Download Annual_Report_for_2007-08

STUDY PERFORMANCE REPORT
State: Michigan
Project No.: F-80-R-9
Study No.: 230723
Title: Genetic diversity of fish populations in Michigan:
Effects of environmental and anthropogenic factors on
recruitment and apportionment of phenotypic and
genetic diversity of fish populations in inland and Great
Lakes habitats
Period Covered: October 1, 2007 to September 30, 2008
Study Objectives: To use molecular markers and population genetics theory to investigate
demographic, ecological, and genetic stock relationships within and among native and naturalized
fish species of importance in Michigan. I seek to estimate recruitment, examine variation in life
history traits, estimate degree of stock structure, and the effects environmental and anthropogenic
factors on population and meta-population levels of genetic diversity.
Summary: Research is ongoing in each of the main focus areas, related to historical and
contemporary factors affecting the genetic diversity of fisheries in Michigan waters. Studies
involve numerous native species of economic and ecological importance that are intensively
managed. This project seeks to provide information to better understand how recent management
practices have affected the genetic diversity of Michigan’s fisheries.
Findings: Jobs 1–9 were scheduled for 2007-08, and progress is reported below.
Job 1. Title: Review literature.–Fish populations in northern latitudes, including the Great Lakes
region have only recently recolonized currently occupied laucustrial and riverine habitats.
Accordingly, levels of genetic differentiation are among populations are generally low. Estimates
of variance in the frequency of alleles at genetic markers such as microsatellites or mitochondrial
(mt)DNA are routinely used in ecological and conservation genetic studies and must be
interpreted in the context of the recency of population separation times and impacts water levels
and degree of connectivity of pro-glacial habitats has had on fish movements and gene flow.
Extensive literature reviews were conducted on the genetic basis for resident and anadromous life
history phenotypes, including brook charr to provide input to Division comments to federal
listings for coaster brook trout. Data from across the species range, including drainages in Lake
Superior are equivocal regarding the degree of distinctiveness of coaster brook trout and of the
genetic bases for phenotypic and behavioral differences observed.
I have conducted extensive literature reviews on genetic and environmental influences of disease
susceptibility in salmonid species, focusing primarily on mechanisms of transmission. The overall
objective of the literature review in this area was to better understand the epidemiology of
Renibacterium salmoninarum (RS) and bacterial kidney disease (BKD) in the Great Lakes basin,
to understand potential genetic factors underlying susceptibility to disease and the importance of
natural stock structure and hatchery management and supplementation in mediating disease
prevalence, transmission, and persistence in BKD-susceptible salmonid species, with particular
emphasis on chinook salmon (Oncorhynchus tshawytscha).
Literature reviews are also supporting ongoing and future work on the effects microbial
communities have on fish recruitment through effects on egg mortality. Scientific articles
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regarding the interaction of the developing sturgeon egg as a model system to investigate
interactions between fish eggs with the microbial community were reviewed. This stage is a
critical point of reference for studies of developmental evolution, for development of predictive
models of consequences of changes in land-use, damming of rivers, and global warming, and for
fish recruitment and conservation. Our focus is on reproductive traits of the sturgeon that interact
both positively and negatively with the microbial populations of streams. Traits can co-evolve
based on the degree of covariance between the selected traits and environmental variables tied to
locations and timing of reproduction. Lake sturgeon represent a unique example of adaptation to
temporal variation in environmental conditions within a breeding season. Stream environmental
conditions including temperature and microbial communities vary temporally within a season and
spatially over different scales, and are expected to affect embryonic development, juvenile traits,
and survival. Our literature review will direct studies seeking to understand how microbes and
temperature affect multivariate selection during embryogenesis and early larval life stages in lake
sturgeon and other economically and recreationally important fish species in Michigan. Ongoing
work will lead to greater understand of how temporal and spatial variation in these agents of
selection affect the evolution of complex phenotypes.
