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Environmental significance of freshets in reducing Perkinsus marinus infection in eastern oysters Crassostrea virginica: Potential management applications

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The effects of extreme freshwater events on Perkinsus marinus–Crassostrea virginica interactions remain unexplored. The effects of freshwater events on P. marinus infection in C. virginica and oyster survival were therefore examined in controlled laboratory experiments and a field study. For the laboratory experiments, oysters were collected in spring, summer and winter from an area in Louisiana where P. marinus is endemic. Oysters were placed in 2 recirculating water systems at a salinity and temperature similar to their collection site. They were subjected to 2 salinity treatments (freshet and control). Freshet events were simulated by reducing the water to salinities of 0 to 1 ppt over a 48 h period, and maintained for a 21 d period. Control oysters were maintained at the initial salinity. Thirty oysters were sampled prior to the freshet event, and 30 oysters per treatment group (freshet, control) were sampled on Days 7, 14 and 21 after initiation of the freshet event. Oyster mortality, P. marinus infection intensities, oyster condition index and oyster plasma osmolality were measured in weekly samples. All 3 simulated freshet events (i.e. spring, summer, winter) resulted in a significant reduction in P. marinus infection intensity, but failed to eliminate infection. The failure of the oyster plasma to reach very low osmolality (<50 mOsm kg–1) provides a likely explanation for the lack of complete P. marinus elimination. The field study involved sampling oysters monthly in the Caloosahatchee estuary, Florida, from September 2000 to February 2002, and determining P. marinus weighted prevalence and condition index of wild oysters, and growth and survival of caged juvenile oysters. The data strongly support the contention that the numerous freshwater releases to the Caloosahatchee River kept P. marinus infection intensities in oysters at low levels, resulting in an overall low weighted prevalence, low oyster mortality and good growth. Data from our field study appear to support the hypothesis that repetitive and well-timed freshet events can prevent infection of oysters with P. marinus or at least maintain P. marinus infections at non-lethal intensities (e.g. <106 parasites g–1 wet tissue) in oyster populations. The use of an adaptive management approach involving control of freshwater inflows could be invaluable to the oyster industry in areas close to freshwater diversion projects.
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MARINE ECOLOGY PROGRESS SERIES
Mar Ecol Prog Ser
Vol. 248: 165–176, 2003 Published February 20
INTRODUCTION
Modern ecological synthesis recognizes that varia-
bility is an inherent part of natural systems: natural and
human induced change and disturbance are consid-
ered the norm rather than the exception (Odum 1969,
Pickett et al. 1992). Nonetheless, in attempts to estab-
lish causal relationships among various environmental
factors, many natural-system studies assume that sys-
tems are in a state of equilibrium that categorically ex-
© Inter-Research 2003 · www.int-res.com
*Email: mlapey@lsu.edu
Environmental significance of freshets in reducing
Perkinsus marinus infection in eastern oysters Crassostrea
virginica: potential management applications
Megan K. La Peyre1,*, Amy D. Nickens2, Aswani K. Volety3, Gregory S. Tolley3,
Jerome F. La Peyre2
1US Geological Survey, Louisiana Fish and Wildlife Cooperative Research Unit, School of Renewable Natural Resources,
Louisiana State University, Baton Rouge, Louisiana 70803, USA
2Cooperative Aquatic Animal Health Research program, Department of Veterinary Science,
Louisiana State University Agricultural Center, Baton Rouge, Louisiana 70803, USA
3Division of Ecological Studies, Florida Gulf Coast University, Fort Myers, Florida 33912, USA
ABSTRACT: The effects of extreme freshwater events on Perkinsus marinus –Crassostrea virginica in-
teractions remain unexplored. The effects of freshwater events on P. marinus infection in C. virginica and
oyster survival were therefore examined in controlled laboratory experiments and a field study. For the
laboratory experiments, oysters were collected in spring, summer and winter from an area in Louisiana
where P. marinus is endemic. Oysters were placed in 2 recirculating water systems at a salinity and tem-
perature similar to their collection site. They were subjected to 2 salinity treatments (freshet and control).
Freshet events were simulated by reducing the water to salinities of 0 to 1 ppt over a 48 h period, and
maintained for a 21 d period. Control oysters were maintained at the initial salinity. Thirty oysters were
sampled prior to the freshet event, and 30 oysters per treatment group (freshet, control) were sampled
on Days 7, 14 and 21 after initiation of the freshet event. Oyster mortality, P. marinus infection intensi-
ties, oyster condition index and oyster plasma osmolality were measured in weekly samples. All 3
simulated freshet events (i.e. spring, summer, winter) resulted in a significant reduction in P. marinus
infection intensity, but failed to eliminate infection. The failure of the oyster plasma to reach very low
osmolality (<50 mOsm kg–1) provides a likely explanation for the lack of complete P. marinus elimina-
tion. The field study involved sampling oysters monthly in the Caloosahatchee estuary, Florida, from
September 2000 to February 2002, and determining P. marinus weighted prevalence and condition
index of wild oysters, and growth and survival of caged juvenile oysters. The data strongly support
the contention that the numerous freshwater releases to the Caloosahatchee River kept P. marinus
infection intensities in oysters at low levels, resulting in an overall low weighted prevalence, low oyster
mortality and good growth. Data from our field study appear to support the hypothesis that repetitive and
well-timed freshet events can prevent infection of oysters with P. marinus or at least maintain P. marinus
infections at non-lethal intensities (e.g. <106parasites g–1 wet tissue) in oyster populations. The use of
an adaptive management approach involving control of freshwater inflows could be invaluable to the
oyster industry in areas close to freshwater diversion projects.
KEY WORDS: Dermo · Perkinsus marinus · Crassostrea virginica · Freshet · Infection intensity ·
Mortality · Osmolality · Condition index
Resale or republication not permitted without written consent of the publisher
Mar Ecol Prog Ser 248: 165–176, 2003
cludes short-term and catastrophic events. While envi-
ronmental conditions have long been held as critical
controls on host-parasite interactions through their im-
pacts on the physiological condition, reproduction and
survival of both hosts and parasites, little attention has
been paid to the effects of short-term events on host-
parasite interactions. Understanding the role these
short-term events may play in influencing host-parasite
interactions is critical to the design, application and
evaluation of disease management strategies for en-
hanced production of wild commercially important spe-
cies, such as the eastern oyster Crassostrea virginica.
The oyster parasite Perkinsus marinus is considered
to be a major cause of mortality in Gulf Coast (Craig
et al. 1989, Soniat 1996) and East Coast subtidal
Crassostrea virginica populations (Burreson & Ragone
Calvo 1996, Ford 1996). In the Chesapeake Bay region,
P. marinus has decimated oyster populations (Burreson
& Ragone Calvo 1996). In the Gulf of Mexico, the
market-size component of oyster populations suffers
an estimated 50% yearly mortality from P. marinus
(Mackin 1962, Hofstetter 1977, Powell et al. 1996).
Temperature and salinity are generally held to be the
dominant environmental factors controlling both the
survival and growth of oysters and P. marinus inde-
pendently, and it is likely that they influence the host-
parasite interaction (Soniat 1985, Soniat & Gauthier
1989, Chu & La Peyre 1993, Chu et al. 1993, Ragone
&Burreson 1993, Burreson & Ragone Calvo 1996).
Early studies concluded that oysters can exist and
grow vigorously in salinities slightly lower than the
minimum tolerated by Perkinsus marinus, although it
was concluded at the time that the differences were so
minimal that for practical purposes they did not exist
(Mackin 1956). Despite this contention, along both the
East and Gulf Coasts, locations characterized by the
regular occurrence of freshet events, are noteworthy
for their lack of P. marinus-infected oysters (Brooks et
al. 1988, Chu & Greene 1989, Soniat & Gauthier 1989).
The term freshet is used broadly to denote a rapid and
short-term freshwater event in normally saline waters.
Our overall goal is to identify the significance of
freshet events in reducing P. marinus infection in their
host, the eastern oyster.
There is a paucity of manipulative field and labora-
tory studies examining effects of short-term acute
events such as freshets on oysters, Perkinsus marinus
and oyster–P. marinus interactions. For example, only
recently have we acquired in vitro evidence to suggest
that acute freshwater events may be important in
oysterP. marinus interactions. The most relevant
recent laboratory study tested the effects of acute ex-
posure (24 h) of in vitro cultured P. marinus parasites to
low salinities and found that mortality was greater than
99% (Burreson et al. 1994). Field surveys and studies
that have discussed short-term events concluded that
variation in oyster physiology (Fisher et al. 1996), oys-
ter production (Livingston et al. 2000) and disease inci-
dence (Soniat 1985) were related to storm events and
ephemeral impacts of human activities.This laboratory
and field evidence combined suggests that the use of
environmental averages (e.g. monthly averages) to fully
understand and predict oyster and P. marinus survival,
and interactions in a natural environment may be lim-
ited: P. marinus –Crassostrea virginica dynamics may be
controlled more by extreme events in the environment
(i.e. freshets) than yearly means (i.e. salinity means).
Extreme variations in salinity frequently occur in
coastal areas due to both natural events (e.g. seasonal
rains, heavy rainstorms and El Niño Southern Oscilla-
tion events) and human actions (e.g. land develop-
ment, water management) (e.g. Livingston et al. 2000).
In particular, in southwest Florida, extreme salinity
variations are common due to management practices
designed to accommodate watershed land uses and
development (Volety et al. 2001a,b). Typically, these
water management practices include a cessation of
weir openings during dry months to conserve water
for human consumption (e.g. agricultural, residential),
and frequent weir openings during the rainy season to
prevent upriver flooding. This pattern of weir openings
results in a general freshening of estuaries in the
summer (rainy months) as large freshwater flows are
allowed into the estuary, contrasted with higher salini-
ties in the winter (dry season), when weirs are not
opened and very little fresh water enters the estuary.
