Figure 3 - uploaded by Frances Cassirer
Content may be subject to copyright.
Spatiotemporal distribution of the 16S-23S intergenic spacer (IGS) genotypes of Mycoplasma ovipneumoniae in the Hells Canyon bighorn sheep metapopulation in Washington, Oregon, and Idaho, USA in a) 1986 and 1992 b) 1995–2013; and c) 2014–2015. Each colored marker represents one strain type, pie charts display strain types within populations and are scaled by sample size. Pie charts containing >1 color indicate that 2 strain types were present in a population during that time interval but not necessarily detected in the same year. Gray shaded polygons denote bighorn sheep populations: AS = Asotin, BB = Black Butte, BC = Big Canyon, BR = Bear Creek, IM = Imnaha, LH = Lower Hells Canyon, LM = Lookout Mountain, MU = Muir, MV = Mountain View, MY = Myers Creek, RB = Redbird, SM = Sheep Mountain, TU = Tucannon, UO = Upper Hells Canyon, Oregon, UI = Upper Hells Canyon Idaho.
Source publication
Transmission of pathogens commonly carried by domestic sheep and goats poses a serious threat to bighorn sheep (Ovis canadensis) populations. All-age pneumonia die-offs usually ensue, followed by asymptomatic carriage of Mycoplasma ovipneumoniae by some of the survivors. Lambs born into these chronically infected populations often succumb to pneumo...
Similar publications
Infectious disease contributed to historical declines and extirpations of bighorn sheep (Ovis canadensis) in North America and continues to impede population restoration and management. Reports of pneumonia outbreaks in free-ranging bighorn sheep following contact with domestic sheep have been validated by the results of 13 captive commingling expe...
Citations
... The genetic diversity of M. ovipneumoniae is high in domestic sheep, indicating their role as a signi cant reservoir and source of infection, while in BHS, it is low, suggesting spillover as the primary source of transmission. Indeed, ancestral state reconstruction from MLST sequences con rmed domestic sheep as the primary source of infection for BHS, emphasizing the importance of strain typing to map transmission dynamics (4). In BHS, an initial outbreak of fatal bronchopneumonia is often followed by recurring fatal outbreaks in lambs. ...
... Recurrent outbreaks have been observed from 2 to 15 years after the initial spillover (2,(5)(6)(7). Recent evidence suggests there may be no cross-strain immunity, leaving surviving animals susceptible to infection (4,8). ...
... DNA-based strain typing is used to document the invasion, persistence, and transmission of M. ovipneumoniae in these populations (7). A previously developed multilocus sequence typing (MLST) scheme targeting four elements, the 16-23S intergenic spacer region (IGS), 16S rRNA region (LM), RNA polymerase β-subunit gene (rpoB), and DNA gyrase subunit-β gene (gyrB), has demonstrated strong differential typing capability in over 600 samples and 270 strain types (1,4). Owing to the rapid emergence of new strains and the extensive diversity of novel types, creating a conventional database of alternative alleles is impractical (1,8,10). ...
Background
Spillover events of Mycoplasma ovipneumoniae have devastating effects on wild bighorn sheep populations. Multilocus sequence typing (MLST), a common method for tracking bacterial lineages, is used to monitor spillover events and the spread of M. ovipneumoniae between populations. Most work involving M. ovipneumoniae typing has used Sanger sequencing, however, this technology is time consuming, expensive, and is not well suited to efficient batch sample processing. Our study aimed to develop and validate a workflow for multilocus sequence typing of M. ovipneumoniae using Nanopore Rapid Barcoding sequencing and multiplex PCR. We compare the workflow with Nanopore Native Barcoding library preparation and Illumina MiSeq amplicon protocols to determine the most accurate and cost-effective method for sequencing multiplex amplicons.
Results
A multiplex PCR was optimized for four housekeeping genes of M. ovipneumoniae using archived DNA samples from wild sheep. Sequences recovered from Nanopore Rapid Barcoding correctly identified all MLST types with the shortest total workflow time, and lowest cost per sample when compared to Nanopore Native Barcoding, and Illumina MiSeq methods.
Conclusion
Our proposed workflow serves as a convenient and effective diagnostic method for strain typing of M. ovipneumoniae, and could be applied to other bacterial MLST schemes. The workflow is suitable for diagnostic settings where reduced hands-on time, cost and multiplexing capabilities are important.
