Supplemental Materials

Strong impact of organic emerging contaminants on prokaryotic communities at the

horizontal but not vertical scales in a subtropical reservoir, China

Running title: Response of reservoir prokaryotic communities to micropollutants

Bob Adyari1,2,3, Dandan Shen4, Shuang Li1,2, Lanping Zhang1,2, Azhar

Rashid1,5, Qian Sun1, Anyi Hu1*, Nengwang Chen6, Chang-Ping Yu1,7

1. CAS Key Laboratory of Urban Pollutant Conversion, Institute of Urban Environment,

Chinese Academy of Sciences, Xiamen 361021, China

2. University of Chinese Academy of Sciences, Beijing 100049, China

3. Department of Environmental Engineering, Universitas Pertamina, Jakarta 12220,

Indonesia

4. Section of Biological Oceanography, Leibniz Institute for Baltic Sea Research,

Warnemünde, Germany

5. Nuclear Institute for Food and Agriculture, Tarnab, Peshawar, Pakistan

6. Fujian Provincial Key Laboratory for Coastal Ecology and Environmental Studies,

College of the Environment and Ecology, Xiamen University, Xiamen 361005, China

7. Graduate Institute of Environmental Engineering, National Taiwan University, Taipei

106, Taiwan

*Correspondence to:

Dr. Anyi Hu, Email: ayhu@iue.ac.cn; Tel: 86-592-6190582.

Taxonomic composition of core and satellite groups within PA or FL communities.

Overall, Cyanobacteria dominated the core sub-communities from both horizontal (surface

water; average 33.3% of PA and 39.2% of FL core sub-communities) and vertical datasets

(PA: 24.7% and FL: 26.7%) (Fig. S3). There was a noticeable decrease in the Cyanobacteria

from surface to vertical core groups. In contrast, satellite sub-communities harbored higher

relative abundance of Proteobacteria, which consists of Alphaproteobacteria (surface PA:

18.7% and surface FL: 18.04%; vertical PA: 16.23% and vertical FL: 17.47%),

Gammaproteobacteria (surface PA: 18.65% and surface FL: 16.44%; vertical PA: 16.77%

and vertical FL: 13.03%), and Deltaproteobacteria (surface PA: 11.58% and surface FL:

12.26%; vertical PA: 8.23% and vertical FL: 12.7%) than those in core sub-communities

(Wilcoxon test, P < 0.05). Despite of their presence in core and satellite sub-communities,

Chloroflexi, Acidobacteria and Firmicutes were generally less abundant in the core sub-

communities (0.36% ~ 1.17%) than their satellite (1.2% ~ 5.31%) counterparts (Wilcoxon

test, P < 0.05). The comparison of core communities between PA and FL fractions indicated

that the members affiliated with Cyanobacteria and Actinobacteria tended to be enriched in

the FL, while, Plancomycetes and Bacteroides were of higher abundance in the PA.

Table S1. Friedman test results of seasonal and depth differences of physico-chemical

variables.

Dataset Parameter Full nameSeason DepthP P

Vertical

Chl-a Chlorophyll-a 0.0314 0.001Cond. Conductivity < 0.001 0.0983DO Dissolved oxygen 0.0037 0.0023DON Dissolved organic nitrogen 0.0034 0.3916DOP Dissolved organic phosphorus < 0.001 0.4484DSi Dissolved silica 0.0018 0.5433DTN Dissolved total nitrogen < 0.001 0.4188DTP Dissolved total phosphorus < 0.001 0.1856NH4-N Ammonium nitrogen 0.001 0.2085NO2-N Nitrite nitrogen 0.0079 0.4188NO3-N Nitrate nitrogen < 0.001 0.3659SRP Soluble reactive phosphorus < 0.001 0.037Temp. Temperature < 0.001 < 0.001TPP Total particulate phosphorus 0.0273 0.0342TSM Total suspended matter 0.0013 0.0386

Horizontal

Chl-a Chlorophyll-a < 0.001 NACond. Conductivity < 0.001 NADO Dissolved oxygen 0.002 NADON Dissolved organic nitrogen 0.224 NADOP Dissolved organic phosphorus < 0.001 NADSi Dissolved silica 0.002 NADTN Dissolved total nitrogen < 0.001 NADTP Dissolved total phosphorus < 0.001 NANH4-N Ammonium nitrogen < 0.001 NANO2-N Nitrite nitrogen 0.0027 NANO3-N Nitrate nitrogen < 0.001 NASRP Soluble reactive phosphorus < 0.001 NATemp. Temperature < 0.001 NATPP Total particulate phosphorus < 0.001 NATSM Total suspended matter < 0.001 NA

NA = non-applicable

Table S2. Friedman test results of seasonal and depth differences of micropollutant

variables.