The Caloosahatchee River in south Florida, USA, is
dominated by a water management approach involv-
ing the extensive use of weirs. The Caloosahatchee
River basin is located in southwest Florida and drains
3700 km2. The entire watershed is dominated by agri-
cultural and rangeland land uses in the upper reaches
of the basin and urban developments in the lower
reaches of the basin. Water releases into the Caloosa-
hatchee Estuary from Lake Okeechobee are controlled
by the opening and closing of weirs and follow the
typical seasonal pattern described above. This water
management practice results in extreme variations
of salinity in the estuary ranging from 0 to 37 ppt
(depending on time of year and location), providing an
ideal setting to investigate effects of water manage-
ment and freshet events on oyster survival and Per-
kinsus marinus infection.
The main objective of this study was to investigate
the significance and potential role of freshet events in
reducing Perkinsus marinus infection in their host, the
eastern oyster. This study focused on subtidal oysters
as these are the dominant oysters along the Gulf Coast.
This study reports the results of (1) controlled labora-
tory experiments in which we determined the effects of
166
La Peyre et al.: Effect of freshets on eastern oysters infected with P. marinus
simulated freshet events occurring in the spring, sum-
mer and winter on oyster P. marinus infection intensity,
condition index, mortality and plasma osmolality, and
(2) a field study quantifying the related effects of sea-
son and freshet events on oyster P. marinus weighted
prevalence, condition index, mortality and growth.
MATERIALS AND METHODS
Laboratory experiments. Oysters and site: Infected
subtidal eastern oysters Crassostrea virginica, 6 to
10 cm in length, were obtained from the Louisiana Sea
Grant oyster hatchery in Grand Isle, Louisiana. The
Grand Isle area is endemic for Perkinsus marinus,
ensuring that oysters collected from this area will have
been exposed naturally to P. marinus during their
grow-out phase. All oysters in this region are subtidal.
At the time of collection, water temperature and salin-
ity were recorded. Oysters were transported to Louis-
iana State University (LSU), Baton Rouge, in April, July
and November 2001 and evenly distributed between
2recirculating water systems.
Experimental design and exposure system: The
experiment was conducted as a controlled laboratory
experiment with 3 seasons (spring, summer, winter)
and 2 salinity treatments (control and freshet). Each
season 280 oysters were randomly placed, in groups of
13 to 14 oysters, into a total of 21 separate containers.
These 21 containers were divided between the 2 re-
circulating systems holding 1000 l artificial seawater
(Hawaiian Marine Imports), resulting in 10 and 11
containers per system. Initial salinity and temperature
conditions in each system were established that were
similar to conditions at Grand Isle at the time of oyster
collection (April: 18°C, 23 ppt; July: 28°C, 20 ppt;
November, 16°C, 26 ppt). Water in each system was
filtered through 10 and 1 µm filters to eliminate cross-
contamination of parasites between containers. Water
in each system was recirculated at least 4 times h–1, ex-
cept during feeding, and was constantly aerated. The
oysters were fed daily with the marine algae Isochrysis
galbani (Reed Mariculture) for about 4 h, during which
time water bypassed each container. After 1 wk of accli-
mation, 30 oysters from 3 containers (2 containers from
the system with 11 containers, and 1 container from the
system with 10 containers) were sampled to determine
initial (Day 0) Perkinsus marinus infection intensities,
oyster condition index and plasma osmolality. At Day 0,
one system, designated the treatment system, was sub-
jected to a simulated freshet event: salinity was re-
duced to 0–1 ppt over a period of 48 h. Water used for
the reduction of salinity was held in a separate tank and
filtered through a carbon filter to dechlorinate the
water. Throughout the course of the experiment, the
other system was maintained at the initial salinity and
temperature as a control. Oyster mortality and water
quality (pH, NO2, NH3) were measured daily in both
control and treatment systems in order to ensure that
water quality parameters were similar among systems,
and not a cause of mortality in oysters. P. marinus infec-
tion intensities, oyster condition index and plasma
osmolality were determined in 30 treatment and 30
control oysters collected from 3 containers in each sys-
tem on Days 7, 14 and 21. When the number of live
treatment or control oysters in the 3 randomly chosen
containers was less than 30, all oysters remaining in the
selected containers were sampled.
Perkinsus marinus infection intensity: The number
of parasites per gram of oyster tissue was determined
using the whole-oyster procedure as described by
Fisher & Oliver (1996) and modified by Coates et al.
(1999). The whole-oyster procedure, although labor
intensive, provides an accurate measure of infection
intensity in individual oysters (Bushek et al. 1994). All
chemicals were from Sigma Chemical unless other-
wise indicated. Briefly, each oyster was weighed
and homogenized in alternate fluid thioglycollate me-
dium (ARFTM) supplemented with 16 g marine salts
(Hawaiian Marine Imports) and 5% of a commercial
lipid concentrate 100×at a ratio of 1 g oyster tissue per
50 ml of ARFTM. After 1 wk of incubation in ARFTM,
samples were centrifuged at 1500 ×gfor 10 min and
the supernatant discarded. The resulting pellets were
incubated in 2 N NaOH at 60°C to digest oyster tissues,
leaving the parasites intact. The samples were rinsed
with 0.1 M phosphate buffer saline containing 0.5 mg
ml–1 of bovine serum albumin to prevent parasite
clumping. Samples were then serially diluted in
96-well plates and parasites stained with Lugol’s solu-
tion. The number of parasites was counted from wells
containing 100 to 400 parasites (i.e. hypnospores) with
an inverted microscope at a magnification of 200 ×.
Infection intensity of individual oysters is reported as
number of parasites per gram of oyster tissue.
Condition index: A 10 ml aliquot of oyster tissue
homogenate in ARFTM was dried at 65°C for 48 h and
the dry weight determined by subtracting the weight
of ARFTM only. The dry weight for the whole oyster
was calculated based on the total volume of homoge-
nized tissue in ARFTM. The ratio of the dry weight of
tissue to the dry weight of the shell was calculated and
multiplied by 100 to determine oyster condition index
(CI). This index has been recommended for measuring
the condition of adult oysters and other bivalves (Mann
1978, Lucas & Beninger 1985).
Oyster plasma osmolality: Oyster plasma osmolality
was measured because plasma thoroughly bathes oys-
ter tissues where Perkinsus marinus proliferate extra-
cellularly. Oyster hemolymph (0.2 ml) was withdrawn
167
Mar Ecol Prog Ser 248: 165–176, 2003
from the pericardial cavity using a 27 gauge needle, fol-
lowing careful removal of the shell of all oysters sam-
pled in July and December. The sampled hemolymph
was immediately transferred into vials on ice and the
cell-free hemolymph or plasma was obtained by cen-
trifugation of hemolymph at 600 ×gfor 15 min at 4°C.
The osmolality of the plasma was measured using a va-
por pressure osmometer (Wescor). During the July 2001
experiment, we measured osmolality on Days 2, 4 and 6
using 10 extra oysters in a separate container each time
in order to examine the immediate change in plasma
osmolality as a response to the simulated freshet event.
Statistical analyses: Results of Perkinsus marinus in-
fection intensity, condition index and osmolality for
each season were analyzed using a 2-factor analysis of
variance (i.e. treatment and sampling time) followed by
Student-Newman-Keul’s test (SNK) when significant
differences were found (p < 0.05) (Zar 1984). To test for
seasonal differences in initial P. marinus infection levels
(i.e. pre-freshet event), we ran a 1-factor ANOVA fol-
lowed by SNK when significant differences were found
(p < 0.05). All data were checked for assumptions of
ANOVA and transformed as needed. Data for P. mari-
nus body burden were log transformed to achieve nor-
mality and homogeneity of variance. Mortality data were
analyzed as percent mortality using chi-square tests.
Field study. Oysters and site: Ten eastern oysters
(6 to 12 cm) were collected monthly from September
2000 through February 2002 from 2 sampling stations
near the Caloosahatchee River, Florida. Piney Point
sampling station is located approximately 2 km up-
stream from the river mouth while the more estuarine
influenced station, Bird Island, is located approximately
6km downstream from the river mouth (Fig. 1). The
total length of the river, from the upstream weir to the
mouth of the river before opening into San Carlos Bay,
and ultimately into the Gulf of Mexico, is 42 km. Dis-
crete measures of temperature and salinity data at both
sampling sites were obtained on each day of sampling.
Salinity data were also obtained throughout the study
from continuous water quality monitoring stations
maintained by the South Florida Water Management
District (SFWMD) at Shell point, located at the river
mouth, also at Fort Myers, 20 km upstream of the river
mouth, and in Sanibel, 10 km southwest of the river in
San Carlos Bay. Daily freshwater discharge data via the
weir from Lake Okeechobee into the Caloosahatchee
River were also obtained from the SFWMD (Fig. 1).
Perkinsus marinus weighted prevalence: Oysters
collected monthly were assayed for the presence of P.
marinus using Ray’s fluid thioglycollate medium (FTM)
technique (Ray 1954). Samples of gill and digestive
diverticulum were incubated in FTM for 5 d, smeared
on a slide and the enlarged parasites stained with
Lugol’s solution. The intensity of infection was rated
according to the categories of Ray (1954) by estimating
the percentage of tissue occupied by the parasite, with
0 = no infection, 1 = light infection, 3 = moderate infec-
tion and 5 = heavy infection. Weighted prevalence was
calculated by averaging the intensity of infection of all
oysters sampled monthly (Mackin 1962, Ragone Calvo
& Burreson 1994). This technique, although less sen-
sitive than the whole-body burden assay described
above, is correlated with the body-burden assay and
enables processing of high numbers of samples re-
quired in field studies (Bushek et al. 1994).
