[From the U.S. Government Printing Office, www.gpo.gov]













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         I   Qll
              430.7
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              B76
              1996



                                                                             Task 19   VCU/DEQ/VMRC
                                                                                                                FINAL PRODUCT
                                                                             FY 1995   FIELD-TESTING OF DISEASE RESISTANT EASTERN OYSTER
                                                                                       STRAINS IN THE CHESAPEAKE BAY
                                          Virginia Coastal Resources Mana-ge-ment-
                           Interim Progress Report for the perio@ October 1995 - September 1996
                                                   Grant 4 NA570ZO561-01
                            Field-testing of disease resistant eastern oysters in Chesapeake Bay
                                    Bonnie L. Brown, Arthur J. Butt and James A. Wesson
                                L

           NARRATIVE:
                 Restoring water quality within Chesapeake Bay will be greatly aided by biological filtration as
           performed by filter feeding organisms in addition to chemical and physical treatments. Oysters are a
           prime candidate to fill the role of reducing the effects of excess phytoplankton and associated
           eutrophication due to their high capacity for fitration. This "top-down" as opposed to the traditional
           "bottom-up" strategy can be highly cost effective. For example, conservative estimates of
           expenditures by participants in the Chesapeake Bay Program during the period 1984 - 1993 totalled
           $2.3 billion for nutrient reductions that included point and non-point source and combined sewer
           overflow. While "bottom-up" measures have significantly reduced phosphorus, they have done little
           to reduce nitrogen which is a primary cause of eutrophication in the Bay. Near the turn of the
           century, prior to being overharvested and stressed by pollution and disease, oysters are estimated to
           have I-lad the capacity to filter the entire volume of Chesapeake Bay in approximately I week,
           removing 20-40% of the daily carbon. No technological achievement can rival this level of
           cleansing! Recent estimates by VMRC indicate that it requires between $6,000 - 65,000 per acre to
           construct viable reefs depending on the types of materials used. Assuming that 5% of the Bay's 11.5
           billion m2 surface area is amenable to oyster survival, then reefs could conceivably cover 142,080
           acres of the Bay. Further assuming that the cost to construct reefs is midway within the estimated
           range, say $20,000 per acre, then the same 10 year expenditure devoted to oysters and other living
           resources would have had a much greater effect by promoting the removal of particulate carbons fro
           the system, contributing to improved clarity of the water column, and enhancing SAV.
                  Unfortunately, the oysters currently residing in the Bay are a mere vestige of the once
           abundant population. Those that remain are intolerant of the exotic diseases Dermo and MS that
  F        now threaten the remaining native Chesapeake oysters. Some individuals who are unaware of the
             gative consequenc                                                                    aparies4ster
  _9                            es of introducing exotic species have suggested introducing the J
           ne
           Crassostrea gigas to fill the ecological void. The known negatives associated with such a proposal
           include gametic competition with the indigenous C. virginica, the fact that C. g'gas does not prosper
  F) I     under the low salinity conditions that prevail in the majority of the Bay, and infestation of C. gigas
           with pests, parasites, and viral diseases that are not native to Chesapeake Bay. There is hope,
    ;@z    however, for eventually restoring oysters to Chesapeake Bay and reaping the free filtration services a
  F        flourishing oyster population would provide. Other geographic strains of th  e indigenous eastern
           oyï¿½ter, C. virginica, have been demonstrated to tolerate the challenge of disease while continuing to
           grow and reproduce at some Bay sites. For example, we know that some oysters that demonstrate
                                                                                                         ,on
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                                                                                                           that
















           outstanding performance at low salinity may perish at high salinity due to the effects of disease.






