Water Quality Monitoring of the Bellinger and Kalang Rivers 2015-16

Water Quality Monitoring of the Bellinger and Kalang Rivers Technical Report for Bellingen Shire Council

Sarah Mika, Adrienne Burns and Darren Ryder

Technical Report for Bellingen Shire Council

Dr Sarah Mika, Dr Adrienne Burns, A/Prof. Darren Ryder. School of Environmental and Rural Science, University of New England, Armidale, NSW, 2351

This report should be cited as: Mika, S., Burns, A. and Ryder, D.S (2016). Water Quality Monitoring of the Bellinger and Kalang Rivers 2015-16. Technical Report for Bellingen Shire Council. University of New England, Armidale.

Project contact: Dr Sarah Mika School of Environmental and Rural Science University of New England, Armidale, NSW, 2351 Email: [email protected]

Cover Photo: Confluence of Bellinger and Kalang Rivers at Urunga (D.Ryder, 2015).

Summary

Water quality including on-site physico-chemical variables and nutrient and algal concentrations were sampled during low flows at eight sites on the Bellinger and Kalang Rivers and their tributaries on 24 November 2015 (T1), 15 February 2016 (T2), 12 April 2016 (T3), and 12 July 2016 (T4), and on 23 June 2016 immediately after a flood event. Microbial Source Tracking (MST) to identify the risk of E.coli contamination was recorded in the lower Bellinger and Kalang estuaries in the post flood period.

Total Nitrogen and Total Phosphorus concentrations exceeded the ANZECC/MER trigger values in all estuary and freshwater sites at some point in the study, and were particularly high in the T3 and T4 low flow periods. Bioavailable nutrients are directly able to be assimilated by aquatic flora and fauna. Available nitrogen (NOx) concentrations were consistently above the ANZECC/MER trigger value in estuary sites for all sample times, peaking at 286 µg/L (19 times trigger value) in the Bellinger estuary. NOx concentrations exceeded MER trigger values in all freshwater sites on all occasions except T1 when values were generally lower. All upper and lower estuary sites in the Bellinger and Kalang Rivers exceeded the ANZECC/MER guidelines for available phosphorus at all sampling times. In contrast, Never Never Creek, Rosewood River and Spicketts Creek did not exceed the ANZECC guideline in the study.

The post flood sampling identified Total Nitrogen concentrations were below the MER trigger values at all sites except Spicketts Creek. In contrast, all Total Phosphorus and available nitrogen and phosphorus concentrations were well above MER trigger values and above the mean low flow concentrations for all freshwater and estuary sites.

Three main themes have emerged: (1) Water quality deteriorated in T3 and T4 sampling events after prolonged periods of low flows; (2) the consistently high nutrient concentrations did not result in chlorophyll a concentrations exceeding the trigger values at any site or time; and (3) Spicketts Creek stood out as the system with consistently elevated nutrient concentrations in both post flood and low flow periods, and the only system to record dissolved oxygen concentrations that may stress biota.

Table of Contents

1. Background ...... 5 2. Study Design and Field Sampling ...... 5 3. Results ...... 7 3.1 NSW MER Water Quality Objectives and ANZECC guidelines during low flows ...... 7 3.11 Nutrients ...... 7 3.12 Physico-chemistry ...... 8 4. Post Flood Sampling Event ...... 10 4.1 Water chemistry...... 10 4.2 Microbial Source Tracking ...... 10 Appendix 1 ...... 12 Appendix 2 ...... 14 Appendix 3 ...... 15 Appendix 4 ...... 22

1. Background

Bellingen Shire Council (BSC) resolved to conduct water quality monitoring to inform the Council, community and various government and non-government agencies of the current water quality of Bellingen Shire’s rivers and estuaries. The aim of the monitoring program was to compare results with the ANZECC and NSW MER water quality guidelines for healthy aquatic ecosystems from sites that were: (1) assessed in the Bellinger-Kalang Rivers Ecohealth Project 2009-10, (2) within the zone of the Bellinger River Snapping Turtle mortality event of 2015, (3) downstream of the 2014 Rock Fillets project, (4) downstream of the 2015-16 Friesians and Fish project zone, (5) downstream of the Bellingen Island River Bank Stabilisation projects 2011 and 2013, (6) downstream of the Arthur Keogh Reserve Riparian Rehabilitation project 2012, (7) downstream of the Moody’s Bridge River Bank Stabilisation project 2011, (8) downstream of the Newry Island Sewage Treatment Works Upgrade 2013-14, and (9) capturing the End-of-System inputs of suspended sediments and nutrients from Spicketts Creek identified as a significant point-source in the Bellinger-Kalang Rivers Ecohealth Project 2009-10.