Condition index: The condition index of oysters col-
lected monthly was determined by calculating the ratio
of the dry weight of the tissue to the dry weight of shell
and multiplying this ration by 100 as recommended by
Lucas & Beninger (1985).
Juvenile oyster growth and mortality: Two hundred
juvenile oysters (17 mm mean size) were placed in wire
mesh cages (5 ×5 mm mesh size) at each sampling site
in August 2000. Growth and mortality of 50 randomly
selected oysters were determined monthly.
Statistical analyses: Monthly differences for weighted
prevalence of Perkinsus marinus infection, condition
index and growth were tested using a 1-factor ANOVA
followed by Tukey’s multiple comparison test when
significant differences were found (p < 0.05). Data
were tested for normality and homogeneity of variance
and transformed as necessary.
RESULTS
Laboratory experiment
Perkinsus marinus infection intensities
In April, a significant interaction in Perkinsus mari-
nus infection intensities was found between the type of
treatment (i.e. control and freshet) and the time of sam-
pling (i.e. 7, 14 and 21 d) (p < 0.0001). P. marinus infec-
tion intensities of oysters exposed to the freshet were
significantly lower than the infection intensities of con-
trol oysters on Days 14 and 21 (Fig. 2). There were sig-
nificant decreases in infection intensities in oysters
exposed to the freshet from Day 7 (4.70 ×104±6.00 ×
104parasites g–1 wet tissue) to Day 14 (1.17 ×104±
1.30 ×105parasites g–1 wet tissue), and from Day 7 to
Day 21 (4.35 ×103±9.20 ×103parasites g–1 wet tissue).
Initial infection intensity of oysters sampled prior to the
freshet was 2.09 ×105±1.70 ×105parasites g–1 wet tis-
sue. The infection intensity of control oysters was sig-
nificantly higher on Day 14 (5.04 ×105±6.5 ×105par-
asites g–1 wet tissue) than on Day 7 (8.84 ×104±9.8 ×
104parasites g–1 wet tissue) or Day 21 (3.96 ×105±
4.8 ×105parasites g–1 wet tissue) (Fig. 2).
168
La Peyre et al.: Effect of freshets on eastern oysters infected with P. marinus
In July, a significant interaction in Perkinsus mari-
nus infection intensities was found between type of
treatment and time of sampling (p < 0.001). P. mari-
nus infection intensities of oysters exposed to the
freshet were significantly lower than the infection
intensities of control oysters on Days 7, 14 and 21
(Fig. 2). Infection intensities of treatment oysters
were significantly reduced from Day 7 (3.54 ×105±
5.90 ×105) to Day 14 (7.22 ×103±1.10 ×104para-
sites g–1 wet tissue) and Day 21 (2.37 ×104±5.10 ×
104parasites g–1 wet tissue). Initial infection inten-
sity of oysters sampled prior to the freshet was 7.06 ×
105±1.82 ×106parasites g–1 wet tissue. The infec-
tion intensity of control oysters tended to increase
with time from 3.52 ×106±6.60 ×106parasites g–1
wet tissue on Day 7 to 4.75 ×106±8.70 ×106para-
sites g–1 wet tissue on Day 14, and 6.95 ×106±9.70 ×
106parasites g–1 wet tissue on Day 21.
In December, no interaction in Perkinsus marinus
infection intensities was found between type of treat-
ment and time of sampling. P. marinus infection inten-
sities of oysters exposed to the freshet were signifi-
cantly lower than control oysters (p < 0.003) (Fig. 2).
No significant effect of time of sampling was found.
Initial infection intensities in oysters sampled prior to
the freshet was 6.66 ×105±1.29 ×106parasites g–1 wet
tissue.
Comparison of initial Perkinsus marinus infection
intensities in oysters sampled prior to the simulated
freshet events revealed that they were not significantly
different between season (p = 0.07), although oysters
collected in April had approximately 3-fold less infec-
tion intensities (2.09 ×105±1.70 ×105) than oysters
collected in July (7.06 ×105±1.82 ×106) or December
(6.66 ×105±1.29 ×106).
Oyster mortality
No significant difference in cumulative oyster mor-
tality was found between oysters exposed to freshet
and control oysters in April or December (Fig. 3). In
July, the cumulative mortality of oysters exposed to the
freshet (69%) was significantly higher than that of
control oysters (12%) (Fig. 3). There was a significant
difference in cumulative mortality between oysters
exposed to a freshet in April (1.6%), July (69%) and
December (11%). Likewise, there was a significant dif-
ference in mortality between control oysters in April
(0.8%), July (12%) and December (5 %).
169
A
B
Fig. 1. (A) Stylized map denoting field
site and nearby South Florida Water
Management District (SFWMD) water
quality monitoring stations. (B) Weekly
salinities at the Fort Myers (20 km from
the mouth of the River), Shell Point
(mouth of the River) and Sanibel
(10 km from the mouth of the river in
San Carlos Bay) SFWMD water quality
monitoring stations and freshwater dis-
charge via weir from Lake Okeechobee
into the Caloosahatchee River (in cfs
[cubic feet per second] = 0.02832 m3
s–1) from January 2000 to February
2002. Piney Point, our upstream field
study site, is 2 km upstream of the river
mouth, while Bird Island, our down-
stream field study site, is 6 km down-
stream of the river mouth
Mar Ecol Prog Ser 248: 165–176, 2003
Oyster condition index
In April, significant interaction in oyster condition
index was found between type of treatment and time
of sampling (p < 0.0001). No significant differences
existed in condition index between oysters exposed to
the freshet and control oysters until Day 21 (Fig. 4). On
Day 21, oysters exposed to the freshet had a high con-
dition index of 2.9 ± 1.00, and control oysters reached a
low condition index of 1.9 ± 1.03. Initial condition index
of oysters sampled prior to the freshet was 2.67 ± 0.56.
In July, a significant interaction was found between
type of treatment and time of sampling for the oyster
condition index (p < 0.0001). Oysters exposed to the
freshet experienced a rapid and significant decline in
condition index from Day 7 (2.40 ± 0.99) to Day 14 (1.50 ±
0.58) and from Day 14 to Day 21 (0.8 ± 0.27) (Fig. 4).
Release of gametes was observed in the tank (i.e. milky
water) in oysters exposed to the freshet. Initial oysters
had a mean condition index of 1.82 ± 1.59 from which
control oysters did not deviate significantly on Day 7
(2.60 ± 1.01), Day 14 (2.40 ± 0.82) or Day 21 (2.20 ± 0.83).
In December, no significant difference in condition
index was found between control oysters and oysters
exposed to the freshet (Fig. 4). Condition index in these
groups of oysters ranged from 1.54 ± 0.44 to 1.79 ±
0.48, and the initial condition index of oysters sampled
prior to the freshet (1.73 ± 0.39) was in this range.
170
Fig. 2. Infection intensities of control oysters and in oysters
exposed to freshet events in the spring, summer and winter,
sampled on Days 7, 14, and 21 of the simulated freshet event.
Within each seasonal graph, bars with different letters were
significantly different (p < 0.05). Error bars represent +SD
Fig. 3. Cumulative mortality of control oysters and in oysters
exposed to a simulated freshet event in the spring, summer
and winter. Within each season, bars with different letters
were significantly different (p < 0.05). Seasonal mortalities
were not statistically compared
Fig. 4. Condition index of control oysters and in oysters ex-
posed to freshet events in the spring, summer and winter,
sampled on Days 7, 14 and 21 of the simulated freshet event.
Within each seasonal graph, bars with different letters were
significantly different (p < 0.05). Error bars represent +SD
La Peyre et al.: Effect of freshets on eastern oysters infected with P. marinus
Plasma osmolality
In July, a significant interaction was found between
the type of treatment and the time of sampling for oys-
ter plasma osmolality (p < 0.0001). Plasma osmolality of
oysters exposed to the freshet was significantly lower
than plasma osmolality of control oysters on Days 7, 14
and 21; Fig. 5). There was a significant decrease in
plasma osmolality of oysters exposed to the freshet
between Day 7 (218 ± 44 mOsm kg–1) and Day 21
(101 ± 29 mOsm kg–1). Initial plasma osmolality of oys-
ters sampled prior to the freshet was 619 ± 116 mOsm
kg–1. In the first week after the simulated freshet event,
plasma osmolality showed a rapid reduction to 268 ±
8mOsm kg–1 on Day 2 and 196 ± 26 mOsm kg–1 by
Day 4 (Fig. 6).
In December, no significant interaction in oyster
plasma osmolality was found between the type of
treatment and the time of sampling. Plasma osmolality
of oysters exposed to the freshet was significantly
lower than in control oysters (p < 0.003) (Fig. 5). Plasma
osmolality in oysters exposed to the freshet ranged
between 330 ± 127 mOs kg–1 and 275 ± 71 mOsm kg–1,
and plasma osmolality of control oysters ranged from
679 ± 15 mOsm kg–1 to 702 ± 51 mOsm kg–1.
No significant effect of time of sampling was found.
Initial oyster plasma osmolality in oysters sampled
prior to the freshet was 663 ± 57 mOsm kg–1.
Field study
Water temperature and salinity
The discrete measures of temperature and salinity
taken at the collection sites on sample days ranged
between 16 and 31°C and 3 and 39 ppt respectively
(Fig. 6). Daily salinity data from the closest SFWMD
water quality monitoring stations suggest that salinity
at Piney Point was lower than 8 ppt and may have
reached 0 ppt on certain days between August and
November 2001 (Fig. 1). Salinities at the Shell Point
monitoring station appear to track closely salinities
at the Sanibel monitoring station, but demonstrate
amuch greater freshening to freshwater releases.
