the tick vector Ixodes scapularis. It is hypothesised that I. scapularis feeding on passerine birds migrating north in spring are important in founding new I. scapularis populations leading to northward range expansion. However, there are no studies on how far north I. scapularis may be carried, only inferences from passive tick surveillance. We used stable hydrogen isotope (δ(2)H) analysis of rectrices collected from northward migrating, I. scapularis-carrying, passerine birds captured in Canada to estimate how far north I. scapularis may be carried. Rectrices are usually grown close to breeding sites and their δ(2)H values reflect those in the environment, which vary strongly with latitude in North America. Passerines usually return to their breeding or natal sites so δ(2)H values of rectrices of northward migrating birds can identify the likely latitudinal bands of their intended destinations. In 2006 we analysed δ(2)H from rectrices of 73 I. scapularis-carrying birds captured at five migration monitoring stations, mainly from southern Ontario. Values of δ(2)H ranged from -33 to -124‰, suggesting 19/71 (26.7%) birds were destined for latitude band B (the most southerly part of Ontario), 40/71 (56.3%) birds were destined for band C (which extends from southern Ontario, Quebec and the Maritimes to southern James Bay) and 12/71 (16.9%) birds were destined for bands D and E (which extend from northern Ontario and Quebec into the southern Canadian Arctic). This indicates that many I. scapularis-carrying migratory birds in spring have destinations far north in Canada, including some farther north than the current region of climatic suitability for I. scapularis. These findings support the hypothesis that I. scapularis may continue to be spread north by spring migrating passerines. Some thrush species may be particularly implicated in far northward dispersion of I. scapularis.

Complex population structure of Borrelia burgdorferi in southeastern and south central Canada as revealed by phylogeographic analysis.

Lyme disease, caused by the bacterium Borrelia burgdorferi sensu stricto, is an emerging zoonotic disease in Canada and is vectored by the blacklegged tick, Ixodes scapularis. Here we used Bayesian analyses of sequence types (STs), determined by multilocus sequence typing (MLST), to investigate the phylogeography of B. burgdorferi populations in southern Canada and the United States by analyzing MLST data from 564 B. burgdorferi-positive samples collected during surveillance. A total of 107 Canadian samples from field sites were characterized as part of this study, and these data were combined with existing MLST data for samples from the United States and Canada. Only 17% of STs were common between both countries, while 49% occurred only in the United States, and 34% occurred only in Canada. However, STs in southeastern Ontario and southwestern Quebec were typically identical to those in the northeastern United States, suggesting a recent introduction into this region from the United States. In contrast, STs in other locations in Canada (the Maritimes; Long Point, Ontario; and southeastern Manitoba) were frequently unique to those locations but were putative descendants of STs previously found in the United States. The picture in Canada is consistent with relatively recent introductions from multiple refugial populations in the United States. These data thus point to a geographic pattern of populations of B. burgdorferi in North America that may be more complex than simply comprising northeastern, midwestern, and Californian groups. We speculate that this reflects the complex ecology and spatial distribution of key reservoir hosts.

Is there a risk of chikungunya transmission in Canada?

An outbreak of chikungunya virus infection is currently underway in the Caribbean, Central America and South America, and autochthonous (local or indigenous) transmission has occurred in the southeastern United States. The mosquito species known to transmit chikungunya-Aedes aegypti and Aedes albopictus-are not known to reside in Canada at this time. But how comfortable can we be that this situation will continue? Here we explore four key conditions that must be met for transmission of chikungunya within Canada. We conclude that not all of these conditions have been met and the risk of chikungunya transmission in most of Canada appears to be very low at present. The risk is slightly higher in warmer areas, such as southern British Columbia and isolated locations in south central and southeastern Canada. However, there are significant gaps in our knowledge and ongoing risk assessment, research and surveillance for vectors of chikungunya are indicated.

Surveillance for Lyme disease in Canada, 2009 to 2012.

OBJECTIVES: To summarize the first four years of national surveillance for Lyme disease in Canada from 2009 to 2012 and to conduct a preliminary comparison of presenting clinical manifestations in Canada and the United States. METHODS: The numbers and incidence of reported cases by province, month, year, age and sex were calculated. Logistic regression was used to examine trends over time. Acquisition locations were mapped and presenting clinical manifestations reported for jurisdictions where data was available. Variations by province, year, age and sex as well as presenting clinical symptoms were explored by logistic regression. An initial comparative analysis was made of presenting symptoms in Canada and the United States. RESULTS: The numbers of reported cases rose significantly from 144 in 2009 to 338 in 2012 (coefficient = 0.34, standard error = 0.07, P <0.05), mostly due to an increased incidence of infections acquired in Canada. More cases were classified as 'confirmed' (71.5%) than 'probable' (28.5%). Most cases occurred in locations where vector tick populations were known to be present. More men than women were affected (53.4% versus 46.6%), incidence was highest in adults aged 55 to 74 years and in children aged five to 14 years. Most cases (95%) were acquired from April to November. Of cases acquired in endemic areas, 39.7% presented with manifestations of early Lyme disease, while 60.3% had manifestations of disseminated Lyme disease. There were significant differences among age groups, sexes and provinces in the frequencies of reported clinical manifestations. The proportion of cases acquired in endemic areas presenting with early Lyme disease was lower than that reported in the US. CONCLUSION: Lyme disease incidence is increasing in Canada. Most cases are acquired where vector tick populations are spreading and this varies geographically within and among provinces. There is also variation in the frequency of age, season and presenting manifestations. The lower proportion of cases presenting with early Lyme disease in Canada compared with the US suggests lower awareness of early Lyme disease in Canada, but this requires further study.