Environmental Characteristics Associated with the Occurrence of Avian Botulism in Wetlands of a Northern California Refuge
Results
Of the 10 wetlands originally included in the study, only the 4 wetlands flooded year-round were included in our analyses. Outbreaks in seasonally flooded wetlands were inconsistent, and the environmental data collected in these wetlands were insufficient to warrant statistical evaluation.
Botulism outbreaks occurred in sentinel mallards in 2 wetland enclosures (P-2, P-8) in both 1987 and 1989 (Fig. 1); no outbreaks were detected in 1988. During these botulism outbreaks, weekly mortality rates in sentinel mallards ranged from 1.0 to 8.8 deaths/100 birds. Although botulinum toxin was found in 2 carcasses in another enclosure (T-F) in 1988, these cases were widely separated in time (>2 months), and botulism was not detected in wild birds in this or any other wetland in 1988. Because our criteria for classifying outbreak wetlands for this study required the detection of >1 carcass with botulinum toxin, the cases in T-F in 1988 were not considered to constitute an outbreak.
The range and variability of wetland conditions in our 4 study sites (means, minimums, maximums) fell within expected ranges (Table 1). Water temperatures varied from 13.0 to 26.8°C, with sediment temperatures generally 1–2 degrees higher. The water was very fresh, with specific conductivity values in the range of 88–896 µmhos/cm, and water pH was neutral to moderately alkaline, within a range of 6.0–8.4. Invertebrates included in our statistical analysis (counts and benthic biomass) included gastropods, nematodes, oligochaetes, eubranchipods, lymnaeid snails, cyclopoid copepods, ostracods, unknown dipterans, ceratopogonids, chaoborids, culicids, tipulids, ephydrids, notonectids, muscids, stratiomyids, unknown chironomids, and chironomids of the genera Goeldichironomus, Glyptotendipes, Tanypus, and Procladius. Other invertebrates had very low biomass or counts and were only included in total biomass calculations.
Table 1. Environmental
variables measured and averaged (no. of sampling intervals [n], mean
[
], minimum [min] and maximum
[max]) in 1.6-ha wetland enclosures where botulism outbreaks occured (outbreak
wetlands) and did not occur (nonoutbreak wetlands) in sentinel mallards at
the Sacramento National Wildlife Refuge, California, 1987-89. Outbreak wetlands
were further separated into outbreak intervals (i.e., when botulism outbreaks
occured in sentinel mallards) and nonoutbreak intervals (i.e., when no outbreaks
occured). For each sampling interval, environmental measurement averaged from
5 locations distributed throughout the wetland enclosure included redox potential
(redox) and standardized redox potential (stand. redox) measured in millivolts,
temperature measured in °C, specific conductivity measured in µmhos/cm,
pH, numbers of benthic invertebrates (benthic counts; log transformed), mass
(g) of benthic invertebrates (benthic biomass; log transformed), mass (g)
of total invertebrates (total biomass; log transformed), dissolved oxygen
in the water measured in mg/L, percent organic matter in the sediments, water
depth measured in centimeters, and water turbidity measured in nepholometer
turbidity units.