Increased freshwater flow at the weir in June 2000 and
October 2000 was followed by a decrease in salinity to
a low of 12.5 and 10.5 ppt at the Shell point monitoring
station, and to a low of 27 and 23.5 ppt at the Sanibel
monitoring station. An even greater decrease in salin-
ity was measured at the Sanibel monitoring station
from mid-July to mid-October 2001, with a low of
16 ppt recorded in September 2001. Unfortunately, the
water quality probe at Shell point monitoring station
was not functioning during that period of time. How-
ever, following the trend observed before July 2001
between salinities at the Sanibel and Shell Point moni-
toring stations, it is likely that salinities at the Shell
Point monitoring station and at Piney Point sample
station, 2 km upstream, would have dropped to near
freshwater for at least a few weeks in September 2001.
Similarly, it is likely that salinity at Bird Island would
have dropped below 10 ppt, and possibly 5 ppt during
this time. This contention is supported by a salinity
model developed to predict the effects of the weir from
Lake Okeechobee on the downstream river and estu-
ary (Bierman 1993). Specifically, the model predicted
that moderate mean monthly discharges of only 2000
cfs (cubic feet per second = ca. 57 m3s–1) would result
in much of the river upstream of Shell Point becoming
nearly fresh water, while inflows greater than 4000 cfs
(= ca. 113 m3s–1) would cause the entire estuary
upstream of Shell Point to become fresh water (Bier-
man 1993).
171
Fig. 5. Osmolality of plasma collected from control oysters and
from oysters exposed to freshet events in summer and winter
sampled on Days 0, 7, 14 and 21 of the simulated freshet
event. Groups of lines with different letters were significantly
different (p < 0.05)
Fig. 6. Plasma osmolality measured prior to the freshet event
(Day 0), and at Days 2, 4 and 6 following the freshet event.
Plasma osmolality was measured in 10 oysters exposed to the
simulated freshet on each day during July 2001. Error bars
represent ± SD
Mar Ecol Prog Ser 248: 165–176, 2003
Perkinsus marinus weighted prevalence
Significant monthly differences were noted in
Perkinsus marinus weighted prevalence (p < 0.001) at
both stations, but no seasonal trends were noted. The
upstream station, Piney Point, had significantly lower
weighted prevalence (mean = 0.20) compared to the
downstream station, Bird Island (mean = 0.46) (Fig. 7).
Condition index
Significant differences were observed in monthly
oyster condition index (p < 0.001). Condition index at
both sampling stations was higher during the cooler
months into spring (January to May) and decreased
from May to October (Fig. 8).
Juvenile oyster mortality and growth
No significant mortality was observed in juvenile
oysters between September 2000 and February 2002.
Mortality of juvenile oysters during the study period,
including after the freshets from July to October
2001, was less than 10%. At the same time, juvenile
oysters showed significant growth (p < 0.0001) dur-
ing the study period, increasing from a mean size of
17 mm in August 2000 to a mean size of 63 mm in
February 2002 at Piney Point, and a mean size of
54 mm at Bird Island (Fig. 9). The mean growth rate
of juvenile oysters at Piney Point was 2.42 ± 8.53 mm
mo–1 compared to 1.95 ± 8.53 mm mo–1 at Bird
Island.
DISCUSSION
Past studies demonstrated that lowered
salinities (below 12 ppt) retarded Perkinsus
marinus disease development in oysters (Ray
1954, Andrews & Hewatt 1957, Chu et al.
1993, Ragone & Burreson 1993). Despite field
observations documenting a lack of P. mari-
nus infection in areas prone to freshet events,
no study, field or laboratory, that we are
aware of has explicitly determined experi-
mentally the impact of freshets on P. marinus
infection intensities in oysters. This con-
trolled quantitative assessment of the effects
of freshet events on both P. marinus infection
intensities and oyster condition in the labora-
tory provides information on the effect of
single freshet events and their potential for
use in controlling P. marinus infections, while
maintaining the viability of oyster cultures.
This field study documents the links between
a highly variable salinity regime marked by
seasonal freshet events, and a maintained
low P. marinus infection level.
Effects of freshet events on Perkinsus
marinus infections and oysters
Inherent to a study examining the response
of both a host and a parasite to external con-
ditions is the fact that the response of the host
can have impacts on the conditions that the
parasite experiences. Therefore, despite evi-
dence demonstrating that acute exposure of
in vitro cultured Perkinsus marinus, trans-
ferred from 22 to 0 ppt, resulted in >99%
mortality (Burreson et al. 1994), the oyster’s
172
Fig. 7. Monthly Perkinsus marinus weighted prevalence, salinity and
temperature at Piney Point and Bird Island, the field study sites in the
Caloosahatchee estuary, from September 2000 to February 2002. Ten
randomly selected oysters were sampled every month at both sites to
determine P. marinus weighted prevalence. Temperature and salinity
values were taken at the surface during sampling
La Peyre et al.: Effect of freshets on eastern oysters infected with P. marinus
response to the freshet event needed to be examined
to understand the conditions that P. marinus experi-
enced in vivo. Along with oyster mortality rates, 2 clues
from the oyster’s response that provide further infor-
mation on the environment that the parasite had to
deal with were examined: oyster plasma osmolality
and oyster condition index.
The controlled laboratory study indicated that Per-
kinsus marinus infection intensities in eastern oysters
were significantly reduced by all 3 simulated freshet
events. The simulated freshet events failed, however,
to completely eliminate P. marinus infections. The lack
of complete P. marinus elimination was likely due to a
combination of factors, including the failure of plasma
to reach very low osmolality (< 50 mOsm kg–1) and the
acclimation of surviving parasites to lowered plasma
osmolalities. Mean plasma osmolalities of
oysters exposed to freshets ranged between
274 and 330 mOsm kg–1 in December and
between 218 and 101 mOsm kg–1 in July.
Interestingly, the largest decrease in P. mari-
nus infection intensity (by 99%) following a
freshet occurred in July when plasma osmo-
lality was lowest, while P. marinus infection
intensity was only reduced to 66% in oysters
exposed to a freshet in December. The result
must be interpreted with caution, however,
because it is also possible that the greater
mortality of oysters in July could have
contributed to the significant decrease in
infection intensities if oysters with heavier
infection intensities died at a faster rate than
oysters with lower infection intensities.
Most of the reduction of Perkinsus marinus
infection intensity is likely due to the rapid
decrease in oyster plasma osmolality. In vitro
studies with cultured P. marinus have shown
that the viability of parasites measured 24 h
after their transfer from a salinity of 22 ppt
(~660 mOsm kg–1) to salinities of 9 ppt
(~270 mOsm kg–1) was reduced to 57%. Via-
bility was 30% after transfer from 22 to 6 ppt
(~180 mOsm kg–1) and 10% after transfer
from 22 to 3 ppt (~90 mOsm kg–1). When
measured every other day, plasma osmolality
was found to decrease rapidly following the
freshet event, and remained low throughout
the freshet event. This decrease in plasma
osmolality, combined with past in vitro find-
ings using cultured P. marinus, explains the
range of reduction of P. marinus infection
intensities observed in our in vivo studies.
The grossest measure of the effects of a
simulated freshet event on Crassostrea virgi-
nica is the mortality rate of oysters. There was
a pronounced seasonal effect of the freshet events on
oyster mortality in the laboratory experiments. The
mortality of oysters exposed to the freshet in April and
December remained low throughout each experiment
and was not significantly different than the mortality
of control oysters. In contrast, oysters exposed to the
freshet in July experienced 69% cumulative mortality
by Day 21 compared to 12% cumulative mortality of
control oysters. It is likely that some confounding fac-
tor, such as high temperature, high initial infection
intensities and spawning stress of oysters collected in
July, led to the high mortality. The laboratory results
suggest that oysters with moderate to heavy Perkinsus
marinus infection intensities in the field would be able
to survive freshet events (at least for 21 d) in winter
and spring but not in summer. This would agree with
173
Fig. 8. Condition index (CI) of oysters at our field study sites, Piney Point
(PP) and Bird Island (BI), in the Caloosahatchee estuary. Ten oysters from
each location were sampled monthly from September 2000 to February
2002 and CI determined according to the procedure of Lucas & Beninger
(1985). Results presented are monthly means ± SD
Fig. 9. Mean size (±SD) of juvenile oysters at our sampling sites, Piney
Point (PP) and Bird Island (BI), in the Caloosahatchee estuary. Two hun-
dred juvenile oysters were placed in a 0.5 m2wire mesh cage (0.5 ×
0.5 mm mesh size) at each sampling station. The length of 50 randomly
selected oysters was measured monthly at each field site from September
2000 to February 2002
Mar Ecol Prog Ser 248: 165–176, 2003
field surveys that have noted that C. virginica can
survive salinities below 5 ppt, especially when water
temperatures are low (Butler 1949, Loosanoff 1953,
Andrews et al. 1959, Galtsoff 1964, Austin et al. 1993,
Winstead 1995).
In the field study, oysters at our more freshwater
dominated sampling station, Piney Point, encountered
salinities of less than 10 ppt for at least 3 mo, mid-July
through mid-October 2001, and probably salinities less
than 3 to 5 ppt for at least 2 to 3 wk in September 2001,
with no significant mortalities in deployed juvenile
oysters. The overall mortality in deployed juvenile
oysters at both sampling stations was less than 10% at
the end of the study. While the mortality of wild adult
oysters collected at both sites was not measured, it is
likely to be less than juvenile oysters, since juvenile
oysters are generally more sensitive to freshets and
other stress factors than adult oysters. The much lower
mortality of Florida oysters in summer 2001, compared
to the Louisiana oysters collected in July 2001 and
exposed to a simulated freshet in the laboratory, could
be due to their much lower initial Perkinsus marinus
infection intensities.