... These pathogens can be present in both bighorn and domestic sheep populations, and contact between bighorn and domestic sheep has been identified as a risk factor for bighorn sheep populations [15][16][17]. Current diagnostics to identify these bacterial pathogens in bighorn sheep include culture and PCR [18], and limited multilocus sequence typing (MLST) [19]. More robust and standardized diagnostics could be useful to provide comprehensive and consistent data for management decisions that affect both bighorn and domestic sheep stakeholders. ...
... We used published MLST schemes (pubMLST [26]) for M. haemolytica and P. multocida as a basis for our MLST approach to the Pasteurellaceae bacteria, with modifications based on preliminary results using these primers, and other literature referencing housekeeping genes in Pasteurellaceae bacteria [27]. Primers previously used for M. ovipneumoniae MLST by others [19,28,29] were included in our MLST scheme for backward comparisons, although this approach targets only three genes (gyrB, rpoB, and 16S rRNA) as well as an intergenic spacer region [19]. We expanded this MLST approach to provide a total of eight targets. ...
... We used published MLST schemes (pubMLST [26]) for M. haemolytica and P. multocida as a basis for our MLST approach to the Pasteurellaceae bacteria, with modifications based on preliminary results using these primers, and other literature referencing housekeeping genes in Pasteurellaceae bacteria [27]. Primers previously used for M. ovipneumoniae MLST by others [19,28,29] were included in our MLST scheme for backward comparisons, although this approach targets only three genes (gyrB, rpoB, and 16S rRNA) as well as an intergenic spacer region [19]. We expanded this MLST approach to provide a total of eight targets. ...
Respiratory disease is responsible for dramatic population declines in bighorn sheep ( Ovis canadensis) , and respiratory pathogen diagnostics contribute to the management of bighorn populations. To create a comprehensive and consistent approach to bighorn sheep respiratory diagnostics, we created a culture-independent assay to detect and strain type Mannheimia haemolytica , Bibersteinia trehalosi , Pasteurella multocida , and Mycoplasma ovipneumoniae . The assay also detects and characterizes the Pasteurellaceae leukotoxin A gene, and broadly assesses the bacterial composition of each sample based on 16S rRNA sequences. The assay is based on a three-step approach: 1) Multiplex PCR to amplify targets including eight loci for each bacterial species, the Pasteurellaceae lktA gene, and the 16S rRNA gene 2) Library preparation, barcoding, and short-read Illumina sequencing to determine the genetic sequences of each target, and 3) Bioinformatics in the form of automated software to analyze genetic sequences. The assay was designed to assess shared pathogens between domestic and bighorn sheep, but could be useful for many applications in bighorn sheep respiratory disease research and management.
... In a study by Kamath and others, 187 genetically different strains of M. ovipneumoniae were identified in 207 domestic sheep, with 77% of individual sheep having unique strains [7]. In bighorn sheep and domestic sheep, immunity to M. ovipneumoniae appears to be strain specific [9,23]. Oxytetracycline, a bacteriostatic antibiotic, administered daily appeared to produce a decline in M. ovipneumoniae DNA but failed to eliminate the bacteria DNA. ...
... Although animals that survive an initial outbreak appear to maintain immunity for some years, juveniles are naïve, and mortality within the first 4-6 months after birth frequently is high for several years following exposure (Cassirer and Sinclair, 2007;Smith et al., 2014), leading to consistent suppression of recruitment (Butler et al., 2018). Moreover, adult immunity is strain-specific (Cassirer et al., 2017). Thus, continued contact with domestic sheep, or with other bighorn sheep infected with differing strains of M. ovipneumoniae, can result in a new cycle of all-age mortality, followed by suppression of juvenile recruitment that slows, or even prevents, population growth (Manlove et al., 2016;Cassirer et al., 2017), as well as reduction in size and growth rate of sexually selected traits (e.g., horn length; Martin et al., 2022). ...