2. Study Design and Field Sampling

Water quality was sampled during low flows at eight sites on the Bellinger and Kalang Rivers and their tributaries (Table 1) on 24 November 2015 (T1), 15 February 2016 (T2), 12 April 2016 (T3), and 12 July 2016 (T4), and on 23 June 2016 immediately after a flood event.

Previous Microbial Source Tracking (MST) analyses from the Bellinger catchment identified a shift from Low to High Risk of Animal Bacteroides during flow events. To capture the MST response, samples were collected from end-of-system sites BELL1 and KALR1 on 23 June 2016 immediately after a flood event. MST analyses were conducted at Australian Water Quality Centre NATA accredited laboratories.

Table 1. Water quality sampling sites. Site codes in brackets indicate those used in the Bellinger-Kalang Rivers Ecohealth Project 2009-10. Site Code River System Location Altitude Easting Northing (m) BELL1 Bellinger d/s of Mylestom (BR8) 0 503625 6628698 BELL4 Bellinger @ Marx Hill, Fernmount (BR4) 4 491836 6631113 BELL6 Bellinger @ 1st bridge on Darkwood Rd (BR2) 41 477971 6633700 KALR1 Kalang @ Pacific Hwy bridge (KR11) 0 501361 6626852 KALR6 Kalang @ bridge on Sunny Corner Rd (KR3) 23 487594 6627840 NNCK1 Never Never @ bridge at Arthur Keough Reserve (NN1) 20 486086 6636107 SPKC1 Spicketts @ Spicketts Creek bridge (SC1) 36 488101 6624416 ROSR1 Rosewood @ bridge on Summervilles Rd (RW1) 45 478649 6635019

Sampling was completed by staff from Bellingen Shire Council (BSC), North Coast Local Land Services (NC LLS), NSW Office of Environment and Heritage (OEH) and the University of New England (UNE) (Appendix 3). Physical water quality variables were measured on site using a calibrated Quanta Hydrolab multiprobe. The following variables were measured at the surface (0.1m) and at every metre until 0.2m above the channel floor:  Temperature (°C)  Specific Conductivity (µS/cm)  pH  Salinity (PPT)  Dissolved Oxygen (mg/L and % Saturation)

Additionally, the following observations were made at each site on each sampling occasion where possible:  Maximum depth (m)  Secchi Depth (m)

Water samples were collected from surface waters on each sampling occasion and transported to UNE for analysis at UNE’s Environmental Chemical Analysis Laboratory. Water chemistry variables analysed from filtered or unfiltered samples were:  Total Nitrogen (µg/L)  Total Phosphorus (µg/L)  Bioavailable Nitrate-Nitrate (µg/L)  Soluble Reactive Phosphorus (µg/L)  Chlorophyll a (µg/L)  Total Suspended Solids (mg/L)  Turbidity (NTU)

Nutrient data were compared with NSW MER and ANZECC guidelines for healthy aquatic ecosystems to provide trigger thresholds beyond which aquatic ecosystems may become stressed. NSW MER trigger values are available for the Bellinger-Kalang Rivers Catchment for Total Nitrogen, Total Phosphorus and chlorophyll a concentrations plus physical parameters ANZECC guidelines were used for parameter exceedances of bioavailable nitrogen (NOx) and phosphorus (SRP) in this report (Table 2).

Table 2. NSW MER trigger values and ANZECC for the Bellinger-Kalang Rivers.

DO EC Turbidity Chla NOx* SRP TN TP Category pH (%) (µScm) (NTU) (µgL) (µgL) (µgL) (µgL) (µgL)

90- Freshwater sites >150m 6.5 – 8.0 30 - 350 2-25 5 25 15 250 20 110

85- Freshwater sites <150m 6.5 – 8.5 125 - 2200 6-50 5 40 20 350 25 110

80- Estuary sites 7.0 - 8.5 no values 0.5-10 4 15 5 300 30 110

3. Results

3.1 NSW MER Water Quality Objectives and ANZECC guidelines during low flows

3.11 Nutrients

Total nitrogen (TN) concentrations remained below MER guidelines at freshwater sites only in T2 (February) 2016 (Figure 1a). Exceedance of TN estuary trigger values were recorded at all sites in T3 (April) and T4 (July) peaking at 626 µg/L at BELL1 in T3. Freshwater sites were also above the trigger values only in T3 and T4, with the highest values of 776 µg/L, 819 µg/L and 605 µg/L recorded in the Rosewood, Never Never and Spicketts sites respectively.