The laboratory experiment results also indicated that
a freshet event of up to 3 wk in length in spring or win-
ter would not adversely affect the condition index of
oysters. April- and December-simulated freshets re-
sulted in no significant differences in oyster condition
index between control and treatment, except for on
Day 21 in April, when control oysters had a lowered
condition index. In contrast, a simulated freshet in July
may exacerbate already stressed oysters (from high
Perkinsus marinus infection intensities, high tempera-
tures and spawning) and be detrimental to the oyster
populations. Clearly, in July, from Day 7 onwards,
there was a reduction in condition index indicating
that the oysters were stressed. Part of the reduction in
condition index is likely due to spawning, since the
release of gametes was observed in the tank following
the freshet event. Oysters in Gulf waters have an
extended spawning season from April to October in
this subtropical region, with gametogenic recycling
and sometimes up to 3 spawning events occurring
during this period (Hayes & Menzel 1981, Supan &
Wilson 2001).
Potential management implications
All natural systems exhibit environmental variabil-
ity, ranging from predictable seasonal variations to
extreme El Niño Southern Oscillation events to in-
tended and unintended human effects. These events,
acting on ‘primary’ environmental variables (i.e. salin-
ity, temperature), have been shown to affect fish stocks
(Houde 1997), survival of aquatic organisms (Hobday
& Boehlert 2001), habitat use (Peebles & Flannery
1992), as well as impact parasite survival (Bataller &
Boghen 2000). Several studies with oysters have found
that environmental variability works to eliminate para-
sites without detrimental effects on the oysters (Haskin
& Ford 1982, Ford 1985, Ford & Haskin 1988, Bataller &
Boghen 2000), suggesting that managed environmen-
tal variability, such as freshet events, has the potential
to be a valuable management tool.
In past studies examining the effects of low salinities
on Perkinsus marinus, results indicated that a reduction
of P. marinus associated with low salinities was quickly
replaced by a rapid proliferation of P. marinus once
more favorable conditions for the parasite (i.e. higher
salinity) returned (Ragone Calvo & Burreson 1994,
Fisher & Oliver 1996, Ford 1996, Ford et al. 1999). For
freshet events to become a useful management tool in
reducing oyster mortality from P.marinus, P. marinus
must either be eliminated in order to prevent its likely
rapid proliferation once the freshet event is over, or the
freshet events must be repeated in order to maintain it
at low levels. Clearly, a single freshet event, similar to
our simulated ones, lasting up to 3 wk will not have any
significant or lasting effects on the intensity of P. mari-
nus infection in an oyster population moderately to
heavily infected with the parasite. The use of repeated
freshet events may be worth investigating.
A number of investigators have noted that locations
characterized by the regular occurrence of freshet
events lack significant Perkinsus marinus infections
(Soniat & Gauthier 1989, Thurston et al. 2001, Volety et
al. 2001a,b). Interestingly, higher oyster densities and
overall oyster bar growth in Apalachicola Estuary were
reported to occur in the vicinity of the confluence of
high salinity water and river-dominated low salinity
water (Livingston et al. 2000), hence in areas prone to
high variation in salinity. In this instance, oyster growth
was positively correlated with variation (i.e. standard
deviation) in salinity. Further investigation into the ac-
tual site characteristics in terms of timing, frequency
and length of these freshet events may provide some
clues as to the extent of environmental variability that
may be correlated, in these instances, with the lack of
significant P. marinus infections in the oysters.
Based on our discrete monthly samples, depicting an
environment characterized by high water tempera-
tures (16 to 31°C) and salinities (3 to 39 ppt) at our sam-
pling sites in the Caloosahatchee estuary, much higher
infection intensities in oysters, similar to other studies,
were expected (Ragone Calvo & Burreson 1994, Soniat
1996). In the warm water months, from May to October
when temperatures exceed 28°C, salinity was variable,
and likely reached extreme lows (below 5 ppt at Piney
Point, and below 8 ppt at Bird Island) several times due
174
La Peyre et al.: Effect of freshets on eastern oysters infected with P. marinus
to the combination of freshwater releases from Lake
Okeechobee and heavy rainfall in the basin (Bierman
1993). The numerous freshwater releases from the
weir and the high rainfall in the Caloosahatchee River
provide a likely explanation for the low Perkinsus mar-
inus weighted prevalences we detected, and the lack
of a strong seasonal (temperature-related) pattern. P.
marinus weighted prevalence in oysters at our fresh
water site, Piney Point, appeared to follow a trend with
decreasing prevalence from 0.5 to 0.1, as salinity was
drastically reduced during the summer of 2001. Our
more saline site, Bird Island, did not show a significant
decline in weighted prevalence, but rather, failed to
exhibit any significant peak in weighted prevalence,
likely because salinity was drastically reduced in 2001
during the months with warmer temperatures.
Similar decreases in Perkinsus marinus weighted
prevalence have also been noted to occur concomi-
tantly with decreased salinities in other southwest
Florida estuaries (Thurston et al. 2001, Volety et al.
2001a,b). For example, P. marinus infection intensities
decreased in Blackwater River, Henderson Creek, and
Faka-Union estuaries during summer months, a period
characterized by heavy rains and the release of fresh-
water from upstream areas resulting in several freshet
events and extremely low salinities. P. marinus infec-
tions decreased from 0.7, 0.8, and 0.6 to 0.01, 0.08, and
0.05 in Blackwater River, Henderson Creek, and Faka-
Union respectively, from July to September 2001.
The overall success of the oyster industry depends on
our ability to manage oyster populations in estuaries
where the parasite is widespread and abundant.
Andrews & Ray (1988) suggested that management
measures that support the diversion of fresh water into
high salinity areas may be an effective means to control
Perkinsus marinus in the Gulf of Mexico. Some limited
freshwater diversions from the Mississippi River in the
1980s have been cited as being effective in enhancing
oyster production, although it is not clear what the
impacts were on P. marinus (see Andrews & Ray 1988).
Recent data from the large Caernarvon diversion water
control project in Louisiana demonstrated a significant
increase in oyster production associated with increased
freshwater flows (Villarubia 19981). Most importantly,
data from our field study seem to support the idea that
repetitive and well-timed freshet events can prevent in-
fection of oysters with P. marinus, or maintain P. marinus
infection to non-lethal intensities (e.g. <106parasites g–1
wet tissue) in oyster populations. The use of an adaptive
management approach involving control of freshwater
inflows could be invaluable to the oyster industry.
Acknowledgements. We thank Casey Barroco, Tracy Brown,
Chawn-Hong Foo and Jessica Stevenson for technical assis-
tance during the laboratory experiments. We thank John
Supan for providing oysters. The laboratory experiments
were funded by the Louisiana Sea Grant College Program,
the National Sea Grant Gulf Oyster Industry Program and the
Louisiana Department of Wildlife and Fisheries through the
USGS Louisiana Fish and Wildlife Cooperative Research Unit.
Funding for the field component of this study was provided by
the South Florida Water Management District. Thanks are
also due to Sharon Thurston for technical assistance during
the field study. We thank 3 anonymous reviewers for helpful
comments.
LITERATURE CITED
Andrews JD, Hewatt WG (1957) Oyster mortality studies in Vir-
ginia. II. The fungus disease caused by Dermocystidium mar-
inum in oysters in Chesapeake Bay. Ecol Monogr 27:1–26
Andrews JD, Ray SR (1988) Management strategies to control
the disease caused by Perkinsus marinus. In: Fisher WS
(ed) Disease processes in marine bivalve molluscs. Ameri-
can Fisheries Society, Special Publication 18, Bethesda,
p257–264
Andrews JD, Haven D, Quayle DB (1959) Freshwater kill of
oysters (Crassostrea virginica) in James River, Virginia,
1958. Proc Natl Shellfish Assoc 49:29–49
Austin H, Haven DS, Moustafa MS (1993) The relationship
between trends in a condition index of the American oys-
ter, Crassostrea virginica, and environmental parameters
in three Virginia estuaries. Estuaries 16(2):362–374
Bataller E, Boghen AD (2000) Elimination of the gill worm
Urastoma cyprinae (Graff) from the eastern oyster Crasso-
strea virginica (Gmelin) using different salinity-tempera-
ture combinations. Aquaculture 182(3-4):199–208
Bierman V (1993) Performance Report for the Caloosahatchee
Estuary salinity modeling. South Florida Water Manage-
ment District (SFWMD) expert assistance contract, Limno-
Tech, Ann Arbor, MI
Burreson EM, Ragone Calvo LM (1996) Epizootiology of Perkin-
sus marinus disease of oysters in Chesapeake Bay, with
emphasis on data since 1985. J Shellfish Res 15(1):17–34
Burreson EM, Ragone Calvo LM, La Peyre JF, Counts F,
Paynter KT Jr (1994) Acute osmotic tolerance of cultured
cells of the oyster pathogen Perkinsus marinus (Apicom-
plexa: Perkinsida). Comp Biochem Physiol 109A(3):575– 582
Bushek D, Ford SE, Allen SK Jr (1994) Evaluation of methods
using Ray’s fluid thioglycollate medium for diagnosis of
Perkinsus marinus infection in the eastern oyster, Cras-
sostrea virginica. Annu Rev Fish Dis 4:201–217
Butler PA (1949) Gametogenesis in the oyster under condi-
tions of depressed salinity. Biol Bull (Woods Hole) 96:
263–269
Chu FE, Greene KH (1989) Effect of temperature and salinity
on in vitro culture of the oyster pathogen, Perkinsus
marinus (Apicomplexa: Perkinsea). J Invertebr Pathol 53:
260–268
Chu FE, La Peyre JF (1993) Perkinsus marinus susceptibility
and defense-related activities in eastern oysters Cras-
sostrea virginica: temperature effects. Dis Aquat Org 16:
223–234
Chu FE, La Peyre JF, Burreson CS (1993) Perkinsus marinus
infection and potential defense-related activities in east-
ern oysters, Crassostrea virginica: salinity effects. J Inver-
tebr Pathol 62:226–232
Coates GM, Cooper RK, La Peyre JF (1999) Improvement of
175
1Ecosystem response to a freshwater diversion: the Caer-
narvon experience; available at www.lacoast.gov/programs/
Caernarvon/index.htm
Mar Ecol Prog Ser 248: 165–176, 2003
the whole-oyster procedure for enumerating Perkinsus
marinus in oyster tissues. J Shellfish Res 18:328
Craig A, Powell EN, Fay RR, Brooks JM (1989) Distribution
of Perkinsus marinus in Gulf Coast oyster populations.