... Moreover, adult immunity is strain-specific (Cassirer et al., 2017). Thus, continued contact with domestic sheep, or with other bighorn sheep infected with differing strains of M. ovipneumoniae, can result in a new cycle of all-age mortality, followed by suppression of juvenile recruitment that slows, or even prevents, population growth (Manlove et al., 2016;Cassirer et al., 2017), as well as reduction in size and growth rate of sexually selected traits (e.g., horn length; Martin et al., 2022). Virulence appears to differ among strains of M. ovipneumoniae (Johnson et al., 2022), and other factors (hostrelated or environmental) likewise also cause epidemics to have widely varying effects on adult and juvenile survival . ...
... Therefore, when re-establishing bighorn sheep across landscapes, a metapopulation is the ideal distribution to enable these processes to approximate demographics observed in largely intact systems by creating local populations that could be linked by natural dispersal. Yet, that same inter-connectedness likewise provides simultaneous opportunities for the spread of pathogens by contact, as observed in Rocky Mountain bighorn sheep in the Hells Canyon system (Cassirer et al., 2017). One strategy to ameliorate this tradeoff would be to establish systems of populations potentially linked by Frontiers in Ecology and Evolution 11 frontiersin.org ...
Mammals are imperiled worldwide, primarily from habitat loss or modification, and exhibit downward trends in their populations and distributions. Likewise, large-bodied herbivores have undergone a collapse in numbers and are at the highest extinction risk of all mammals. Bighorn sheep (Ovis canadensis) are among those large-bodied herbivores that possess a slow-paced life history, suffer from debilitating diseases, and have experienced range contractions across their historical distribution since the late 1800s. Translocations and reintroductions of these mountain ungulates are key aspects of restoration and often are used to re-establish populations in historical habitat or to supplement declining herds. Millions of US dollars and much effort by state and federal natural resource agencies, as well as public and private organizations, have been expended to restore bighorn sheep. Despite those efforts, translocated populations of bighorn sheep have not always been successful. We assessed restoration of bighorn sheep to provide insights in the context of conservation of populations of bighorn sheep, because this management tool is a frequently used to re-establish populations. We focused briefly on past efforts to restore bighorn sheep populations and followed with updates on the value of habitat enhancements, genetic issues, the importance of ecotypic or phenotypic adaptations when restoring populations, predation, and disease transmission. We also raised issues and posed questions that have potential to affect future decisions regarding the restoration of bighorn sheep. This information will help conservationists improve the success of conserving these iconic large mammals.
... Swabs were tested via polymerase chain reaction (PCR) to detect M. ovipneumoniae specific DNA sequences by Washington Animal Disease Diagnostic Laboratory (WADDL; Pullman, WA). Strain typing consisted of multi-locus sequence typing based on partial DNA sequences of the 16S-23S intergenic spacer region, the 16S ribosomal subunit, and RNA polymerase B and gyrase B genes, as described in Cassirer et al. [53]. Additionally, we aged animals and classified them as less than or greater than 5 years based on horn growth (i.e. ...
Animal movements among habitat patches or populations are important for maintaining long-term genetic and demographic viability, but connectivity may also facilitate disease spread and persistence. Understanding factors that influence animal movements is critical to understanding potential transmission risk and persistence of communicable disease in spatially structured systems. We evaluated effects of sex, age and Mycoplasma ovipneumoniae infection status at capture on intermountain movements and seasonal movement rates observed in desert bighorn sheep (Ovis canadensis nelsoni) using global positioning system collar data from 135 individuals (27 males, 108 females) in 14 populations between 2013 and 2018, following a pneumonia outbreak linked to the pathogen M. ovipneumoniae in the Mojave Desert, California, USA. Based on logistic regression analysis, intermountain movements were influenced by sex, age and most notably, infection status at capture: males, older animals and uninfected individuals were most likely to make such movements. Based on multiple linear regression analysis, females that tested positive for M. ovipneumoniae at capture also had lower mean daily movement rates that were further influenced by season. Our study provides empirical evidence of a pathogenic infection decreasing an individual's future mobility, presumably limiting that pathogen's ability to spread, and ultimately influencing transmission risk within a spatially structured system.
... The long-term population-level effect of exposure to Movi varies from full recovery to functional or local extinction (i.e., mortality of ≥90% of the population; Singer et al., 2000;Sells et al., 2015). Most surviving bighorn sheep clear Movi infections with immunity restricted to that strain Cassirer et al., 2017). Nonetheless, some become chronic carriers, and despite apparent immunity from clinical disease, do not clear the infection, consistently test positive for Movi carriage and pose infection risk to other individuals Garwood et al., 2020). ...