        Likewise, an oyster strain that performs well at high salinity may perish when subjected to synergistic
        stresses of low salinity and low temperature. In fact, it is likely that introduction of several strains of
        oyster will be required for the most effective restoration and culture of oysters in the wide variety of
        lower Chesapeake regions. This study involves testing of disease tolerant oyster stocks best suited to
        the lower Bay and will identify several optimum locations of suitable habitat for aquaculture and
        restoration purposes. Growth, disease tolerance, and survival are being assessed monthly for each of
        six oyster strains cultivated under three salinity regimes: low, moderate, and high.
                The specific objectives of this project are fourfold:

                T  Evaluate several potentially suitable oyster strains for aquaculture and replinishment
                   programs in lower Chesapeake Bay;
                (D Evaluate growth rates, disease tolerance, and survivability of various oyster stocks over an
                   18 month period;
                3  Identify oyster strains most suited for culture in the wide range of salinities that occur in
                   the lower coastal zone; and
                   Demonstrate the beneficial economic application of aquaculture to restoration of this local

                   resource.


        Experimental Design:
                Broodstock oysters were collected from geographically distinct eastern oyster populations
        (Table 1). They were conditioned and spawned at Horn Point Environmental Laboratory in Maryland
        by cooperative agreement and disease-free spat were shipped directly to the Ecological Genetics Lab
        at VCU. Spat from each group were deployed in the Fall of 1995 at three coastal zone sites in lower
        Chesapeake Bay. The sites were selected by accessibility and salinity (low: 10 O/oo, moderate: 20 '/oo,
        and high: 32 O/oo). Growth rates, disease tolerance and survivability of six strains have been
        monitored monthly and will continue for a total of approximately 18 months.
                The six oyster groups are being cultivated side-by-side in floating rafts as previously described
        by Brown et al. 1993, 1994, and 1995. Spat were held initially in small-mesh bags within floating
        trays. As they have grown, they have been transferred to successively larger mesh sizes in the
        floating trays. Growth is determined monthly by measuring both shell length (mm) and wet/dry tissue
        weight (g). All measurements except mortality are determined by randomly selecting approximately
        25 oysters from each strain at each site. Mortality is recorded monthly for all members of each
        treatment. Individual representatives. of each group at each site are sacrificed monthly for condition
        .assessment and disease analysis according to Paynter and DiMichele (1990). All of these operations
        are conducted through the Ecological Genetics Lab at VCU.







                        Table 1. Oyster strains currently being tested in the coastal zone/lower
                                  Chesapeake Bay for growth, disease, and survival.

                                   Strain   Origin of Broodstock


                                     1 .    Upper Chesapeake Bay
                                     2.     North Carolina
                                     3.     South Carolina
                                     4.     Texas
                                     5.     Louisiana
                                     6.     Louisiana (Triploid)

          Progress:
                The project was begun by introducing five strains of eastern oyster, Crassostrea virginica, during
          September, October and November 1995 at four lower Chesapeake Bay sites'. The geographic range of
          native C virginica strains tested includes synchronously spawned oysters from Texas, Louisiana, South
          Carolina, North Carolina, and Chesapeake Bay. Each of the four geographically estranged strains was
          hy bridized to Chesapeake Bay oysters to produce the four hybrid groups and one pure Chesapeake Bay
          group introduced in September which we are now tracking. A sixth strain, triploid C. virginica denoted
          3N, was introduced in July 1996. To avoid differential treatment of any one set of oysters, the five strains
          were identified by the numbers 16, 17, 18, 19, and 20. At present, with the exception of the triploids, the
          study is blind and we cannot yet determine which of the five strains originated from which geographic
          location.
                 Non-disease mortality was noted at various times during the study due to pests and predators,
          noteably crabs, tunicates, and an as yet unclassified parasitic worm. Crab induced mortality was limited to
          very small oysters (< 10 mm) at a single site. Tunicates compete with the oysters for space during specific
          times of the year at one site. Exceedingly high barnacle sets occasionally occur on the trays requiring tray
          maintenance. The worm infestation, which also occurred at only one site, was terminated by chemical
          treatment of the oyster shells. This infestation was one of several times that unscheduled (non-
          monitoring) trips were necessary. Other unscheduled trips have been necessary to retrieve trays that have
          broken loose from their tethers and to salvage repair damage following Hurricane Fran and other severe

