SARS-CoV-2 viral titer measurements in Ontario, Canada wastewaters throughout the COVID-19 pandemic.

During the COVID-19 pandemic, the Province of Ontario, Canada, launched a wastewater surveillance program to monitor SARS-CoV-2, inspired by the early work and successful forecasts of COVID-19 waves in the city of Ottawa, Ontario. This manuscript presents a dataset from January 1, 2021, to March 31, 2023, with RT-qPCR results for SARS-CoV-2 genes and PMMoV from 107 sites across all 34 public health units in Ontario, covering 72% of the province's and 26.2% of Canada's population. Sampling occurred 2-7 times weekly, including geographical coordinates, serviced populations, physico-chemical water characteristics, and flowrates. In doing so, this manuscript ensures data availability and metadata preservation to support future research and epidemic preparedness through detailed analyses and modeling. The dataset has been crucial for public health in tracking disease locally, especially with the rise of the Omicron variant and the decline in clinical testing, highlighting wastewater-based surveillance's role in estimating disease incidence in Ontario.

Wastewater surveillance for earlier detection of seniors congregate living COVID-19 outbreaks in Peterborough, Ontario.

The coronavirus disease 2019 (COVID-19) pandemic has disproportionately affected seniors living in congregate living settings. The evolving surveillance context has led to novel use of wastewater surveillance to monitor levels of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in these settings. This study presents a pilot of upstream congregate living wastewater surveillance of SARS-CoV-2 for the detection of COVID-19 outbreaks and the effects of early public health interventions. We monitored localized wastewater SARS-CoV-2 levels from four congregate living settings March 15, 2021 to October 1, 2022 and correlated these levels with suspected and confirmed COVID-19 outbreaks determined by other methods. We identified five wastewater signals that correlated with confirmed outbreaks and three wastewater signals that did not correlate with subsequent outbreaks. In the five confirmed outbreaks, the wastewater signal was detected 2-10 days (median, five days) prior to confirmation of the outbreak by case testing. This pilot demonstrates upstream sampling for SARS-CoV-2 in wastewater may effectively detect outbreaks prior to their detection through symptomatic case testing and could support a balanced approach to outbreak response in congregate living settings, leading to increased wellbeing of these residents.

A review of sebum in mammals in relation to skin diseases, skin function, and the skin microbiome.

Diseases vary among and within species but the causes of this variation can be unclear. Immune responses are an important driver of disease variation, but mechanisms on how the body resists pathogen establishment before activation of immune responses are understudied. Skin surfaces of mammals are the first line of defense against abiotic stressors and pathogens, and skin attributes such as pH, microbiomes, and lipids influence disease outcomes. Sebaceous glands produce sebum composed of multiple types of lipids with species-specific compositions. Sebum affects skin barrier function by contributing to minimizing water loss, supporting thermoregulation, protecting against pathogens, and preventing UV-induced damage. Sebum also affects skin microbiome composition both via its antimicrobial properties, and by providing potential nutrient sources. Intra- and interspecific variation in sebum composition influences skin disease outcomes in humans and domestic mammal species but is not well-characterized in wildlife. We synthesized knowledge on sebum function in mammals in relation to skin diseases and the skin microbiome. We found that sebum composition was described for only 29 live, wild mammalian species. Sebum is important in dermatophilosis, various forms of dermatitis, demodicosis, and potentially white-nose syndrome. Sebum composition likely affects disease susceptibility, as lipid components can have antimicrobial functions against specific pathogens. It is unclear why sebum composition is species-specific, but both phylogeny and environmental effects may drive differences. Our review illustrates the role of mammal sebum function and influence on skin microbes in the context of skin diseases, providing a baseline for future studies to elucidate mechanisms of disease resistance beyond immune responses.

The bear necessities: A sensitive qPCR assay for bear DNA detection from bile and derived products to complement wildlife forensic enforcement.