Dataset Parameter micropollutants group Full nameSeason DepthP P

Vertical

BPA Plasticizer Bisphenol_A < 0.001 0.4717BPAF Plasticizer Bisphenol_AF < 0.001 0.02BPS Plasticizer Bisphenol_S < 0.001 0.8013BZP UV filter Benzophenon-3 < 0.001 0.7736CAF Stimulant Caffeine 0.0045 0.185CBZ Anticonvulsant Carbamazepine < 0.001 0.217DCF Anti-inflammatory Diclofenac 0.0917 0.1116ERF Antibiotics Enrofloxacin  0.9765 0.091FNP Anti-inflammatory Fenoprofen 0.2043 0.4336LST Hypertensive agent Losartan 0.0072 0.771MCZ Anti-fungal Miconazole 0.0034 0.6444MTP Preservative Methyl_paraben 0.0038 0.9221PRC Anti-inflammatory Paracetamol 0.036 0.8214SFM Antibiotics Sulfamonomethoxine < 0.001 0.0719SFP Antibiotics Sulfapyridine 0.0074 0.3373SFZ Antibiotics Sulfamethoxazole 0.0019 0.224TMP Antibiotics Trimethoprim < 0.001 0.0772

Horizontal

BPA Plasticizer Bisphenol A 0.002 NABPAF Plasticizer Bisphenol AF < 0.001 NABPS Plasticizer Bisphenol S < 0.001 NABZP UV filter Benzophenon-3 < 0.001 NACAF Stimulant Caffeine 0.0035 NACBZ Anticonvulsant Carbamazepine < 0.001 NADCF Anti-inflammatory Diclofenac 0.0113 NAERF Antibiotics Enrofloxacin  0.3507 NAFNP Anti-inflammatory Fenoprofen 0.456 NALST Hypertensive agent Losartan 0.0102 NAMCZ Anti-fungal Miconazole 0.0049 NAMTP Preservative Methyl_paraben 0.0071 NAPRC Anti-inflammatory Paracetamol 0.0393 NASFM Antibiotics Sulfamonomethoxine < 0.001 NASFP Antibiotics Sulfapyridine 0.0049 NASFZ Antibiotics Sulfamethoxazole 0.003 NATMP Antibiotics Trimethoprim 0.0012 NA

NA = non-applicable.

Table S3. The relative abundance of core and satellite groups in the total prokaryotic

communities.

Scale FractionRelative to total abundance Relative to OTU richness

Core Satellite Core Satellite

Horizontal PA 98.54% 1.07% 35.16% 30.20%FL 98.90% 0.68% 39.13% 24.24%

Vertical PA 98.39% 1.10% 38.66% 30.79%FL 98.59% 0.68% 40.31% 23.73%

Table S4. DistLM results of significant micropollutant variables.

Category Abbreviation Full nameVertical profile Surface profile

Free-living Particle-associated Free-living Particle-associatedCore Satellite Core Satellite Core Satellite Core Satellite

Plasticizer BPA Bisphenol A 6.41%*** 5.89%*** 13.21%*** 6.47%*** NS NS 2.79%* 4.32%**Plasticizer BPAF Bisphenol AF 17.29%*** 7.2%*** 15.5%*** 7.13%*** 32.57%*** 12.9%*** 31.5%*** 12.68%***Plasticizer BPS Bisphenol S 9.01%*** NS NS 3.79%** NS NS 21.41%*** 9.79%***UV filter BZP Benzophenon NS NS NS NS NS 3.64%* 2.25%* 3.76%*Stimulant CAF Caffeine 4.01%** 4.11%** 4.92%** NS 8.73%*** 4.09%** 3.02%* 6.76%***Anticonvulsant CBZ Carbamazepine 4.56%** 4.63%*** 8.22%*** 4.75%*** 2.91%* 4.15%** NS NSAnti-fungal MCZ Miconazole NS NS NS NS 2.58%* NS NS NSAntibiotics SFM Sulfamonomethoxine NS NS NS NS 20.61%*** 8.36%*** 3.11%** 4.2%**Antibiotics SFP Sulfapyridine 8.07%*** 3.55%* 4.98%** 3.72%** NS NS NS NSAntibiotics SFZ Sulfamethoxazole NS NS 5.01%* NS NS 8.31%*** 10.04%*** NSAntibiotics TMP Trimethoprim NS NS 3.01%* NS NS NS NS 3.93%*