| Outbreak wetlands (P-2, P-8) | ||||||||||||
| Outbreak intervals | Nonoutbreak intervals | Nonoutbreak wetlands (T-F, T-19) | ||||||||||
| Variable | n |  |
min | max | n | |
min | max | n | |
min | max |
| Redox Wa | 15 | 208 | 50 | 338 | 40 | 196 | -26 | 467 | 56 | 219 | 8 | 477 |
| Redox S1b | 15 | 111 | -24 | 303 | 40 | 19 | -171 | 254 | 56 | 80 | -80 | 337 |
| Redox S2c | 15 | 62 | -40 | 256 | 39 | 47 | -165 | 289 | 56 | 49 | -174 | 375 |
| Stand. redox W | 15 | 251 | 86 | 367 | 40 | 227 | 16 | 505 | 56 | 248 | 17 | 485 |
| Stand. redox S1 | 15 | 109 | -43 | 252 | 40 | 28 | -177 | 274 | 56 | 86 | -65 | 317 |
| Stand. redox S2 | 15 | 61 | -43 | 252 | 39 | 47 | -165 | 289 | 56 | 63 | -160 | 359 |
| Temperature W | 15 | 22.4 | 18.3 | 26.5 | 41 | 20.6 | 13.0 | 26.6 | 57 | 20.4 | 12.1 | 26.8 |
| Temperature S1 | 15 | 23.9 | 19.4 | 28.3 | 40 | 22.1 | 13.5 | 28.0 | 56 | 21.7 | 13.7 | 27.6 |
| Temperature S2 | 15 | 23.8 | 19.5 | 27.6 | 39 | 22.1 | 13.9 | 28.0 | 56 | 21.8 | 14.4 | 27.3 |
| Conductivity W | 15 | 474 | 285 | 882 | 41 | 428 | 231 | 896 | 57 | 336 | 88 | 864 |
| Conductivity S1 | 15 | 793 | 584 | 1086 | 40 | 733 | 530 | 1018 | 56 | 670 | 222 | 1186 |
| Conductivity S2 | 15 | 1028 | 745 | 1275 | 39 | 1020 | 688 | 1510 | 56 | 1140 | 679 | 1819 |
| pH W | 15 | 7.73 | 7.09 | 8.80 | 41 | 7.54 | 7.00 | 8.77 | 57 | 7.51 | 6.38 | 8.75 |
| pH S1 | 13 | 7.26 | 7.00 | 7.47 | 38 | 7.26 | 6.75 | 8.03 | 53 | 7.19 | 6.50 | 8.35 |
| pH S2 | 13 | 7.14 | 6.94 | 7.53 | 37 | 7.30 | 6.73 | 8.02 | 53 | 7.28 | 5.99 | 8.87 |
| Benthic counts | 15 | 1.68 | 1.24 | 2.15 | 38 | 1.53 | 0.83 | 2.41 | 55 | 1.72 | 0.87 | 2.64 |
| Benthic biomass | 15 | 0.010 | 0.001 | 0.042 | 38 | 0.006 | 0.000 | 0.049 | 55 | 0.005 | 0.00 | 0.02 |
| Total biomass | 15 | 0.012 | 0.003 | 0.045 | 38 | 0.009 | 0.002 | 0.051 | 55 | 0.008 | 0.00 | 0.08 |
| Dissolved oxygen | 15 | 3.94 | 1.51 | 7.05 | 41 | 4.20 | 0.92 | 10.32 | 57 | 4.20 | 1.00 | 10.05 |
| % organic matter | 15 | 12.1 | 4.5 | 22.5 | 39 | 10.8 | 4.7 | 18.3 | 57 | 6.1 | 3.8 | 7.8 |
| Depth | 15 | 52.5 | 35.8 | 100.8 | 41 | 53.6 | 22.8 | 105.7 | 57 | 53.8 | 34.8 | 85.6 |
| Turbidity | 15 | 11.4 | 2.4 | 34.3 | 40 | 23.8 | 2.2 | 150.0 | 57 | 25.6 | 2.1 | 78.2 |
| aW refers to water samples collected 7.5 cm above the sediment-water interface. | ||||||||||||
| bS1 refers to interstitial water samples collected 2.5 cm below the sediment-water interface. | ||||||||||||
| cS2 refers to interstitial wtaer samples collected 11.25 cm below the sediment-water interface. | ||||||||||||
Seven principal components identified by factor analysis accounted for 79% of the variation in the original 22 environmental parameters. Rotated factor loadings showed that environmental parameters in the soil and water were highly correlated (Table 2). The resulting 7 environmental factors had a relatively simple interpretation in relation to the original environmental parameters. Redox potential variables (REDOX) explained the largest portion of the variation (23%) found in the environmental measurements. Temperature variables (TEMP) and a factor consisting of specific conductivity and water pH (SC–pHW) accounted for an additional 26% of the variation. The fourth factor (pHSOIL) represented soil pH, and the fifth factor (INVERT) represented variables related to invertebrates. The sixth factor (DO–DEPTH–POM) was positively associated with dissolved oxygen and water depth but negatively associated with percent organic matter. Water turbidity was the only environmental variable strongly represented by the seventh factor (TURB).