Demand for bear bile, a prized component of traditional Asian medicines, threaten Asiatic and sun bear population sustainability. While laws exist to prevent poaching and trafficking of bear parts and derivatives, smuggling persists with demand extending to surrogate species, including American black bears (Ursus americanus). Mitochondrial DNA (mtDNA) sequencing can identify products putatively containing biological bear material but can be undermined by PCR inhibitors in bile and a lack of sensitivity at trace levels. Quantitative PCR (qPCR) assays can be used to distinguish between closely related target species, while concomitantly evaluating inhibition and false negative results in low quality/quantity DNA applications. Herein, we develop a multiplexed qPCR assay to detect and differentiate among bear species, including highly diluted bile samples mixed within liquors as common dilutants. The assay detects as little as 10 locus copies/reaction of bear DNA with 95% confidence, distinguishing among sun, Asiatic and American black bears. Demonstrating the sensitivity and applicability of this assay in context of current bile mixture recipes, dilutions of 1:5,000 bile with ethanol, red wine, and spirits, all yielded clear quantifiable detections, where our data suggests as little as 1 drop of bile per 750 mL bottle of alcohol would still exceed the limits of detection (e.g., 1:15000 dilution or <0.05 mL bile per 750 mL bottle). Overall, this study provides a rapid, sensitive, and specific test to identify and distinguish among bear species commonly used for bile production to aid wildlife enforcement applications.

The genome sequence of the Loggerhead sea turtle, Caretta caretta Linnaeus 1758.

We present a genome assembly of Caretta caretta (the Loggerhead sea turtle; Chordata, Testudines, Cheloniidae), generated from genomic data from two unrelated females. The genome sequence is 2.13 gigabases in size. The assembly has a busco completion score of 96.1% and N50 of 130.95 Mb. The majority of the assembly is scaffolded into 28 chromosomal representations with a remaining 2% of the assembly being excluded from these.

Metabarcoding of fecal pellets in wild muskox populations reveals negative relationships between microbiome and diet alpha diversity.

Microbiome diversity and diet composition concomitantly influence species health, fitness, immunity, and digestion. In environments where diet varies spatially and temporally, microbiome plasticity may promote rapid host adaptation to available resources. For northern ungulates in particular, metabarcoding of noninvasively collected fecal pellets presents unprecedented insights into their diverse ecological requirements and niches by clarifying the interrelationships of microbiomes, key to deriving nutrients, in context of altered forage availability in changing climates. Muskoxen (Ovibos moschatus) are Arctic-adapted species that experience fluctuating qualities and quantities of vegetation. Geography and seasonality have been noted to influence microbiome composition and diversity in muskoxen, yet it is unclear how their microbiomes intersect with diet. Following observations from other species, we hypothesized increasing diet diversity would result in higher microbiome diversity in muskoxen. We assessed diet composition in muskoxen using three common plant metabarcoding markers and explored correlations with microbiome data. Patterns of dietary diversity and composition were not fully concordant among the markers used, yet all reflected the primary consumption of willows and sedges. Individuals with similar diets had more similar microbiomes, yet in contrast to most literature, yielded negative relationships between microbiome and diet alpha diversity. This negative correlation may reflect the unique capacities of muskoxen to survive solely on high-fiber Arctic forage and provide insight into their resiliency to exploit changing dietary resources in a rapidly warming Arctic altering vegetation diversity.

Impact of Continuous Renal Replacement Therapy Initiation on Urine Output and Fluid Balance: A Multicenter Study.

INTRODUCTION: The effect of continuous renal replacement therapy (CRRT) on renal function is poorly understood. However, the initiation of CRRT may induce oliguria. We aimed to investigate the impact of CRRT commencement on urine output (UO). METHODS: This was a retrospective cohort study in two intensive care units. We included all patients who underwent CRRT and collected data on hourly UO and fluid balance before and after CRRT commencement. We performed an interrupted time series analysis using segmented regression to assess the relationship between CRRT commencement and UO. RESULTS: We studied 1,057 patients. Median age was 60.7 years (interquartile range [IQR], 48.3-70.6), and the median APACHE III was 95 (IQR, 76-115). Median time to CRRT was 17 h (IQR, 5-49). With start of CRRT, the absolute difference in mean hourly UO and mean hourly fluid balance was -27.0 mL/h (95% CI: -32.1 to -21.8; p value < 0.01) and - 129.3 mL/h (95% CI: -169.2 to -133.3), respectively. When controlling for pre-CRRT temporal trends and patient characteristics, there was a rapid post-initiation decrease in UO (-0.12 mL/kg/h; 95% CI: -0.17 to -0.08; p value < 0.01) and fluid balance (-78.1 mL/h; 95% CI: -87.9 to -68.3; p value < 0.01), which was sustained over the first 24 h of CRRT. Change in UO and fluid balance were only weakly correlated (r -0.29; 95% CI: -0.35 to -0.23; p value < 0.01). CONCLUSION: Commencement of CRRT was associated with a significant decrease in UO that could not be explained by extracorporeal fluid removal.

Tracing Eastern Wolf Origins From Whole-Genome Data in Context of Extensive Hybridization.