Figure 1. Mean (red line), median (black line), 25th and 75th percentiles for (a) total nitrogen, (b) total phosphorus, (c) bioavailable nitrogen and (d) soluble reactive phosphorus in the eight study sites. Red areas represent exceedances of ANZECC and NSW MER trigger thresholds for lowland freshwater

7 systems (sites represented by blue bars) or estuarine systems (sites represented by green bars). Nutrients have maximum trigger thresholds.

Total phosphorus (TP) concentration consistently exceeded the freshwater and estuary trigger values in all 2016 sample periods, but was generally lower than trigger values in November 2015 (Figure 1b). TP exceeded the MER trigger at all sites in April and July 2016, and by more than four times in the upper estuary site BELL4 at 134 µg/L, and in the upper Bellinger River (BELL6) at 138 µg/L (Figure 1b).

Bioavailable nutrients are directly able to be assimilated by aquatic flora and fauna. NOx concentrations were consistently above the trigger value in estuary sites for all sample times, peaking at 286 µg/L (19 times trigger value) in the Bellinger and 241 µg/L (16 times the trigger value) in the Kalang estuaries (Figure1c). NOx concentrations exceeded MER trigger values in all freshwater sites on all occasions except T1 when values were generally lower. Linked to TN, NOx concentrations were highest in T4 (July 2016) peaking at 353 µg/L in Spicketts Creek and 331 µg/L in the Rosewood River.

All upper and lower estuary sites in the Bellinger and Kalang Rivers exceeded the MER guidelines for SRP at all sampling times with maximum bioavailable phosphorus (SRP) concentrations of 84 µg/L at BELL4 and 49 µg/L at KALR1 in T4 (Figure 1d). Never Never Creek (NNCK1), Rosewood River (ROSR1) and Spicketts Creek (SPKC1) did not exceed the ANZECC guideline for SRP in the study. In contrast, the upper Bellinger River recorded the maximum SRP concentration of 86 µg/L in T4.

3.12 Physico-chemistry

The MER trigger threshold for pH for healthy aquatic ecosystems is 6.5-8.5 for freshwater systems and 7.0-8.5 for estuarine systems. The main trend evident is a great variety in the pH over time and among sites, ranging from 5.9 in T3 to 10.88 in T1 in Spicketts Creek. Sites consistently exceeded the maximum trigger threshold for pH (i.e. more alkaline) in November 2015, February2016 and April 2016 (Figure 2a), but may be a common occurrence for streams draining basalt catchments. The lower Kalang estuary (KALR1) fell below the minimum trigger threshold for pH (i.e. more acidic) in November 2015. In Spicketts Creek (SPKC1) during April 2016, pH fell below the trigger values at <6, with SPKC1 and KALR1 the only sites to have recorded an acidic pH in the study.

The MER trigger threshold for dissolved oxygen saturation for healthy aquatic ecosystems is 85-110% in freshwater reaches and 80-110 in estuaries. Estuary sites were consistently within the trigger value except BELL4 in T1 when dissolved oxygen of 78% was recorded (Figure 2b). The upper Bellinger (BELL6) was also only below the freshwater trigger value in T1. Spicketts (SPKC1), Never Never (NNR1) and the upper Kalang (KALR1) were below the freshwater trigger value on all sample occasions. Dissolved oxygen concentrations in Spicketts were low enough to negatively impact aquatic fauna and promote the release of nutrients and contaminants from sediments, and were associated with the low pH recorded in T3. The extremely low saturation observed in April 2016 may be in response to the high algal bloom as the microbial breakdown of dead algal cells consumes the available oxygen from the water column, or an influx of decomposing organic matter to the site. Low DO% in the upper Kalang River

(KALR6), Never Never Creek (NNCK1) and Rosewood River (ROSR1) in February and April 2016 are likely due to the extremely low discharges during this time. As discharge decreases velocity also decreases, reducing the amount of oxygenation due to turbulent water movement.