| Table 2. Seven principal factor patterns after varimax rotation of environmental variables measured in wetland enclosures at the Sacremento National Wildlife Refuge, California, 1987-89. Standardized regression coefficients indicate sign and relative strength of environmental variables influencing each factor. Only coefficients are included. | ||
| Factor | Environmental variable | Coefficient |
| REDOX | Redox potential Wa | 0.858 |
| Redox potential S1b | 0.925 | |
| Redox potential S2c | 0.937 | |
| Standardized redox potential W | 0.863 | |
| Standardized redox potential S1 | 0.924 | |
| Standardized redox potential S2 | 0.923 | |
| TEMP | Temperature W | 0.962 |
| Temperature S1 | 0.951 | |
| Temperature S2 | 0.948 | |
| SC–pHW | Specific conductivity W | 0.743 |
| Specific conductivity S1 | 0.850 | |
| Specific conductivity S2 | 0.760 | |
| pHSOIL | pH W | 0.558 |
| pH S1 | 0.926 | |
| pH S2 | 0.947 | |
| INVERT | Benthic invertebrates | 0.466 |
| Benthic ivertebrate biomass | 0.910 | |
| Total invertebrate biomass | 0.905 | |
| DO–DEPTH–POM | Dissolved Oxygen | 0.575 |
| Depth | 0.556 | |
| Percent organic matter | -0.647 | |
| TURB | Turbidity | 0.906 |
| aW refers to water samples collected 7.5 cm above the sediment-water interface. | ||
| bS1 refers to interstitial water samples collected 2.5 cm below the sediment-water interface. | ||
| cS2 refers to interstitial water saples collected 11.25 cm below the sediment-water interface. | ||
Comparisons Between Outbreak and Nonoutbreak Wetlands
Repeated measures ANOVA indicated outbreak wetlands had lower REDOX factor scores (F1,2 = 18.3, P = 0.05) than nonoutbreak wetlands during our study (Fig. 2). None of the remaining environmental factors showed a consistent difference (Ps > 0.20) between the outbreak and nonoutbreak wetlands. Significant changes in environmental factors among sampling intervals were found for REDOX (F19,38 = 12.6, P < 0.01; Fig. 2), TEMP (F19,38 = 24.8, P < 0.01; Fig. 3), pHSOIL (F19,38 = 2.88; P < 0.01; Fig. 4), and SC–pHW (F19,38 = 3.42, P < 0.01; Fig. 5). No sampling interval differences (Ps > 0.20) were found for INVERT, DO–DEPTH–POM, or TURB factors. The only significant interaction between botulism outbreaks and sampling intervals was found for SC–pHW (F19,38 = 2.14; P = 0.02).
Comparisons Within Outbreak Wetlands
In outbreak wetlands (P-2, P-8), logistic regression analyses showed an association between the probability of botulism in sentinel mallards and TEMP (X 21 = 8.95, P < 0.01), INVERT (X 21 = 5.64, P = 0.02), and TURB (X 21 = 12.78, P < 0.01) factors. No differences (Ps > 0.10) were found for other environmental factors or serial correlation among sequential sampling intervals, although REDOX ( X 21 = 2.47, P = 0.12) and pHSOIL (X 21 = 2.61, P = 0.11) approached our significance criteria. Coefficients from the logistic regression model indicated increasing values of TEMP and INVERT were correlated with a higher probability of botulism outbreaks. However, water turbidity appeared to have the opposite relation: decreasing levels of turbidity were correlated with a higher probability of botulism outbreaks. When REDOX and pHSOIL were included in the logistic regression model, they showed opposite effects. Botulism outbreaks were positively related to REDOX and negatively related to pHSOIL. Preliminary examination of daily mortality rates during outbreaks indicated
a larger range of weekly mortality rates in P-8 than in P-2 (Fig.
1). Therefore, correlations between mortality rates and the 7 principal
factors were conducted separately for each enclosure. For P-2, daily mortality
rates during botulism outbreaks (excluding Jul 1987 due to missing factor
data) were negatively correlated with SC–pHW (
= -0.87, P = 0.02, n = 6) and INVERT (
= -0.90, P = 0.01, n = 6) factors. In contrast, daily botulism
mortality rates for P-8 (excluding Jul 1987) were positively correlated with
TURB ( = 0.83, P =
0.02, n = 7).
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