Southeastern Canada is inhabited by an amalgam of hybridizing wolf-like canids, raising fundamental questions regarding their taxonomy, origins, and timing of hybridization events. Eastern wolves (Canis lycaon), specifically, have been the subject of significant controversy, being viewed as either a distinct taxonomic entity of conservation concern or a recent hybrid of coyotes (C. latrans) and grey wolves (C. lupus). Mitochondrial DNA analyses show some evidence of eastern wolves being North American evolved canids. In contrast, nuclear genome studies indicate eastern wolves are best described as a hybrid entity, but with unclear timing of hybridization events. To test hypotheses related to these competing findings we sequenced whole genomes of 25 individuals, representative of extant Canadian wolf-like canid types of known origin and levels of contemporary hybridization. Here we present data describing eastern wolves as a distinct taxonomic entity that evolved separately from grey wolves for the past ∼67,000 years with an admixture event with coyotes ∼37,000 years ago. We show that Great Lakes wolves originated as a product of admixture between grey wolves and eastern wolves after the last glaciation (∼8,000 years ago) while eastern coyotes originated as a product of admixture between "western" coyotes and eastern wolves during the last century. Eastern wolf nuclear genomes appear shaped by historical and contemporary gene flow with grey wolves and coyotes, yet evolutionary uniqueness remains among eastern wolves currently inhabiting a restricted range in southeastern Canada.

Draft Genome Assembly of an Iconic Arctic Species: Muskox (Ovibos moschatus).

Muskoxen (Ovibos moschatus) are Arctic species within the Caprinae subfamily that are economically and culturally significant to northern Indigenous communities. Low genetic diversity from repeated genetic bottlenecks, coupled with the effects of Arctic warming (e.g., heat stress, changing forage, pathogen range expansions), present conservation concerns for this species. Reference genome assemblies enhance our ecological and evolutionary understanding of species (which in turn aid conservation efforts). Herein, we provide a full draft reference genome of muskox using Illumina Hiseq data and cross-species scaffolding. The final reference assembly yielded a genome of 2,621,890,883 bp in length, a scaffold N50 of ~13.2 million, and an annotation identifying ~19.3 k genes. The muskox genome assembly and annotation were then used to reconstruct a phylogenetic tree which estimated muskoxen diverged from other ungulate species~12 Mya. To gain insight into the demographic history of muskoxen we also performed pairwise sequentially Markovian coalescent (PSMC) that identified two population bottlenecks coinciding with major glaciation events contributing to the notoriously low genetic variation observed in muskoxen. Overall, this genome assembly provides a foundation for future population genomic studies, such as latitudinal analyses, to explore the capacity of muskoxen to adapt to rapidly changing environments.

Erratum to: Skin pH varies among bat species seasons and between wild and captive bats.

[This corrects the article DOI: 10.1093/conphys/coab088.].

Skin pH varies among bat species and seasons and between wild and captive bats.

Skin is a key aspect of the immune system in the defence against pathogens. Skin pH regulates the activity of enzymes produced both by hosts and by microbes on host skin, thus implicating pH in disease susceptibility. Skin pH varies inter- and intra-specifically and is influenced by a variety of intrinsic and extrinsic variables. Increased skin alkalinity is associated with a predisposition to cutaneous infections in humans and dogs, and inter-specific and inter-individual variation in skin pH is implicated in differential susceptibility to some skin diseases. The cutaneous pH of bats has not been characterized but is postulated to play a role in susceptibility to white-nose syndrome (WNS), a fungal infection that has decimated several Nearctic bat species. We used non-invasive probes to measure the pH of bat flight membranes in five species with differing susceptibility to WNS. Skin pH ranged from 4.67 to 8.59 and varied among bat species, geographic locations, body parts, age classes, sexes and seasons. Wild Eptesicus fuscus were consistently more acidic than wild Myotis lucifugus, Myotis leibii and Perimyotis subflavus. Juvenile bats had more acidic skin than adults during maternity season but did not differ during swarming. Male M. lucifugus were more acidic than females during maternity season, yet this trend reversed during swarming. Bat skin was more acidic in summer compared to winter, a pattern also reported in humans. Skin pH was more acidic in captive than wild E. fuscus, suggesting environmental impacts on skin pH. The pH of roosting substrates affects skin pH in captive bats and may partially explain seasonal patterns in wild bats that use different roost types across seasons. Future research on the influence of pH on microbial pathogenic factors and skin barrier function may provide valuable insights on new therapeutic targets for treating bat skin conditions.