Job 2. Title: Consultation.–Considerable effort was made to work with personnel from Hatchery,
Management, and Research Sections within Fisheries Division to obtain information, coordinate
research activities, and to communicate results.
Research activities involving sturgeon, including capture and data collection on spawning adults
has been coordinated with personnel from the Gaylord Management office and personnel in the
Lake Huron Basin group. I also worked with the Alpena Research Station to collect and analyze
Chinook salmon scale samples at several candidate genes we have shown to be associated with
different expression in bacterial kidney disease challenges. Consultation was also provided to the
Marquette Research Station and members of the Lake Superior Basin Team on coaster brook
trout including analyses of coaster and resident fish.
Numerous presentations were given at meetings including presentations on sturgeon culture at the
Wolf Lake State Hatchery and at the Hatchery Section Meeting. I consult monthly with the head
of Hatchery Section and I have worked with members of the Lake Huron Basin Team to
communicate future plans, current project status, and future plans associated with a long-standing
project on lake sturgeon. I have worked closely with Management and Hatchery Section
biologists and technicians associated with weir operations in the Lake Michigan Basin. I made a
presentation on use of fisheries genetics in management to graduate students associated with the
Institute of Fisheries Research (IFR) in Ann Arbor. I have also made a presentation in the area of
landscape genetics and applications to river inventory and monitoring to researchers, staff, and
students at IFR.
I have communicated results of our Great Lakes landscape genetics work at regional meetings.
Scientists from Department of Fisheries and Oceans and the Ontario Ministry of Natural
Resources are very interested in using data collected for the deepwater sculpin as they consider
the status of its COSEWIC listing.
Job 3. Title: Conduct studies.–We have conducted studies in streams and in the Great Lakes
focusing on important biotic and physical features that affect stock structure and recruitment. The
Great Lakes portion of the work represents a major collaboration between researchers from
Michigan State University and the U.S. Geological Survey (USGS) Great Lakes Science Center.
Data were collected, analyzed and interpretation was offered by biologists associated with USGS
research stations in the upper Great Lakes. Members of an interagency consortium of researchers
involved with collection, interpretation, and application of Great Lakes remotely censused data
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have been involved in information and technology transfer and in the sharing of ideas. The
collection and analysis of spatial and environmental data was facilitated through collaboration
with the USGS Great Lakes Science Center, Eastern Michigan University, and the Great Lakes
GIS project at the University of Michigan.
We sampled and genotyped 4,009 individuals across five codistributed native Great Lakes forage
fish species (lake herring, bloater, nine-spine stickleback, slimy sculpin, and deepwater sculpin)
from 27 locations in Lakes Huron, Michigan, and Superior. Genetic characterizations were made
to determine presence and location of major genetic stocks within and among each lake, and to
determine whether levels of spatial genetic differentiation among stocks could be attributed to
historical biogeographic events and to hydrogeomorphological features of lake habitats (e.g.,
depth, temperature, productivity, currents) that influence movements and gene flow.
Analyses of mitochondrial DNA (mtDNA) sequences and microsatellite loci revealed a
fundamental genetic discordance for all five species between samples collected from Lake
Superior and samples collected from Lakes Michigan and Huron. Results suggest that historical
vicariance and reproductive isolation since the basins were initially colonized were largely
responsible for the major genetic differences. The existence of multiple phylogenetic lineages
within each species suggests that the Great Lakes were colonized by individuals from multiple
glacial refugia, as has been suggested for other fish and invertebrate species.
Bloater and lake herring are managed as two distinct species in the Great Lakes and differ
markedly in a number of ecological characteristics including depth preference and timing and
location of spawning. Interestingly, both species share the same mtDNA haplotypes, including
haplotypes from both putative glacial refugia. Data suggest that both species have a shared recent
evolutionary history and that ecological specialization may be a recent phenomena. Great interspecific differences in Lakes Michigan and Huron relative to Superior likely reflect smaller
effective population sizes in the lower lakes, especially for lake herring. However, differences in
allele and haplotype frequency between species greatly exceed the variance among locations or
lake basins within a species. In most locations where both species were sampled, differences in
genetic characteristics between species were sufficient to allow individuals to be assigned to
species solely on the basis of genotype. With few exceptions, biologists appear to be able to
distinguish species based on pre-defined morphological criteria.