Estuaries 12(2):82– 91
Fisher WS, Oliver LM (1996) A whole-oyster procedure for
diagnosis of Perkinsus marinus disease using Ray’s fluid
thioglycollate culture medium. J Shellfish Res 15(1):
109–118
Fisher WS, Winstead JT, Oliver LM, Edmiston HL, Bailey GO
(1996) Physiologic variability of eastern oysters from
Apalachicola Bay, Florida. J Shellfish Res 15(3):543–553
Ford SE (1985) Effects of salinity on survival of the MSX
parasite Haplosporidium nelsoni (Haskin, Stauber, and
Mackin) in oysters. J Shellfish Res 5(2):85– 90
Ford SE (1996) Range extension by the oyster parasite Perkin-
sus marinus into the northeastern United States: response
to climate change? J Shellfish Res 15(1):45– 56
Ford SE, Haskin HH (1988) Comparison of in vitro salinity
tolerance of the oyster parasite, Haplosporidium nelsoni
(MSX) and hemocytes from the host, Crassostrea virgi-
nica. Comp Biochem Physiol 90A(1):183–187
Ford SE, Schotthoefer A, Spruck C (1999) In vivo dynamics of
the microparasite Perkinsus marinus during progression
and regression of infections in eastern oysters. J Parasitol
85(2):273–282
Galtsoff PS (1964) The American oyster Crassostrea virginica
Gmelin. US Fish Bull 64:1–480
Haskin HH, Ford SE (1982) Haplosporidium nelsoni (MSX) on
Delaware Bay seed oyster beds: a host-parasite relation-
ship along a salinity gradient. J Invertebr Pathol 40(3):
388– 405
Hayes PF, Menzel W (1981) The reproductive cycle of early
setting Crassostrea virginica (Gmelin) in the northern Gulf
of Mexico, and its implications for population recruitment.
Biol Bull (Woods Hole) 160:80– 88
Hobday AJ, Boehlert GW (2001) The role of coastal ocean
variation in spatial and temporal patterns in survival and
size of coho salmon (Oncorhynchus kisutch). Can J Fish
Aquat Sci 58:2021–2036
Hofstetter RP (1977) Trends in population levels of the Amer-
ican oyster, Crassostrea virginica (Gmelin) on public reefs
in Galveston Bay. Texas Parks and Wildlife Dept, Tech Ser
No. 24, Austin, TX
Houde ED (1997) Patterns and trends in larval-stage growth
and mortality of teleost fish. J Fish Biol 51(Suppl A):52–83
Livingston RJ, Lewis FG, Woodsum GC, Niu XF and 8 others
(2000) Modeling oyster population response to variation in
freshwater input. Estuar Coast Shelf Sci 50:655–672
Loosanoff VL (1953) Behavior of oysters in water of low sal-
inities. Proc Natl Shellfish Assoc 43:135–151
Lucas A, Beninger PG (1985) The use of physiological condi-
tion indices in marine bivalve aquaculture. Aquaculture
44:187–200
Mackin JG (1956) Dermocystidium marinum and salinity.
Proc Natl Shellfish Assoc 46:116–128
Mackin JG (1962) Oyster disease caused by Dermocystidum
marinum and other microorganisms in Louisiana. In:
Mackin JG, Hopkins SH (eds) Studies in oysters in relation
to the oil industry, Vol 7. Publ Inst Mar Sci Univ Texas,
p132–299
Mann R (1978) A comparison of morphometric, biochemical,
and physiological indices of condition in marine bivalve
molluscs. In: Thorp JH, Gibbons JW (eds) Early and envi-
ronmental stress in aquatic systems. US Department of
Energy, Symposium Series (771114), Woods Hole Oceano-
graphic Institute, Woods Hole, MA, p 484– 497
Odum PE (1969) The strategy of ecosystem development.
Science 164:262–269
Peebles EB, Flannery MS (1992) Fish nursery use of the Little
Manatee River estuary (Florida): relationships with fresh-
water discharge. Final Report for Southwest Florida Water
Management District, Tampa Bay Estuary Program, St.
Petersburg, FL
Pickett STA, Parker VT, Fiedler P (1992) The new paradigm in
ecology: implications for conservation biology above the
species level. In: Fiedler P, Jain S (eds) Conservation
biology: the theory and practice of nature conservation,
preservation and management. Chapman & Hall, New
York, p 65–88
Powell EN, Klink JM, Hofmann EE (1996) Modeling diseased
oyster populations. II. Triggering mechanisms for Perkin-
sus marinus epizootics. J Shellfish Res 15:141–165
Ragone LM, Burreson EM (1993) Effect of salinity on infection
progression and pathogenicity of Perkinsus marinus in the
eastern oyster, Crassostrea virginica (Gmelin). J Shellfish
Res 12(1):1–7
Ragone Calvo LM, Burreson EM (1994) Characterization of
overwintering infections of Perkinsus marinus (Apicom-
plexa) in Chesapeake Bay oysters. J Shellfish Res 13:
123–130
Ray SM (1954) Biological studies of Dermocystidium mari-
num, a fungus parasite of oysters. The Rice Institute Pam-
phlet, Special Issue, November 114. The Rice Institute,
Houston, TX
Soniat TM (1985) Changes in levels of infection of oysters by
Perkinsus marinus, with special reference to the inter-
action of temperature and salinity upon parasitism. NE
Gulf Sci 7(2):171–174
Soniat TM (1996) Epizootiology of Perkinsus marinus disease
of eastern oysters in the Gulf of Mexico. J Shellfish Res
15(1):35– 43
Soniat TM, Gauthier JD (1989) The prevalence and intensity
of Perkinsus marinus from the mid northern Gulf of
Mexico, with comments on the relationship of the oyster
parasite to temperature and salinity. Tul Stud Zool Bot 27:
21–27
Supan JE, Wilson CA (2001) Analyses of gonadal cycling
by oyster broodstock, Crassostrea virginica (Gmelin), in
Louisiana. J Shellfish Res 20:215–220
Thurston S, Volety AK, Savarese M, Lindland E, Bankston S,
Grindberg R, Benolkin M (2001) Monitoring the impact
of water management practices on estuarine health. II.
Oyster growth, recruitment and disease. 16th Biennial
Conference of the Estuarine Research Federation, Book of
Abstracts. University of South Florida, Tampa, FL, p 139
Volety AK, Tolley SG, Winstead JT (2001a) Effects of season
and water quality on oysters (Crassostrea virginica) and
associated fish assemblages. 16th Biennial Conference of
the Estuarine Research Federation, Book of Abstracts.
University of South Florida, Tampa, FL, p 145
Volety AK, Savarese M, Tolley SG (2001b) Disease status and
physiological responses of oysters as indicators of water-
shed alteration effects in southwest Florida estuaries.
JShellfish Res 20(1):558
Winstead JT (1995) Digestive tubule atrophy in eastern oys-
ters, Crassostrea virginica (Gmelin, 1791) exposed to sal-
inity and starvation stress. J Shellfish Res 14(1):105–111
Zar JH (1984) Biostatistical analysis. Prentice-Hall, Upper
Saddle River, NJ
176
Editorial responsibility: Otto Kinne (Editor),
Oldendorf/Luhe, Germany
Submitted: June 11, 2002; Accepted: November 19, 2002
Proofs received from author(s): February 14, 2003
... Additionally, the timing of large magnitude changes can further limit or enhance survival during exposure, making the timing of freshwater release just as critical as the magnitude and duration (La Peyre et al., 2009. Extreme decreases in salinity can have a greater impact, even for short durations, when they occur during periods in which oysters normally have maximal gametogenic activity (Andrews et al., 1959;La Peyre et al., 2003Loosanoff, 1952) or if they occur during typical south Florida summer high temperatures, as increased metabolic stress can compound the effects of osmotic stress, amplifying the negative response Rybovich et al., 2016). By contrast, prolonged high salinity events resulting from a lack of sufficient freshwater input can also be stressful to oysters. ...
... Likewise, Loosanoff (1952) reported high mortality rates when low salinity exposure occurred during advanced gametogenesis. Laboratory exposures to 3-week long freshets (salinity drop from 20 to 1 over 48 h), resulted in significant mortality of oysters collected and exposed in the summer (July) but not during winter or spring (December and April) (La Peyre et al., 2003). Similarly, McCarty et al. (2020) observed a doubling of mortality rate when 2-year old oysters were exposed to freshets (2.7) in the summer compared to spring. ...