Figure 2. Mean (red line), median (black line), 25th and 75th percentiles for (a) pH, (b) saturated DO, (c) chlorophyll a and (d) turbidity in the eight study sites. Red areas represent exceedances of ANZECC and NSW MER trigger thresholds for lowland freshwater systems (sites represented by blue bars) or estuarine systems (sites represented by green bars). pH and DO% have minimum (lower) and maximum (upper) trigger thresholds while chlorophyll a and turbidity have maximum trigger thresholds.

Concentrations of chlorophyll a are used to estimate algal productivity in the water column. Chlorophyll a remained well below MER guidelines in estuary sites (Figure 2c), and were below trigger values in freshwater sites throughout the study despite the very high nitrogen and phosphorus concentrations recorded. All sites recorded turbidities below trigger thresholds at all times (Figure 2d).

4. Post Flood Sampling Event

4.1 Water chemistry

Water quality was sampled on 23 June 2016 immediately following a flood event (~53,000ML/d at Fosters) on 23 June. Total Nitrogen (TN) concentrations were below the MER trigger values at all sites except Spicketts Creek where 360 µg/L was recorded. In contrast, all TP concentrations were well above MER trigger values and above the mean low flow concentrations for all sites, between 138 µg/L at BELL4 and 375 µg/L at SPCK1, or between 4 and 15 times the trigger value.

The event concentrations of bioavailable nitrogen (NOx) and phosphorous (SRP) exceeded the MER trigger values and mean low flow values at all freshwater and estuary sites. NOx and SRP concentrations exceeded the maximum low flow values at all sites except the lower Bellinger estuary (BELL1). The concentrations of NOx significantly exceeded the maximum low flow value in Never Never Creek, and the SRP concentration significantly exceeded those of low flows in Spicketts Creek (Table A2.1).

The chlorophyll a concentrations were lower than all mean low flow values at all the sites after the high flow event (Table A2.1). Although turbidity was not measured during this event, total suspended solid (TSS) concentrations were high in both the lower estuaries of the Bellinger and Kalang Rivers (Table A2.1).

Sites were consistently in the MER guideline range for pH after the high flow event with the exception of Spicketts Creek: SPKC1 had an alkaline reading higher than the mean low flow value for that site, but lower than the maximum recorded value (Table A2.2). Dissolved oxygen concentrations were marginally lower than the MER trigger values in the upper Bellinger estuary (BELL4), and the lowland sites in the Kalang River (KALR6), Never Never Creek (NNCK1), and Spicketts Creek (SPCK1; Table A2.2).

4.2 Microbial Source Tracking

Escherichia coli (E. coli) are normal inhabitants of the gastrointestinal tract of humans and animals. However, these bacteria are ecologically versatile and have adapted to a variety of host species and environmental condition with some species and strains being considered low risk and “Environmental” types. Microbial Source Tracking to detect E.coli concentration and source was undertaken at two end- of-system sites in the Bellinger (BELL1) and Kalang (KALR1) Rivers in June 2016 following a high flow event. Both sites recorded a low risk of fecal contamination, with concentrations of 180 CFU/100mL and 260 CFU/mL at BELL1 and KALR1 respectively.

There were no E. coli from human origin detected in the study. The A1 (Mammalian/Mammalian Omnivore), B1 (Environmental Herbivore) and A0 (Avian) phylo-groups were recorded in the Bellinger River, with only the A1 and B1 strains in the Kalang. None of the E.coli recorded are infective strains, therefore they would be constituted as low risk. There were 5 dominant sources of E.coli in the Bellinger; 3 Avian, 1 Mammalian Omnivore and 1 Environmental Herbivore. The Kalang River also

10 recorded 5 dominant sources of E.coli with 1 Mammalian Omnivore and 4 separate Environmental Herbivores.

The E.coli fingerprints have some correlation (85 - 91.7%) indicating a similar source in both the Bellinger and Kalang Rivers, most likely birds or cows and/or another type of herbivore (e.g. kangaroos). Another possibility is the flow of the E.coli from upstream, but the source cannot be determined from single locations on each river. However, the E.coli coming from the mammalian gut would not last more than 2 weeks in the environment. Therefore, the E. coli counts (180-260 cfu/100mL) would quickly diminish in this time period to 0 cfu/100mL or very low counts. Further sampling would confirm if ongoing faecal contamination was evident if E.coli counts increased with distance downstream or with flood flows.