          storm events.
                 Evaluation of growth rate, survivability, and disease tolerance began in October 1995. Oyster
          groups ranged from 7 - 16 mm average length at the time they were deployed. Growth has progressed at
          each site at anticipated rates resulting in a current range of sizes from 30 mm at the low salinity site to 59
          min at the high salinity site. Growth, condition, disease and survival differences among the groups are
          minor at this point (Figures 1, 2, 3, and 4). Cumulative non-diseaï¿½e mortality (due to pests and predators;
          in particular, smothering by tunicates) is currently 30% or less regardless of site. Distinct site-dependent
          mortality differences became apparent during September 1996, being extremely limited at low salinity


           Gloucester County (low salinity), Matthews County (low to moderate salinity) and two sites in Virginia Beach (moderate
            salinitv and high salinity)






          sites (< 20%) and increasing rapidly at the moderate and high salinity sites to approximately 60% (Figure
          4). Our disease analy ses indicate that Dermo is a substantial cause of mortality in the moderate and high
          salinity sites (Figure 3), as predicted.


          Prod ucts/Deliverables:
                We projected that by the end of year one of the project, several potentially suitable oyster strains
          would be identified and deployed for use in aquaculture and replenishment in lower Chesapeake Bay
          regions of various salinities. This has been accomplished for six strains and four sites. Substantial
          progress has been made toward completing the products and deliverables projected for year two of the
          proj ect.


          COMMENTS:
                We had originally created the budget in anticipation that the VIMS disease analysis facility could
          be contracted to analyze our 375 oysters per month as follows:
                   F. Contractual. Disease analyses (Dermo and MSX) will be contracted to the
                   lowest qualified bidder in accordance with the Virginia Public Procurement Act.
                   The request for bids will be handled by the VCU Procurement Office and will be
                   preapproved by the VCRMP staff prior to execution.
          With the departure of Dr. Perkins, VIMS experienced a tremendous increase in the numbers of oysters to
          be analyzed. By the time we received funding approval, VIMS could not accommodate our request.
          Therefore, VINIS generously trained our own staff to perform the necessary disease analyses. Dermo
          (Perkinsus) analyses are very time-consuming, requiring approximately 2 persons to spend 3.5 - 4 days per
          month to prepare, incubate, and microscopically examine 375 individual oyster samples. For MSX,
          duplicates of each sample are being archived for later analysis if it is suspected that Haplosporidium
          might be the cause of any otherwise unexplained mortalities. The necessity of doing disease analyses
          caused us to spend considerably more than was originally budgeted in the supply and personnel categories
          during the first and second quarters of the project. The VCU lab is now set up and running smoothly and
          much less is expected to be spent on supplies during the remainder of the project.
                 The cost for VCU to perform the disease analyses in our own lab is currently $782 per month
          ($0.50 per sample in supplies and 64 hours total time spent to perform the analyses) which totals $9390
          per year in unanticipated cost to successfully complete the study. As a result of the increased work load,
          NOAA funds were totally expended as of I July 1996 leaving a shortfall of approximately 3 months. To
          partially address the shortfall, the budget was amended by moving the funds originally budgeted for
          contracting disease analysis ($2,200) to the personnel category and funds from travel ($1,200) to supplies.
          The difference has been paid by contributions from Dr. Brown ($ 11,489) and Chesapeake Scientific
          ($1200) totalling $12,689 to date. The ammended budget outlines a 1:1 state match for this project. At
          the completion of year one, state matching has been exceeded by 53%. Without assistance to correct this
          disparity, it is expected that total state matching ftinds for the project will exceed $70,261 by the end of.
          the second year. This translates to a 1: 1.6 match.