Genetic structure of immunologically associated candidate genes suggests arctic rabies variants exert differential selection in arctic fox populations.

Patterns of local adaptation can emerge in response to the selective pressures diseases exert on host populations as reflected in increased frequencies of respective, advantageous genotypes. Elucidating patterns of local adaptation enhance our understanding of mechanisms of disease spread and the capacity for species to adapt in context of rapidly changing environments such as the Arctic. Arctic rabies is a lethal disease that largely persists in northern climates and overlaps with the distribution of its natural host, arctic fox. Arctic fox populations display little neutral genetic structure across their North American range, whereas phylogenetically unique arctic rabies variants are restricted in their geographic distributions. It remains unknown if arctic rabies variants impose differential selection upon host populations, nor what role different rabies variants play in the maintenance and spread of this disease. Using a targeted, genotyping-by-sequencing assay, we assessed correlations of arctic fox immunogenetic variation with arctic rabies variants to gain further insight into the epidemiology of this disease. Corroborating past research, we found no neutral genetic structure between sampled regions, but did find moderate immunogenetic structuring between foxes predominated by different arctic rabies variants. FST outliers associated with host immunogenetic structure included SNPs within interleukin and Toll-like receptor coding regions (IL12B, IL5, TLR3 and NFKB1); genes known to mediate host responses to rabies. While these data do not necessarily reflect causation, nor a direct link to arctic rabies, the contrasting genetic structure of immunologically associated candidate genes with neutral loci is suggestive of differential selection and patterns of local adaptation in this system. These data are somewhat unexpected given the long-lived nature and dispersal capacities of arctic fox; traits expected to undermine local adaptation. Overall, these data contribute to our understanding of the co-evolutionary relationships between arctic rabies and their primary host and provide data relevant to the management of this disease.

The role of a mechanistic host in maintaining arctic rabies variant distributions: Assessment of functional genetic diversity in Alaskan red fox (Vulpes vulpes).

Populations are exposed to different types and strains of pathogens across heterogeneous landscapes, where local interactions between host and pathogen may present reciprocal selective forces leading to correlated patterns of spatial genetic structure. Understanding these coevolutionary patterns provides insight into mechanisms of disease spread and maintenance. Arctic rabies (AR) is a lethal disease with viral variants that occupy distinct geographic distributions across North America and Europe. Red fox (Vulpes vulpes) are a highly susceptible AR host, whose range overlaps both geographically distinct AR strains and regions where AR is absent. It is unclear if genetic structure exists among red fox populations relative to the presence/absence of AR or the spatial distribution of AR variants. Acquiring these data may enhance our understanding of the role of red fox in AR maintenance/spread and inform disease control strategies. Using a genotyping-by-sequencing assay targeting 116 genomic regions of immunogenetic relevance, we screened for sequence variation among red fox populations from Alaska and an outgroup from Ontario, including areas with different AR variants, and regions where the disease was absent. Presumed neutral SNP data from the assay found negligible levels of neutral genetic structure among Alaskan populations. The immunogenetically-associated data identified 30 outlier SNPs supporting weak to moderate genetic structure between regions with and without AR in Alaska. The outliers included SNPs with the potential to cause missense mutations within several toll-like receptor genes that have been associated with AR outcome. In contrast, there was a lack of genetic structure between regions with different AR variants. Combined, we interpret these data to suggest red fox populations respond differently to the presence of AR, but not AR variants. This research increases our understanding of AR dynamics in the Arctic, where host/disease patterns are undergoing flux in a rapidly changing Arctic landscape, including the continued northward expansion of red fox into regions previously predominated by the arctic fox (Vulpes lagopus).

Transcriptional host-pathogen responses of Pseudogymnoascus destructans and three species of bats with white-nose syndrome.

Understanding how context (e.g., host species, environmental conditions) drives disease susceptibility is an essential goal of disease ecology. We hypothesized that in bat white-nose syndrome (WNS), species-specific host-pathogen interactions may partly explain varying disease outcomes among host species. We characterized bat and pathogen transcriptomes in paired samples of lesion-positive and lesion-negative wing tissue from bats infected with Pseudogymnoascus destructans in three parallel experiments. The first two experiments analyzed samples collected from the susceptible Nearctic Myotis lucifugus and the less-susceptible Nearctic Eptesicus fuscus, following experimental infection and hibernation in captivity under controlled conditions. The third experiment applied the same analyses to paired samples from infected, free-ranging Myotis myotis, a less susceptible, Palearctic species, following natural infection and hibernation (n = 8 sample pairs/species). Gene expression by P. destructans was similar among the three host species despite varying environmental conditions among the three experiments and was similar within each host species between saprophytic contexts (superficial growth on wings) and pathogenic contexts (growth in lesions on the same wings). In contrast, we observed qualitative variation in host response: M. lucifugus and M. myotis exhibited systemic responses to infection, while E. fuscus up-regulated a remarkably localized response. Our results suggest potential phylogenetic determinants of response to WNS and can inform further studies of context-dependent host-pathogen interactions.