Appendix 1 – Detailed water quality results for all sites for the low flow period Table A1.1 Minimums, maximums and means of measured water quality variables for the Bellinger River

BELL1 BELL4 BELL6 Variable Min Max Mean Min Max Mean Min Max Mean Temperature (°C) 17.50 27.10 22.65 15.20 26.11 22.44 12.91 54.57 28.35 pH 8.06 9.82 8.94 7.21 9.91 8.18 7.20 18.15 10.08

EC (µS/cm) 22.40 59.50 49.81 0.07 11.80 5.65 0.06 0.15 0.09

Salinity (PPT) 17.04 39.35 32.80 0.04 6.78 3.20 0.04 0.08 0.05

DO (mg/L) 6.02 32.30 12.59 5.64 9.64 7.36 5.50 12.08 8.50

DO (%) 91.30 99.0 88.61 77.90 126.1 87.24 79.60 169.5 103.3

Turbidity (NTU) 3.30 7.50 5.00 2.70 8.70 4.83 0.00 0.35 0.18

Chla (μg/L) 0.12 1.40 0.49 0.00 4.68 1.69 3.50 13.20 6.83

TSS (mg/L) 10.90 17.28 13.31 0.16 5.81 2.77 0.56 1.60 1.04 TN (μg/L) 20.96 236.2 81.24 20.12 511.8 149.7 19.28 354.3 109.9

TP (μg/L) 49.35 145.2 79.76 16.12 148.6 73.65 17.41 148.8 93.83

NOx (μg/L) 14.47 55.35 32.25 14.47 44.68 23.02 1.32 44.69 21.05

SRP (μg/L) 11.89 88.33 33.57 9.86 90.27 33.85 8.51 88.48 28.90

Table A1.2 Minimums, maximums and means of measured water quality variables for the Kalang River and Spicketts Creek.

KALR1 KALR6 SPKC1 Variable Min Max Mean Min Max Mean Min Max Mean Temperature (°C) 18.17 26.70 22.77 13.5 25.9 19.5 12.68 21.73 18.71 pH 6.06 9.74 8.23 7.2 10.0 8.1 5.90 10.68 8.44

EC (µS/cm) 47.30 58.90 53.71 0.1 0.1 0.1 0.11 0.16 0.13

Salinity (PPT) 30.50 38.91 35.33 0.0 0.1 0.0 0.06 0.08 0.07

DO (mg/L) 6.33 29.53 12.23 4.6 8.8 6.5 0.36 7.76 4.08

DO (%) 84.50 100.2 94.18 47.0 82.3 68.3 4.20 72.70 44.40

Turbidity (NTU) 3.80 4.20 4.00 4.2 6.1 5.2 2.70 22.40 11.63

Chla (μg/L) 0.23 3.26 1.32 0.3 4.9 1.9 0.00 7.92 2.48

TSS (mg/L) 12.70 18.56 15.99 1.1 3.6 2.2 0.40 1.70 1.18 TN (μg/L) 20.12 656.8 230.16 46.9 167.6 125.07 20.12 157.5 62.82

TP (μg/L) 12.22 145.2 72.59 15.27 145.2 81.73 15.88 144.1 96.88

NOx (μg/L) 1.32 44.68 22.21 1.3 44.7 16.44 1.32 46.80 21.58

SRP (μg/L) 7.72 87.69 40.59 10.03 88.2 33.68 13.18 85.74 37.69

Table A1.3 Minimums, maximums and means of measured water quality variables for the Rosewood River and Never Never Creek.

ROSR1 NNCK1 Variable Min Max Mean Min Max Mean

Temperature (°C) 12.6 23.3 19.3 14.03 22.24 19.73 pH 7.7 10.1 8.6 7.59 9.87 8.73

EC (µS/cm) 0.0 0.1 0.1 0.04 0.05 0.04

Salinity (PPT) 0.0 0.0 0.0 0.02 0.03 0.03

DO (mg/L) 4.9 9.4 7.1 4.42 8.58 6.37

DO (%) 62.5 104.1 76.9 49.00 84.90 67.60

Turbidity (NTU) 3.2 9.0 5.3 0.23 5.35 1.57

Chla (μg/L) 0.2 0.6 0.4 0.20 2.30 1.13

TSS (mg/L) 0.3 1.3 0.7 28.50 196.9 75.18 TN (μg/L) 20.1 236.2 101.1 27.48 144.1 81.16

TP (μg/L) 15.3 144.4 81.1 1.32 73.80 30.28

NOx (μg/L) 1.3 55.4 33.8 2.10 8.80 5.00

SRP (μg/L) 12.1 87.6 45.8 12.27 87.47 34.09

Table A1.4 Exceedances1 observed at sites for pH, conductivity (EC), percent saturated dissolved oxygen (DO), turbidity, chlorophyll a (Chl-a), total nitrogen (TN) and total phosphorus (TP), and the site-level WQ grades.