... Individual host response, resulting dynamics and negative effects of disease within bighorn sheep populations is shaped by heterogeneities in Movi strain-specific virulence, exposure dosages, and prior Movi exposure histories (Cassirer et al., 2017(Cassirer et al., , 2018. Describing the effects of exposure history and adaptive immune responses on disease severity and persistence, addresses an important knowledge gap regarding host-pathogen dynamics in wildlife. ...
... Specifically, sheep from the Hells Canyon subpopulations: Asotin (n = 9), Lostine (n = 4), and Sheep Mountain (n = 2) of Washington, Oregon, and Idaho, respectively, had been exposed to the HC-404 strain of Movi. Sheep transported in October 2014 from the Black Butte herd (n = 8) of Washington within Hells Canyon carried the HC-404 strain from 1995 until a novel Movi strain, BB-393, was detected in 2014 (Cassirer et al., 2017). We refer to this strain exposure as BB-393/HC-404. ...
Introduction
Throughout their range, bighorn sheep (Ovis canadensis) populations have seen significant disease-associated declines. Unfortunately, understanding of the underlying epidemiological processes driving the disease dynamics in this species has hindered conservation efforts aimed at improving the health and long-term viability of these populations. Individual response to pathogen exposure emerges from dynamic interactions between competing evolutionary processes within the host and pathogen. The host’s adaptive immune system recognizes pathogens and mounts a defensive response. Pathogens have evolved strategies to overcome adaptive immune defenses including maintaining high genetic diversity through rapid evolution. The outcomes of this evolutionary warfare determine the success of pathogen invasion of the host and ultimately the success of conservation efforts.
Methods
During an epizootic dominated by a single strain, we explore these host-pathogen dynamics by examining the variation in effects of pathogen invasion on captive bighorn sheep with differing histories of exposure to genetically diverse strains of Mycoplasma ovipneumoniae (Movi). We monitored clinical signs of disease and sampled animals and their environment to detect spread of Movi among 37 bighorn sheep separated into nine pens based on known exposure histories.
Results
We documented Movi transmission within and across pens and we detected Movi DNA in air, water, and invertebrate samples. Higher levels of antibody to Movi prior to the epizootic were associated with a lower likelihood of presenting clinical signs of pneumonia. Nonetheless, higher antibody levels in symptomatic individuals were associated with more severe progressive disease, increased probability and speed of pneumonia-induced mortality, and reduced likelihood of returning to a healthy state. Bighorn sheep with previous exposure to a strain other than the predominant epizootic strain were more likely to recover.
Discussion
Our results indicate that Movi-strain variability was sufficient to overwhelm the adaptive host immunological defenses. This outcome indicates, in free-ranging herds, past exposure is likely insufficient to protect bighorn sheep from infection by new Movi strains, although it influences the progression of disease and recovery within the herd. Therefore, given Movi-strain variability and the lack of immunological protection from past exposure, focusing management efforts on minimizing the introduction of Movi into bighorn herds, through separation of domestic and bighorn sheep and avoidance of management activities that create commingling of bighorn sheep carrying differing Movi strains, will likely be the most effective approach for reducing the effects of disease and achieving bighorn sheep conservation goals.
... Whole-genome single-nucleotide-polymorphism (SNP) phylogeny demonstrated clustering of strains based on their host species, regardless of their geographical origin, and suggested co-evolution of M. ovipneumoniae with its host species [19]. Recent studies, using multi-locus sequence typing (MLST), in the USA, also showed that domestic goat strains were distinct from those in domestic sheep [81,82], which had high genetic diversity. However, strains from both domestic ruminant species were found in bighorn sheep [82]. ...