High prevalence of subclinical frog virus 3 infection in freshwater turtles of Ontario, Canada.

Ranaviruses have been associated with chelonian mortality. In Canada, the first two cases of ranavirus were detected in turtles in 2018 in Ontario, although a subsequent survey of its prevalence failed to detect additional positive cases. To confirm the prevalence of ranavirus in turtles in Ontario, we used a more sensitive method to investigate if lower level persistent infection was present in the population. Here we report results via a combination of qPCR, PCR, Sanger sequencing and genome sequencing from turtles from across Ontario, with no clinical signs of illness. We found 2 positives with high viral load and 5 positives with low viral load. Histopathology found subtle histological changes. DNA sequences identified two types of frog virus 3 (FV3), and genome sequencing identified a ranavirus similar to wild-type FV3. Our results show that the virus has been present in Ontario's turtles as subclinical infections.

Geography, seasonality, and host-associated population structure influence the fecal microbiome of a genetically depauparate Arctic mammal.

The Canadian Arctic is an extreme environment with low floral and faunal diversity characterized by major seasonal shifts in temperature, moisture, and daylight. Muskoxen (Ovibos moschatus) are one of few large herbivores able to survive this harsh environment. Microbiome research of the gastrointestinal tract may hold clues as to how muskoxen exist in the Arctic, but also how this species may respond to rapid environmental changes. In this study, we investigated the effects of season (spring/summer/winter), year (2007-2016), and host genetic structure on population-level microbiome variation in muskoxen from the Canadian Arctic. We utilized 16S rRNA gene sequencing to characterize the fecal microbial communities of 78 male muskoxen encompassing two population genetic clusters. These clusters are defined by Arctic Mainland and Island populations, including the following: (a) two mainland sampling locations of the Northwest Territories and Nunavut and (b) four locations of Victoria Island. Between these geographic populations, we found that differences in the microbiome reflected host-associated genetic cluster with evidence of migration. Within populations, seasonality influenced bacterial diversity with no significant differences between years of sampling. We found evidence of pathogenic bacteria, with significantly higher presence in mainland samples. Our findings demonstrate the effects of seasonality and the role of host population-level structure in driving fecal microbiome differences in a large Arctic mammal.

Frog Virus 3 Genomes Reveal Prevalent Recombination between Ranavirus Lineages and Their Origins in Canada.

Ranaviruses are pathogens associated with the decline of amphibian populations across much of their distribution. In North America, frog virus 3 (FV3) is a widely distributed pathogen with wild populations of amphibians harboring different lineages and putative recombinants between FV3 and common midwife toad virus (CMTV). These recombinants have higher pathogenicity, and CMTV-derived genes associated with virulence are reported in wild strains in Canada. However, while FV3 is linked to amphibian die-offs in North America, CMTVs have been reported only in commercial frog farms in North America. We sequenced complete genomes of 18 FV3 isolates from three amphibian species to characterize genetic diversity of the lineages in Canada and infer possible recombinant regions. The 18 FV3 isolates displayed different signals of recombination, varying from none to interspersed recombination with previously isolated CMTV-like viruses. In general, most recombination breakpoints were located within open reading frames (ORFs), generating new ORFs and proteins that were a mixture between FV3 and CMTV. A combined spatial and temporal phylogeny suggests the presence of the FV3 lineage in Canada is relatively contemporary (<100 years), corroborating the hypothesis that both CMTV- and FV3-like viruses spread to North America when the international commercial amphibian trade started. Our results highlight the importance of pathogen surveillance and viral dynamics using full genomes to more clearly understand the mechanisms of disease origin and spread.IMPORTANCE Amphibian populations are declining worldwide, and these declines have been linked to a number of anthropogenic factors, including disease. Among the pathogens associated with amphibian mortality, ranaviruses have caused massive die-offs across continents. In North America, frog virus 3 (FV3) is a widespread ranavirus that can infect wild and captive amphibians. In this study, we sequenced full FV3 genomes isolated from frogs in Canada. We report widespread recombination between FV3 and common midwife toad virus (CMTV). Phylogenies indicate a recent origin for FV3 in Canada, possibly as a result of international amphibian trade.