        mm m mmIm m mm m m m m mm mm mm
                Fig I                       Size of oysters at four sites in lower Chesapeake Bay
                                                                                                  @16 -17                                 18 -19 @20 -3N.]
             Size (mm)                                                                                                                                   70
                  70
                                                                                                                                                                                                     Register
                                                              Wesson
                  60                                                                                                                                     60


                                                                                                                                                         50      - ----- -
                  50


                  40                                                                                                                                     40


                  30                                                                                                                                     30      - -------


                                                                                                                                                         20
                  20


                                                                                                                                                         10
                  10

                    0                                                                                                                                                   10       11        12         1        2         3        4         5        6         7         8         9
                       9        10       11        12         1        2        3         4         5        6         7        8         9                     9
                                                      Month (1994 - 1995)


                  70                                                                                                                                     70
                  60                                             Savage                                                                                  60                                                     Hill

                  50                                                                                                                                     50


                  40                                                                                                                                     40


                                                                                                                                                         30
                  30


                  20                                                                                                                                     20


                                                                                                                                                         10
                  10


                                                                                                                                                            0
                     0 9        10       11        12         1        2        3         4         5        6         7         8        9                     9       10       11        12         1        2         3        4         5         6         7        8         9




                  Fig 2, Cumulative disease-induced mortality of oysters
                                               at four lower Chesapeake Bay sites
          Cum. % Mortality                                       @1 6 @1 7 -18 @ 19                              20         3N:]
                                            Wesson                                                                                Register
               0.8                                                                                   0.8


               0.6                                                                                   0.6


               0.4                                                                                   0.4


               0.2                                                                                   0.2
                                                                            7@'
                0                                                                                       0
                  9     10    11    12     1     2     3      4     5     6     7     a      9           9     10    11    12     1     2      3     4     5     6     7      8     9
                                           Month (1994 - 1995)
               0.8                            Savage                                                  0.8                                H-i I I

               0.6                                                                                    0.6

               0.4                                                                                    0.4

               0.2                                                                                    0.2
                                                                                        AP





                                                                                                        0
                 0
                   9     10    11    12     1     2     3     4     5      6     7     8     9            9     10    11    12     1     2      3     4     5     6     7      8     9



                            Mon m                                          m m                     m m
                  Fig 3. Condition index at four lower Chesapeake Bay
                                                                         sites
                                                      @16 @17              18 -19 @20 -3N
       Condition Index
          0.7                     Wesson                                       0.7                   Register
          0.6                                                                  0.6                                     - --- ----


           .0.5                              - ------                          0.5


          0.4 -                                                                0.4


          0.3                                                                  0.3  - -


          0.2                                                                  0.2


          0.1
                                                                               0.1


            0 -                                                                  0
             9    10   11   12    1   2    3    4    5    6    7    8    9         9   10   11   12   1    2    3    4    5    6    7    8    9
                                  Month (1994 - 1995)
          0.7                       Savage                                     0.7                         Hill
          0.6                                                                  0.6  -

          0.5                                                                  0.5

          0.4                                                                  0.4

          0.3                                                                  0.3

                                                                               0.2
          0.2


                                                                               0.1 -
          0.1

             0                                                                   0
              9   10   11   12    1   2    3    4    5    6    7    8    9         9   10   11   12   1    2    3    4    5    6    7    8    9




           Fig 4. Incidence of disease (Dermo) at four
                  lower Chesapeake Bay sites

                      @16 @17  18 @ 19" 20
   Weighted Prevalence of Dermo
    5                            5
              Wesson                       Register






     10 11 1212  3 4  5 6 7  8 9  10 11 121 2 3 4  5 6  7 8 9
             Month (1994 - 1995)
    5                             5
               Savage                         Hill
    4'                            4



CD
CO






0)
                                                           4-0,


     10 11 121 2 3 4  5 6 7  8 9  10 11 121 2 3  4 5 6  7 8 9