This study evaluated the presence of
The main objectives of this study were to evaluate the effects of seasonal variations on the diversity of
A total of 136 water samples were collected from 34 sampling locations along Puzih River (23°28′N, 120°13′E) in Southern Taiwan. The Puzih River lies within a subtropical monsoon region located near the Tropic of Cancer. A map of the study area with sampling locations is presented in Figure
Sampling locations for
Physical water quality parameters were measured for each water sample on location at the time of sample collection. Water temperature and pH level were measured
For each collected water sample, 2 L was concentrated via membrane filtration method using GN-6 Metricel mixed cellulose ester (MCE) membranes (0.45 µm pore diameter, Pall, Port Washington, NY, USA). After filtration, the membranes were scraped, and the collected material was washed with 100 mL eluted fluid consisting of phosphate-buffered saline (PBS; 7.5 mM Na2HPO4, 3.3 mM NaH2PO4, and 108 mM NaCl, pH 7.2), 1% Tween 80, and 1% sodium dodecyl sulfate. The resulting solution was then transferred into two 50 mL conical centrifuge tubes and centrifuged at 2,600 ×g for 30 min. Subsequently, the top 45 mL supernatant fluid was removed, and the remaining 5 mL pellet was preserved with PBS at 4°C for further culture procedure and PCR analyses.
The resulting pellet was transferred to a culture plate containing 1.5% nonnutrient agar (NNA) and a lawn of heat killed
DNA extraction was done with the concentrated pellet using MagNA Pure LC instruments (Roche, USA) with MagNA Pure LC DNA isolation kits III (Roche, USA) following automated mode specified by the kit manufacturer’s instructions manual. The resulting solution was analyzed for the presence of
The PCR assay genus-specific primers’ sets JDP1 and JDP2 used in this study were designed for
PCR reaction solution was prepared with 5
Negative DNA controls (template DNA replaced with distilled water) and positive controls (
A total of 136 water samples were collected in this study. The detection results of
Distribution of
Sampling season | Sample size | Number of detection for |
Percent of detection for |
Positive sample no. | Species name | Genotypes |
---|---|---|---|---|---|---|
Spring | 34 | 1 | 2.9% | A19 |
|
T4 |
Summer | 34 | 11 | 32.4% | B7 |
|
T4 |
B9 |
|
T4 | ||||
B14 |
|
T4 | ||||
B21 |
|
T11 | ||||
B24 |
|
T4 | ||||
B25 |
|
T4 | ||||
B26 |
|
T4 | ||||
B30 |
|
T4 | ||||
B32 |
|
T4 | ||||
B33 |
|
T4 | ||||
B34 |
|
T2 | ||||
Autumn | 34 | 1 | 2.9% | C23 |
|
T5 |
Winter | 34 | 3 | 8.8% | D9 |
|
T4 |
D16 |
|
T15 | ||||
D18 |
|
T12 | ||||
| ||||||
Total | 136 | 16 | 11.7% |
The sixteen
For the seasonal distribution results, sample A19 was detected
Quarterly monitoring revealed that the diversity of
Water quality parameters were compared with respect to
Ranges, mean, and standard deviation of water quality parameters during different seasons.
Water quality parameter | Heterotrophic plate counts (CFU/mL) | Total coliforms (CFU/100 mL) | Turbidity (NTU) | Temperature (°C) | pH value | |||||
---|---|---|---|---|---|---|---|---|---|---|
Range | Mean |
Range | Mean |
Range | Mean |
Range | Mean |
Range | Mean |
|
Spring |
|
|
0– |
|
25–151 |
|
19.8–22.9 |
|
7.64–8.23 |
|
Summer |
|
|
12– |
|
34–206 |
|
31.0–33.2 |
|
7.67–8.09 |
|
Autumn |
|
|
|
|
25–471 |
|
25–25.3 |
|
7.71–8.40 |
|
Winter | 18.5– |
|
22– |
|
40–155 |
|
18.4–22.9 |
|
8.11–8.76 |
|
| ||||||||||
Total | 18.5– |
|
0– |
|
25–471 |
|
18.4–33.2 |
|
7.64–8.76 |
|
Nonparametric test results for difference and correlation for
Water quality parameter | Spring | Summer | Autumn | Winter | Total | |||||
---|---|---|---|---|---|---|---|---|---|---|
Mann-Whitney |
Spearman rank test | Mann-Whitney |
Spearman rank test | Mann-Whitney |
Spearman rank test | Mann-Whitney |
Spearman rank test | Mann-Whitney |
Spearman rank test | |
Heterotrophic plate counts (CFU/mL) |
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Total coliforms (CFU/100 mL) |
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Turbidity (NTU) |
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Temperature (°C) |
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pH value |
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Note:
The total detection rate was 11.7% for
The contribution of the first authors Ming-Yuan Chou, Chi-Wei Tao, and Wen-Chien Huang is considered equal.
This work was supported by a research Grant from the National Science Council of Taiwan, Taiwan (NSC 100-2116-M-194-004-MY2) and Cheng Hsin General Hospital (102-10), Taiwan.