ONRAB® oral rabies vaccine is shed from, but does not persist in, captive mammals.

ONRAB® is a human adenovirus rabies glycoprotein recombinant vaccine developed to control rabies in wildlife. To support licensing and widespread use of the vaccine, safety studies are needed to assess its potential residual impact on wildlife populations. We examined the persistence of the ONRAB® vaccine virus in captive rabies vector and non-target mammals. This research complements work on important rabies vector species (raccoon, striped skunk, and red fox) but also adds to previous findings with the addition of some non-target species (Virginia opossum, Norway rats, and cotton rats) and a prolonged period of post vaccination monitoring (41 days). Animals were directly inoculated orally with the vaccine and vaccine shedding was monitored using quantitative real-time PCR applied to oral and rectal swabs. ONRAB® DNA was detected in both oral and rectal swabs from 6 h to 3 days post-inoculation in most animals, followed by a resurgence of shedding between days 17 and 34 in some species. Overall, the duration over which ONRAB® DNA was detectable was shorter for non-target mammals, and by day 41, no animal had detectable DNA in either oral or rectal swabs. All target species, as well as cotton rats and laboratory-bred Norway rats, developed robust humoral immune responses as measured by competitive ELISA, with all individuals being seropositive at day 31. Similarly, opossums showed good response (89% seropositive; 8/9), whereas only one of nine wild caught Norway rats was seropositive at day 31. These results support findings of other safety studies suggesting that ONRAB® does not persist in vector and non-target mammals exposed to the vaccine. As such, we interpret these data to reflect a low risk of adverse effects to wild populations following distribution of ONRAB® to control sylvatic rabies.

Urban and Rural Spatial Delineations in Blow Fly Species (Diptera: Calliphoridae) Across Canada: Implications for Forensic Entomology.

Blow fly (Diptera: Calliphoridae) larvae are commonly used in forensic cases to determine postmortem intervals using development rates and successional changes in community composition. Studies are conducted from different regions to provide these data. We wanted to know how widely applicable these data are. We examined whether urbanized landscapes have distinct urban blow fly communities or whether the community composition in urbanized areas is simply a variation of that found in the surrounding habitat or ecozone. Using liver baited traps, we sampled 7,272 flies from 32 sites across Canada and used mapping analysis to assess urban and rural landcover classifications, and compared urban and rural species abundance and composition. Blow fly species communities from urban areas across Canada were made up of similar species and differed from the communities found in nearby rural sites. Trapping at rural sites caught more blow flies compared with urban sites (mean flies/site 59.5 and 12.4). Of the 14 species caught, 8 were caught at urban sites, 61% of these being Cynomya cadaverina Robineau-Desvoidy, 14% Phormia regina Meigen, and 11% Lucilia sericata (Meigen). In rural sites, all 14 species were caught, 41% of specimens caught were P. regina, 21% C. cadaverina, 10% Calliphora vomitoria (Linnaeus), with only 4% L. sericata. These data suggest that regional studies are appropriate for forensic entomology applications in urban landscapes, given the similar trends across Canada, less so for wilderness or rural landscapes.

Technical note: A rapid, non-invasive method for measuring live or preserved insect specimens using digital image analysis.

The measurement of insects is an important component of many entomological applications, including forensic evidence, where larvae size is used as a proxy for developmental stage, and hence time since colonization/death. Current methods for measuring insects are confounded by varying preservation techniques, biased and non-standardized measurements, and often a lack of sample size given practical constraints. Towards enhanced accuracy and precision in measuring live insects to help avoid these variables, and that allows for different measurements to be analyzed, we developed a non-invasive, digital method using widely available free analytical software to measure live blow fly larvae. Using crime scene photographic equipment currently standard in investigation protocols, we measured the live length of 282 Phormia regina larvae. Repeated measurements of maggots, for all instars, were performed for several orientations and images. Most accurate measurements were obtained when maggots were oriented in their natural full extension. Killed specimens resulted in greater length measurements (Mean 1.79 ±â€¯1.11 mm) when compared to live length. Herein, we report a technically simple, fast, and accurate measurement technique adapted for field and lab-based measurements, as well as, a simple linear equation for conversion of live length to standard killed length measurements. We propose this method be utilized for the standardization of forensic entomological evidence collection and development model creation.