Total: 49.35% 25.38% 54.85% 25.86% 67.40% 37.81% 74.12% 45.44%Significance codes: ***P < 0.001, ** P < 0.01, * P < 0.05, NS P > 0.05.

Table S5. DistLM results of significant physico-chemical variables.

Abbreviation Full nameVertical profile Surface profile

Free-living Particle-associated Free-living Particle-associatedCore Satellite Core Satellite Core Satellite Core Satellite

Chl-a Chlorophyll-a 10.18%*** NS NS NS NS NS NS NSCond. Conductivity 5.79%*** 6.98%*** 15.26%*** 6.92%*** NS NS NS NSDepth Depth 3.21%** 3.71%*** NS 3.77%*** NS NS NS NSDO Dissolved oxygen 4.82%*** 5.48%*** 9.87%*** 5.53%*** NS 5.74%*** NS 7.28%***DON Dissolved organic nitrogen NS NS NS NS NS NS NS 3.74%*DOP Dissolved organic phosphorus 13.89%*** NS NS 4.21%*** NS NS NS NSDSi Dissolved silica 3.27%** NS NS NS NS NS NS NSDTN Dissolved total nitrogen NS NS 6.43%*** NS 3.9%** NS 5.01%*** NSDTP Dissolved total phosphorus NS 7.33%*** 3.18%* NS NS NS NS NSNH4-N Ammonium nitrogen 3.18%** NS NS NS 38.08%*** 13.89%*** 35.59%*** 10.23%***NO2-N Nitrite nitrogen NS NS NS NS NS NS NS NSNO3-N Nitrate nitrogen NS NS NS NS NS 10.16%*** NS NSSRP Soluble reactive phosphorus NS NS NS 3.09%* NS NS NS NSTemp Temperature 18.84%*** 3.99%*** 18.82%*** 7.76%*** 19.95%*** NS 23.66%*** 13.29%***TPP Total particulate phosphorus NS NS 2.73%* NS NS NS NS NSTSM Total suspended matter 2.45%* NS 4.41%** NS 2.8%* 4.45%*** 2.58%* NS  Total: 65.63% 27.49% 60.70% 31.28% 64.73% 34.24% 66.84% 34.54%

Significance codes: ***P < 0.001, ** P < 0.01, * P < 0.05, NS P > 0.05.

Table S6. Node properties of DIABLO selected variables for core PA sub-communities (in excel)

Table S7. Edge properties of DIABLO selected variables for core PA sub-communities (in excel)

Table S8. Node properties of DIABLO selected variables for core FL sub-communities (in excel)

Table S9. Edge properties of DIABLO selected variables for core FL sub-communities (in excel)

Table S10. Samples description (in excel)

Fig. S1. Principal coordinate analysis (PCoA) of vertical prokaryoplankton assemblages

grouped by depth (A) and season (B).

Fig. S2. Venn diagram showing the distribution of shared OTUs between PA or FL

communities within core or satellite groups. Asterisk indicates significant difference between

PA and FL based on PERMANOVA test. Significance codes: ** P < 0.01, * P < 0.05.

Fig. S3. Taxonomic composition of four sub-communities at the horizontal scale (i.e.,

horizontal core PA, horizontal satellite PA, horizontal core FL, and horizontal satellite FL),

and four sub-communities at the vertical scale (i.e., vertical core PA, vertical satellite PA,

vertical core FL, and vertical satellite FL). The Phylum Proteobacteria is shown at the Class

level.

Fig. S4. Rarefaction curve of original samples reads