Mycoplasma ovipneumoniae, a well-established respiratory pathogen of sheep and goats, has gained increased importance recently because of its detection in wild ruminants including members of the Cervidae family. Despite its frequent isolation from apparently healthy animals, it is responsible for outbreaks of severe respiratory disease which are often linked to infections with multiple heterologous strains. Furthermore, M. ovipneumoniae is characterized by an unusually wide host range, a high degree of phenotypic, biochemical, and genomic heterogeneity, and variable and limited growth in mycoplasma media. A number of mechanisms have been proposed for its patho-genicity, including the production of hydrogen peroxide, reactive oxygen species production, and toxins. It shows wide metabolic activity in vitro, being able to utilize substrates such as glucose, pyruvate, and isopropanol; these patterns can be used to differentiate strains. Treatment of infections in the field is complicated by large variations in the susceptibility of strains to antimicrobials, with many showing high minimum inhibitory concentrations. The lack of commercially available vaccines is probably due to the high cost of developing vaccines for diseases in small ruminants not presently seen as high priority. Multiple strains found in affected sheep and goats may also hamper the development of effective vaccines. This review summarizes the current knowledge and identifies gaps in research on M. ovipneumoniae, including its epidemiology in sheep and goats, pathology and clinical presentation, infection in wild ruminants, virulence factors, metabolism, comparative genomics, genotypic variability, phenotypic variability, evolutionary mechanisms, isolation and culture, detection and identification, antimicrobial susceptibility, variations in antimicrobial susceptibility profiles, vaccines, and control.
... We then calculated an expected value prediction for the number of mountain goat populations achieving trend objectives 1 generation from 2020 under each AMS as the weighted mean of this distribution.Disease risks to mountain goat and bighorn sheep populationsWe measured performance of fundamental objectives 3 and 4 using a value function for the number of bighorn sheep or mountain goat herds with pathogen transmission risks, respectively, at present or following management actions. The causes of respiratory disease in alpine ungulates are likely polymicrobial or strain-specific(Cassirer et al. 2017(Cassirer et al. , 2018, and mixing microbial communities during translocations may initiate a pneumonia epidemic. Our risk value function represented risk associated with pneumonia outbreaks arising from pathogen presence or mixing, notwithstanding other factors that may affect disease expression(Gullis and Fujino 2015, Sells et al. 2015, Butler et al. 2018:KP M NM Risk value = + ( × ),where KP was the predicted number of populations(either species) harboring pneumonia-associated pathogens, NM was the number of populations mixed through translocations under each AMS, and M was an expression of risk tolerance for uncertainty surrounding consequences of mixing pathogens through translocations. ...
Climate change is expected to disproportionately affect species occupying ecosystems with relatively hard boundaries, such as alpine ecosystems. Wildlife managers must identify actions to conserve and manage alpine species into the future, while considering other issues and uncertainties. Climate change and respiratory pathogens associated with widespread pneumonia epidemics in bighorn sheep (Ovis canadensis) may negatively affect mountain goat (Oreamnos americanus) populations. Mountain goat demographic and population data are challenging to collect and sparsely available, making population management decisions difficult. We developed predictive models incorporating these uncertainties and analyzed results within a structured decision making framework to make management recommendations and identify priority information needs in Montana, USA. We built resource selection models to forecast occupied mountain goat habitat and account for uncertainty in effects of climate change, and a Leslie matrix projection model to predict population trends while accounting for uncertainty in population demographics and dynamics. We predicted disease risks while accounting for uncertainty about presence of pneumonia pathogens and risk tolerance for mixing populations during translocations. Our analysis predicted that new introductions would produce more area occupied by mountain goats at mid‐century, regardless of the effects of climate change. Population augmentations, carnivore management, and harvest management may improve population trends, although this was associated with considerable uncertainty. Tolerance for risk of disease transmission affected optimal management choices because translocations are expected to increase disease risks for mountain goats and sympatric bighorn sheep. Expected value of information analyses revealed that reducing uncertainty related to population dynamics would affect the optimal choice among management strategies to improve mountain goat trends. Reducing uncertainty related to the presence of pneumonia‐associated pathogens and consequences of mixing microbial communities should reduce disease risks if translocations are included in future management strategies. We recommend managers determine tolerance for disease risks associated with translocations that they and constituents are willing to accept. From this, an adaptive management program can be constructed wherein a portfolio of management actions are chosen based on risk tolerance in each population range, combined with the amount that uncertainty is reduced when paired with monitoring, to ultimately improve achievement of fundamental objectives. Mountain goat management in Montana is complicated by climate, demographic, and disease uncertainty and multiple, competing fundamental objectives. Addressing disease risk tolerance within an adaptive management program to reduce demographic and disease uncertainty should improve achievement of fundamental objectives for mountain goat management in Montana.