Analyses are ongoing of spawning habitat use by lake sturgeon in the Black River and of habitatspecific differences in recruitment. Preliminary data from individuals captured in multiple years
revealed that with high probability lake sturgeon spawn at the same date within a season and
location in multiple years. Timing and duration of larval dispersal is related to the time and
locations of spawning by females and contributes significantly to inter-individual variance in
reproductive success and thus to recruitment.
Job 4. Title: Examine effects of landscape features.–For the bloater and lake herring work
described in Job 3, the degree of genetic stock structuring within each basin varied greatly
between species, likely reflecting species differences in effective population size, species’
ecology, and propensity for dispersal over spatially heterogeneous basin landscapes. Significant
spatial variation was found between sampling locations within each basin for lake herring,
bloater, slimy sculpin and stickleback but not deepwater sculpin.
The locations of areas of spatial genetic differences were not concordant among species,
suggesting that basin landscape features differentially impact movements and gene flow of the
different species. Lake herring occupying major embayments in Lake Superior are significantly
differentiated. Slimy sculpin from northern and southern areas of Lake Superior are also
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genetically differentiated. Distance, current patterns, and fall temperature differences between
sampling locales were significant predictors of inter-location genetic differences in lake herring.
Location differences in spring and fall chlorophyll concentrations were important predictors of
inter-location genetic differences for bloaters. Inter-location distance and current patterns also
explained a significant proportion of inter-location variation in slimy sculpin and stickleback.
Job 5. Title: Design hatchery monitoring protocols.–We have implemented collection regimes for
all species, strains, and year classes in the Michigan DNR hatchery system. Samples from adults
and juveniles will be used to assess the effectiveness of current gamete-take methods for retention
of levels of genetic diversity.
We have evaluated the effectiveness of culling adult Chinook salmon at weirs as a means of
reducing probabilities of spawning individuals with bacterial pathogens. Data reveal that of 174
males checked by MDNR personnel at weir and subsequently examined for bacteriology and
genetically based on PCR for BKD and Aeromonas salmonicida (AS), 50 or 174 males examined
at the weir were culled (discarded and gametes not used; 28.7%). Five of 21 cases of BKD (based
on nPCR) from these 174 males were among the fish culled (23.4%). Thirty of 50 fish culled
were found to be infected by multiple bacteria species or strain (60%). Five of 50 fish culled were
found to be infected by a single bacterial species or strain not including AS (10%). Three of the
fish culled were founded to contain AS or other motile Aeromonids of a total of 38 fish infected
with As or motile Aeromonids (7.95%). Of 96 females checked by MDNR personnel at weir and
subsequently examined for bacteriology and genetically, 31 of 96 females examined at the weir
were culled (discarded and gametes not used (32.3%). Nine of the fish and 9 of 27 fish found
based on nested PCR to have BKD were culled (33% of known infected BKD+fish). Five of the
29 fish that were found to contain either AS or other motile Aerominids of 29 fish infected with
As or Aeromonids (17.2%) were culled. Thirteen culled fish of 32 fish (40.6%) found to be
infected by multiple bacteria species or strains. Seven of 19 fish (37%) were found to be infected
by as single bacterial species or strain not including AS were culled.
Findings suggest that using a single diagnostic assay to detect, and consequently cull, spawning
Chinook salmon is not sufficient since each of assay (i.e., bacteriology vs nPCR) has a different
target to measure. However, performing all assays is important to evaluate the progress of BKD
infection in a particular stock. Culling based on visual inspection of gross physical condition can
decrease probability taking gametes from fish with bacterial infection though the specificity of
the technique is relatively low.