... Mid-estuary moderate salinity site BI is marked in green. Asterisks (*) mark years with significant mortality events at IC during which no oysters were alive to be collected for analysis temperature and low salinity is amplified in recently settled juvenile oysters and larvae, which experience higher mortality rates under these conditions compared to adult oysters (La Peyre et al., 2003;Volety et al., 2009Volety et al., , 2016. Therefore, acute exposure to extreme low salinity events in the late summer, when peaks in spawning and recruitment are observed, may not only inhibit gametogenesis and induce large scale mortality events in the breeding population but could also increase morality rates in free swimming larvae and newly settled juveniles. ...
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Few estuaries remain unaffected by water management and altered freshwater deliveries. The Caloosahatchee River Estuary is a perfect case study for assessing the impact of altered hydrology on natural oyster reef (Crassostrea virginica) populations. The watershed has been highly modified and greatly enlarged by an artificial connection to Lake Okeechobee. Accordingly, to generate data to support water management recommendations, this study monitored various oyster biometrics over 15 years along the primary salinity gradient. Oyster reef densities were significantly affected by both prolonged high volume freshwater releases creating hyposaline conditions at upstream sites and by a lack of freshwater input creating hypersaline conditions at downstream sites. Low freshwater input led to an increase in disease caused by Perkinsus marinus and predation. Moderate (< 2000 cfs) and properly timed (winter/spring) freshets benefited oysters with increased gametogenesis, good larval mixing, and a reprieve from disease. If high volume freshets occurred in the late summer, extensive mortality occurred at the upstream site due to low salinity. These findings suggest freshwater releases in the late summer, when reproductive stress is at its peak and pelagic larvae are most vulnerable, should be limited to < 2000 cfs, but that longer freshets (1–3 weeks) in the winter and early spring (e.g., December–April) benefit oysters by reducing salinity and lessening disease intensity. Similar strategies can be employed in other managed systems, and patterns regarding the timing of high volume flows are applicable to all estuaries where the management of healthy oyster reefs is a priority.
... Resulting enlarged, spherical parasite hypnospores or pre-zoosporangia measure 10-250 µm, with optically refractile walls that also stain blue-black in 20-30% (v/v) Lugol's iodine by a reaction that does not involve starch ( Figure 2). Alternative Ray's fluid thioglycollate media (ARFTM) that lack agar and includes additional metabolytes are superior for some applications (La Peyre et al. 2003), though ARFTM and RFTM media are treated as equivalent and synonymous in the protocols that follow. ...
... Strong correlations are reported between sample prevalences and infection intensities of individual oysters that were analyzed in parallel by RFTM assays and several qPCR assays (Yarnall et al. 2000, Gauthier et al. 2006. For RFTM methods and reviews, see Ray (1966), Bushek et al. (1994), La Peyre et al. (2003), and Dungan and Bushek (2015). ...
... Parasite cell counts are normalized to recorded host tissue wet weights, and sensitivities reach 1 parasite cell/host when host tissues are analyzed exhaustively (Bushek et al. 1994). The use of ARFTM facilitates differential centrifugation steps of these assays by eliminating agar, a medium component that does not dissolve during alkaline hydrolysis (La Peyre et al. 2003). ...
Chapter
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This is a revision of Reece, KS and CF Dungan. 2006. Perkinsus sp. Infections of Marine Molluscs. In AFS-FHS (American Fisheries Society-Fish Health Section). FHS Blue Book: suggested procedures for the detection and identification of certain finfish and shellfish pathogens, 2016 edition. Bethesda, Maryland.
... In October 2020, near the completion of this study, 20 oysters from each stock (5/basket) were haphazardly collected to determine Perkinsus marinus infection intensity (parasites per gram wet tissue), infection prevalence (number of infected oysters/total number of oysters sampled × 100), and body condition index [100 × dry tissue weight/ (whole weight − shell weight)] as described by La Peyre et al. (2003, 2019b. The P. marinus infection intensity was categorized as either no infection (0 parasites/g wet tissue), light infection (1 to <10 4 parasites/g wet tissue), moderate infection (10 4 to 5 × 10 5 parasites/g wet tissue), or heavy infection (>5 × 10 5 parasites/g wet tissue) . ...
... Oysters at the lower-salinity site experienced both lighter infection intensity and lower overall prevalence of infection compared with the moderate-salinity site, which can be attributed to limited or delayed development of P. marinus at lower salinities (Chu and La Peyre 1993;La Peyre et al. 2003;Ragone Calvo and Burreson 2003;Bushek et al. 2012). Additionally, with higher mortalities seen at the site with lighter infection (LUMCON), we concluded that P. marinus infection was not a leading cause of differing mortality between stocks in this study. ...
Article
Eastern oysters Crassostrea virginica support a critical commercial industry and provide many ecosystem services to coastal estuaries yet are currently threatened by changing estuarine conditions. A changing climate and the effects of river and coastal management are altering freshwater inflows into productive oyster areas, causing more frequent and extreme salinity exposure. Although eastern oysters are tolerant to a wide range of salinity means and variations, more frequent and extreme exposure to low salinity (<5‰) impacts oyster populations and aquaculture operations. This study assessed four Louisiana eastern oyster stocks to explore population‐specific responses to low‐salinity exposure. Hatchery‐produced progeny (10–25 mm) were deployed in baskets kept off‐bottom on longline systems in a low‐salinity (mean ± 1 standard error of the mean daily salinity = 8.7 ± 0.2‰; range = 1.2–19.0‰) and a moderate‐salinity (16.8 ± 0.3‰; 4.8–30.0‰) environment for 1 year, beginning in December 2019, with growth and mortality determined monthly. Significant differences in cumulative mortality between stocks at the end of the study were found at the low‐salinity site, with the greatest increase in cumulative mortality occurring mid‐July to mid‐August. Mortality differences between stocks suggest that some eastern oyster populations (i.e., stocks) may be better suited to low salinity or low‐salinity events than others. This difference may be attributed to similarity between site of origin and grow‐out site conditions and/or to greater salinity variability and therefore higher phenotypic plasticity in some eastern oyster populations compared with others. The identification of oyster stocks able to survive under extreme low‐salinity conditions may facilitate the development of “low‐salinity‐tolerant” broodstock to support aquaculture in areas experiencing and predicted to experience low‐salinity events.
... Average condition indices were generally higher in the lower temperature treatments (18℃ and 21℃) as compared to oyster exposed at 30℃, albeit not statistically so. These results were similar to results reported by La Peyre et al. (2003) where average condition indices of oysters exposed to freshwater were observed to be significantly lower after two weeks of exposure in summer season conditions. ...
... Under winter conditions, La Peyre et al. (2003) observed average condition indices were similar to the controls, indicating that cooler temperatures may benefit the overall health of oysters when exposed to stressful conditions. Volety et al. (2016) observed that condition indices were at their lowest in the late summer/early fall and found a negative correlation with temperature. ...
... Dermo disease risk is largely limited to high and moderate salinity environments (≥ 12 psu) because of the pathogen's environmental limitations (Powell et al. 1996;Levinton et al. 2011). Thus, while oyster gametogenesis is inhibited in extreme low salinity environments, such areas can also serve as a refuge from disease (La Peyre et al. 2003. Documenting Dermo prevalence can not only help to interpret potential mechanisms limiting gametogenesis between high and low salinity sites, but it also provides a critical baseline metric to inform restoration planning and monitoring the health of restored populations. ...
Article
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Low salinity can negatively affect reproduction in estuarine bivalves. The spatial and temporal extents of these effects are important to inform models of population dynamics, environmental risk assessments, restoration efforts, and predictions of climate change effects. A hypothesis of delayed gametogenesis for oysters at low salinity sites was tested relative to their higher salinity counterparts in downstream experimental cages. In 2018, the timing of gametogenesis and spawning was observed June–August for 2-year-old oysters from three distinct ancestries (native, hatchery, aquaculture), outplanted at age 1 month along the salinity gradient (3–30 psu) of a temperate estuary. A second season of data was collected in 2019 from a 3-year-old aquaculture line and mixed-age native adult oysters dredged and transplanted 1 year prior. Dermo was tested in 2019 and prevalence was 1.3% ( n = 240). Gametogenesis and spawning were retarded for all ancestries at low salinity relative to higher salinity sites during July and August. The reverse pattern was found in June, with low salinity sites having more advanced gonad index than at a high salinity site. This difference in average gonad index was 2.65 vs 1.46, respectively, for the native line and 2.62 vs 2.08 for aquaculture. Low salinity seemed to not only induce earlier gametogenesis in June, but also extended the reproductive season relative to higher salinity sites. Among oyster ancestries, the aquaculture line stood out as having 30–48% lower gametogenic synchrony within sites, but only in 2018. Because the native oysters used in this study have been restricted to low salinity conditions for many generations, demonstration of their reproductive plasticity across salinities is notable and broadens the range of potential future restoration strategies.
... The Aquaculture Zone was designed to capture conditions best suited for high oyster growth and low mortality while de-emphasizing reproduction and predation (oysters would be grown in predatorexcluding baskets from hatchery provided seed). The annual mean salinity was defined as equal to or greater than 12 for at least four out of five years to represent ideal conditions for growth and survival (La Peyre et al., 2003;Bushek et al., 2012;Lowe et al., 2017). No maximum salinity was applied as the primary threat from higher salinity would be mortality from Perkinsus marinus and, given the fast growth and harvest within less than 1 year of oysters grown off-bottom in Louisiana waters, P. marinus is not generally a concern Leonhardt et al., 2017). ...