... Consult your local agency veterinarian for potential pathogens in your study area. Swab the nasal cavity or the mouth for bacteria that are associated with pneumonia(Cassirer et al., 2017) and tuberculosis(Barasona et al., 2017). The recent development of molecular methods provides opportunities to collect DNA as well, which in combination with results from the mortality site investigations, can confirm the presence of specific predators (seeMumma et al., 2014 for a general description of DNA protocols). ...
Mortality site investigations of telemetered wildlife are important for cause‐specific survival analyses and understanding underlying causes of observed population dynamics. Yet, eroding ecoliteracy and a lack of quality control in data collection can lead researchers to make incorrect conclusions, which may negatively impact management decisions for wildlife populations. We reviewed a random sample of 50 peer‐reviewed studies published between 2000 and 2019 on survival and cause‐specific mortality of ungulates monitored with telemetry devices. This concise review revealed extensive variation in reporting of field procedures, with many studies omitting critical information for the cause of mortality inference. Field protocols used to investigate mortality sites and ascertain the cause of mortality are often minimally described and frequently fail to address how investigators dealt with uncertainty. We outline a step‐by‐step procedure for mortality site investigations of telemetered ungulates, including evidence that should be documented in the field. Specifically, we highlight data that can be useful to differentiate predation from scavenging and more conclusively identify the predator species that killed the ungulate. We also outline how uncertainty in identifying the cause of mortality could be acknowledged and reported. We demonstrate the importance of rigorous protocols and prompt site investigations using data from our 5‐year study on survival and cause‐specific mortality of telemetered mule deer (Odocoileus hemionus) in northern California. Over the course of our study, we visited mortality sites of neonates (n = 91) and adults (n = 23) to ascertain the cause of mortality. Rapid site visitations significantly improved the successful identification of the cause of mortality and confidence levels for neonates. We discuss the need for rigorous and standardized protocols that include measures of confidence for mortality site investigations. We invite reviewers and journal editors to encourage authors to provide supportive information associated with the identification of causes of mortality, including uncertainty.
... The MLST approach involves selecting a small number (often 5-7) of the highly conserved housekeeping genes, spread across the genome, to evaluate potential genetic differences (Maiden et al. 1998). Cassirer et al. (2017) developed a MLST method for differentiating strains of M. ovipneumoniae that consists of four loci: a region of the small ribosomal subunit (16S rRNA), a region in the 16S-23S intergenic space, and regions encoding two housekeeping genes (RNA polymerase B and DNA gyrase B). Kamath et al. (2019), applying this method to samples from domestic and wild Caprinae from the western US, concluded that this very diverse bacterium includes hundreds of strain types. ...
... The WADDL extracted DNA from the samples using the MagMAX-96 Viral RNA Isolation Kit (ThermoFisher Scientific, Wal-tham, Massachusetts, USA) and real-time PCR was performed for M. ovipneumoniae as previously described ). The extracted DNA included at least one M. ovipneumoniae-positive individual from each population for each year detected and were genotyped using MLST, as previously described (Cassirer et al. 2017). Sequencing of the amplified PCR product was carried out by a commercial company (Genewiz, South Plainfield, New Jersey, USA). ...
In 2018, Mycoplasma ovipneumoniae was detected in free-ranging caribou (Rangifer tarandus grantii) and Dall's sheep (Ovis dalli dalli) in Alaska, US. Evaluation of additional nasal swabs and archived tissues for M. ovipneumoniae suggested that this bacterium was widespread geographically and temporally in populations of both species. Multilocus sequence typing (MLST) of four loci identified a single, novel, apparently stable strain type of M. ovipneumoniae in 11 Dall's sheep and 15 caribou in multiple populations across Alaska sampled over a period of 15 yr (2004-2019). This strain type differs from those detected to date from wild or domestic sheep (Ovis aries) or goats (Capra aegagrus hircus) tested in Alaska or the lower 48 US states. Although the population health implications of this strain are unknown, it has not been associated with population-wide mortality events. The presence of this strain does not decrease the potential risk from the introduction of a pathogenic M. ovipneumoniae strain associated with severe disease in other wildlife populations; therefore, continued monitoring for signs of disease and additional strains is important.