Job 6. Title: Examine effects of stocking.–Scale samples were collected at each of the broodstock
weirs to evaluate the potential genetic implications of stocking practices. These samples were
archived for evaluation at a later date.
Job 7. Title: Estimate effects of environmental pathogens.–Emerging and resurging infectious
agents pose significant challenges to the health and persistence of ecological communities they
inhabit. Dynamics of disease emergence are poorly understood within the context of evolutionary
forces, ecological interactions, epidemiological processes, or in terms of genetic (innate) factors
affecting immunological response. It is important to understand the responses of pathogen
populations to selective processes, both imposed by the host’s immune system and in the natural
environment. We embraced this synergistic approach. During the past fiscal year, we have
considered several complementary areas including molecular, quantitative, and population
genetics, epidemiology, pathology, laboratory manipulation, and ecosystem-based surveys to
infer mechanisms of susceptibility and methods of disease transport and persistence. Work has
principally focused on Chinook salmon (Oncorhynchus tshawytscha) and Renibacterium
salmoninarum (RS) the causative agent of bacterial kidney disease.
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Using cDNA microarrays we identified 312 genes that were statistically differentially expressed
between infected and non-infected juvenile Chinook salmon. The ratio of genes detected as
significantly up-regulated vs. down-regulated in kidneys from infected fish was approximately
3:1 (237:75). The genes selected for real-time qPCR validation reflect this trend. Lysozyme and a
number of other genes related to the phagocytic destruction of pathogens by macrophages during
the immune response were detected as up-regulated by microarray analysis. Genes involved in the
cell-mediated adaptive immune response such as MHC were significantly differentially expressed
as were immunoglobulin genes involved in humoral response. A large number of genes of
unknown function were significantly differentially expressed and should be investigated further.
A large number of genes identified from the literature and our array were found to contain
polymorphisms in Chinook and other species. We developed rapids screening protocols as a
means to survey polymorphisms in genes of potential adaptive function in natural and hatchery
populations. Genes were sequenced and found to be homologous in other fish species and can be
used to provide a physical map of networks of genes with known immunological function.
Incidence of RS varied greatly in Chinook collected among Lake Michigan sampling locations,
collection periods and between males and females. Disease incidence generally increased over
time during the spawning season and differed geographically suggesting that fish from different
regions of the Lake Michigan basin may be arriving at spawning areas at different times and from
areas with different prevalence of disease. Incidence of other pathogens was also quantified.
Statistical associations were found between RS and motile Aeromonids, suggesting that
probabilities of secondary coinfection by other bacterial species is more likely give presence of
RS. Incidence of fish differing in stage or intensity of infection was also detected across locations
and sampling period.
Fish in advanced stages of disease were more likely to have gametes also infected with RS. In
addition, fish with no evidence of RS infection in the kidney were found to have gametes infected
with RS, suggesting that vertical transfer from soma to gametes occurs during gameteogenesis,
and that somatic tissues can be cleared of infection while RS remains in the gonads.
Experimental density, handling and nutritional treatments were designed to evaluate the relative
impacts and probabilities of acquiring RS from the water and changes in incidence over time.
Experimental conditions simulating different hatchery practices including different densities,
nutritional condition, and handling were found to significantly affect probabilities of acquiring
RS from the environment. Data on RS incidence in native and introduced non-salmonid species
suggests that species that comprise the forage base of predator species of recreational and
commercial importance are significant reservoirs of infection and mechanisms of disease
transmission in the open lake and adjoining environments.
Job 8. Title: Prepare annual progress report and communicate results.–Findings of activities are
communicated in this document and to Michigan DNR and regional fisheries biologists via oral
communications at meetings, professional meetings, and written reports.
Job 9. Title: Publish report.–Because of recent changes in the granting and proposing of this work
from a research to a monitoring type of study, there was some confusion about the completion of
this job. This job was not completed this year and will be addressed with an appropriate
amendment for this project in the coming year.
Prepared by: Kim Scribner
Date: September 30, 2008
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