Article
Eastern oysters (Crassostrea virginica) are a critical ecological and commercial resource in the northern Gulf of Mexico facing changing environmental conditions from river management and climate change. In Louisiana, USA, development of restored reefs, and off-bottom aquaculture would benefit from the identification of locations supportive of sustainable oyster populations (i.e., metapopulations) and high consistent production. This study defines four oyster resource zones across coastal Louisiana based on environmental conditions known to affect oyster survival, growth, and reproduction. Daily data from 2015 to 2019 were interpolated to generate salinity and temperature profiles across Louisiana's estuaries, which were then used to classify zones based on monthly and annual salinity mean and variance. Zones were classified as supportive of (1) broodstock sanctuary reefs (i.e., support reproductive populations), (2) productive reefs during dry (salty) years, (3) productive reefs during wet (fresh) years, and (4) off-bottom aquaculture development. Of the 38,000 km² investigated, over 11,000 km² of potential oyster zone area was identified across the Louisiana coast. The Broodstock Sanctuary Zone was the smallest (∼540 km²), as salinity variance limited this zone in many areas, as it is driven largely by riverine inputs across many estuaries. Located up-estuary (Dry Restoration Zone) and down-estuary (Wet Restoration Zone) of the Broodstock Sanctuary Zone, Dry and Wet Restoration Zone areas covered ∼2400 km² and ∼3900 km², respectively. Mapped reefs in Louisiana currently exist largely within the Dry Restoration zones, suggesting a potential strategy to focus reef development in Wet Restoration zones to ensure reef network sustainability through years with high precipitation and river inflow. The off-bottom Aquaculture Zone was the largest (∼6400 km²) zone identified, with much of this area located more down-estuary and off-shore. Accounting for variable water quality conditions enables the development of a network of reefs resilient to environmental variability, and more stable areas for consistent off-bottom aquaculture production. Spatial planning and identification of oyster resource zones reduces focus on individual reef success and supports management of oyster metapopulation outcomes, while identifying zones supportive of off-bottom aquaculture.
... Bivalves including oysters can cope with low salinity for a short period by releasing a suite of stress response genes, closing their valves, and shifting to anaerobic metabolism to isolate their internal tissues in order to maximize their survival under extreme conditions (Michaelidis et al., 2005;Zhang and Zhang, 2012). Short-term exposure to low salinity sometimes may even enhance the population by reducing predators (e.g., oyster drill, whelk) and parasites (e.g., Perkinsus marinus) (La Peyre et al., 2003;Levinton et al., 2011;Pollack et al., 2011). However, if they need to close their valves for too long, the accumulation of toxic compounds and the unfavorable energetic balance inside the shell will eventually lead to their mortality. ...
Article
Extreme precipitation events are projected to occur more frequently under a warming climate, posing increasing threats to coastal ecosystems. Hurricane Harvey (2017), the wettest tropical cyclone in the U.S. history that caused a 1000-year flood in the Houston metropolitan area, provides an opportunity to study the response of coastal ecosystems to extreme events. As sessile, epibenthic filter-feeding organisms, oysters are inherently sensitive to changes in environmental and water quality conditions, making them a good indicator for ecosystem health. Oyster measurements at 130 sites in Galveston Bay show that the mean oyster mortality drastically increased from 11% before Harvey to 48% after Harvey. Post-Harvey oyster mortality exhibited large spatial variability and was up to 100% at some major reef complexes. For all the oyster sampling sites, brown shells were dominant, while black shells indicating mud burial were rare. Considering the little impact from sediment deposit, we hypothesized the low-salinity exposure as the main cause for the massive oyster kill. We conducted a multidisciplinary (biological-geological-physical) investigation combining the oyster data with bay-wide sediment core data and results of a previously-validated high-resolution numerical model. Oyster mortality was found to be significantly and positively correlated with the bottom low-salinity exposure time (duration of bottom salinity continuously less than 5 PSU), while there was no significant relationship with the thickness of storm-induced sediment deposit. The physiological aspects for the impact of low-salinity exposure, the underlying physical mechanisms for the prolonged salinity recovery, and wider implications of oyster kill in Galveston Bay in the context of global oyster reef conditions were discussed. The worldwide reported oyster kill events due to extreme weather events suggest additional pressure posed by future climate on the native coastal oyster reefs that are already at the brink of functional extinction worldwide due to centuries of resource extraction and coastal habitat degradation.
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Low salinity can negatively affect reproduction in estuarine bivalves. The spatial and temporal extent of these effects are important to inform models of population dynamics, environmental risk assessments, restoration efforts, and predictions of climate change effects. We hypothesized that oysters at low salinity sites would have delayed gametogenesis compared to their higher salinity counterparts in downstream experimental cages. The timing of gametogenesis and spawning was observed June-August for 2-year-old oysters from three distinct ancestries (Native, Hatchery, Aquaculture), outplanted at age 1 month along the salinity gradient (3-30 psu) of a temperate estuary. A second season of data was collected from 3-year-old Aquaculture oysters (comparable to year 1 data) and Native adult oysters transplanted one year prior. Dermo was very low both years. A delay in gametogenesis and spawning was observed for all ancestries at low salinity relative to higher salinity sites during July and August of the first year but not the second year. In contrast, June showed the reverse pattern with northern low salinity sites having more advanced gonad index (2.65) than a high salinity site (1.46). This difference in average gonad index was 2.65 vs 1.46, respectively, for the Native line and 2.62 vs 2.08 for Aquaculture. Low salinity seemed to not only induce earlier gametogenesis in June, but also extended the reproductive season relative to higher salinity sites. Among oyster ancestries, the Aquaculture line stood out as having 30 — 48% lower gametogenic synchrony within sites, but only in 2018. Despite some dependence of reproductive phenology on salinity variation, the Native low salinity population demonstrates notable reproductive plasticity in the completion of a reproductive cycle across a wide range of salinities, an encouraging result for potential future restoration strategies.
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The eastern oyster Crassostrea virginica has a wide salinity tolerance, but all life stages are vulnerable to environmental extremes and elevated temperatures can truncate the expected salinity tolerance. The rising water temperatures and more intense and variable storm events predicted to accompany global climate change therefore raise concerns for habitat suitability for ecologically important species such as the eastern oyster. To better understand environmental limitations, oysters of all life stages were exposed to a range of salinities (0–40) and temperatures (25 and 30°C) to test physiological tolerance to the combined effects of osmotic and thermal stresses. Elevated temperatures (30°C) amplified negative effects during exposure to salinity extremes at all life stages; however, tolerance to extremes increased with developmental stage (gametes < embryos < larvae < spat < adult). Gradual changes allow for a wider tolerance in juvenile oysters (spat) compared to acute changes, and short-term reprieves during low salinity exposure improved survival rates for adults. Overall, the present study found a threshold salinity of 15 for polyhaline oyster populations and highlights the importance of both rate of change and temperature as critical components of salinity tolerance. Additionally, salinities < 10 during the summer months could result in negative population effects, especially if extreme low salinity occurs during peak reproduction when pelagic gametes, embryos, and larvae are most vulnerable to environmental stresses. This work will benefit population models and inform resource management decisions regarding the timing of controlled freshwater releases.
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Bivalves leave behind articulated valves when they die that can be used to estimate natural mortality. However, a common method used to estimate natural mortality in bivalves, known as the box count method, includes several assumptions that may be violated for eastern oysters Crassostrea virginica in Chesapeake Bay, Maryland. We developed a Bayesian model alternative to the box count method that included modifications to account for potential violations of assumptions. The model was applied to oysters in 32 areas in Maryland during 1991–2017 using dredge survey data, and the natural mortality estimates from the model were compared to ones derived from the box count method. The spatial and temporal trends in natural mortality from the model were summarized using dynamic factor analysis. Natural mortality showed considerable spatial and temporal variation, with median mortality rates ranging between 0.00 and 0.96 yr⁻¹. Natural mortality spiked in most regions in 2002 and was lower in more recent years. The Bayesian model estimated slightly higher (0.02 yr⁻¹ on average) natural mortality than the box count method, except for years following high natural mortality, after which the Bayesian model estimated lower natural mortality than the box count method. The dynamic factor analysis revealed two common trends in natural mortality and a north-south gradient in the loadings on the trends. This work improves our understanding of the variability of oyster natural mortality in Maryland and the Bayesian model could be modified for use with oysters in other regions or with other species.
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Diagnosis of Perkinsus marinus disease of eastern oysters Crassostrea virginica has been routinely accomplished by incubating oyster tissues in a fluid thioglycollate medium described by Ray in the early 1950s. At least three modifications of the technique are available with applications to different diagnostic needs. Of these, the quantitative whole-oyster technique is potentially the most valuable because it includes all oyster tissues and does not rely on subjective estimates of intensity. A variety of protocols and approaches were examined in an attempt to develop a standardized procedure for quantitative whole-oyster diagnosis that optimizes sensitivity, specificity, precision and accuracy. A recommended procedure, with possible variations, is presented here with the expectation that its presentation will foster further refinement and improvement. We conclude that the recommended whole-oyster diagnostic technique is capable of providing reliable quantifications of prevalence and intensity and has great potential for examining correlations of total P. marinus body burdens with measurements of oyster biology and for evaluating or calibrating other diagnostic techniques.
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Oysters held near-shore in Caminada Bay, Louisiana during the summer, exhibit hypertrophic gonads with prominent genital canals beneath transparent mantle tissue about four weeks post-hatchery spawning, indicating recycling. Broodstock (N = 200) were analyzed histologically over a two-year period to document such gametogenesis, using Gonad/Body Ratios (GBR) and developmental stages. Ten oysters were randomly selected from a broodstock pool prior to each spawning attempt, and monthly during the winter-spring. As expected, the mean GBR before successful spawning attempts was significantly greater (P ≤ 0.05) than the mean GBR before unsuccessful attempts. A dramatic drop in the percent occurrence of the advanced spawning and regression stage from May to June, a >40% spawning stage occurrence from May to October, and fluctuations in the percent occurrence of early and late developmental stages during the summer months illustrates gonadal recycling.