Site pH EC DO % Turbidity Chl-a TN TP NOx SRP BELL1 8(50%)0,8 na 2(13%)2,1 0(0%) 0(0%) 0(0%) 4(100%) 2(50%) 4(100%) BELL4 2(20%)0,2 na 3(30%)2,1 0(0%) 1(25%) 1(25%) 3(75%) 2(50%) 4(100%) BELL6 1(20%)1,2 0(0%) 2(40%) 1,1 0(0%) 1(25%) 1(25%) 3(75%) 1(25%) 1(25%) KALR1 8(50%)5,3 na 0(0%) 0(0%) 0(0%) 1(25%) 3(75%) 2(50%) 4(100%) KALR6 1(20%)0,1 0(0%) 5(100%) 5,0 0(0%) 0(0%) 0(0%) 3(100%) 1(33%) 1(33%) SPKC1 6(75%)2,4 0(0%) 8(100%) 8,0 1(33%) 1(25%) 0(0%) 3(75%) 1(25%) 2(50%) ROSR1 3(38%)0,3 0(0%) 6(75%) 6,0 0(0%) 0(0%) 0(0%) 2(67%) 2(67%) 2(67%) NNCK1 2(50%)0,2 0(0%) 4(100%) 4,0 0(0%) 0(0%) 0(0%) 1(25%) 0(0%) 2(50%) 1 Numbers in black represent the total number and percent of exceedances. Numbers in blue and red represent the numbers of measurements lower than the minimum threshold and higher than the maximum threshold, respectively. The number of exceedances includes all depths sampled so may be greater than the number of times sampled. Turbidity, chlorophyll a, and nutrients (TN, TP, NOx and SRP) only have maximum trigger thresholds.

Appendix 2 - Detailed water quality results for all sites for the post flood sampling event

Table A2.1. Nutrient and water quality variables Post Flood Sampling Event for the Bellinger River, Kalang River, Never Never Creek, Rosewood Creek and Spicketts Creek. Exceedances of MER trigger values are in red.

Site TN (µg/L) TP (µg/L) NOx (µg/L) SRP (µg/L) Chla (µg/L) TSS (mg/L) BELL1 62.92 152.76 46.40 73.11 0.12 28.40 BELL4 204.49 138.07 47.40 133.19 0.23 2.20 BELL6 182.02 144.53 47.40 116.96 0.23 1.30 KALR1 182.02 162.76 48.40 100.49 0.35 13.20 KALR6 294.38 187.60 46.40 160.45 0.29 1.74 NNCK1 226.97 170.29 47.40 139.47 0.35 2.10 ROSR1 182.02 183.98 46.40 135.92 1.28 1.00 SPKC1 137.08 375.48 46.40 245.88 0.47 1.28

Table A2.2 Water quality variables Post Flood Sampling Event for the Bellinger River, Kalang River, Never Never Creek, Rosewood Creek and Spicketts Creek, (mean values indicate samples at multiple depths (0.1m, 1m and 1m intervals thereafter). Exceedances of MER trigger values are in red.

Site Temp pH Cond (mS/cm) Salinity (PSS) DO (mg/L) DO% BELL1 (mean, n=3) 19.80 8.41 60.0 39.99 6.54 90.75 BELL4 (mean, n=3) 14.69 8.72 0.10 0.09 8.21 79.5 BELL6 (n=1) 13.52 7.55 0.07 0.03 10.20 96.0 KALR1 (mean, n=5) 19.08 8.15 57.38 37.78 6.51 88.5 KALR6 (n=1) 14.68 7.72 0.09 0.05 8.30 82.3 NNCK1 (n=1) 14.14 7.70 0.04 0.02 8.63 84.3 ROSR1 (mean, n=2) 13.19 7.67 0.05 0.03 9.32 89.0 SPKC1 (mean, n=2) 14.36 9.40 0.13 0.06 7.22 70.5

Appendix 3 – Photos of sites and WQ sampling

Plate 1. NSW Office of Environment & Heritage provided a boat and skipper to access estuarine sample sites (12 July 2016, J. Eales, BSC).

Plate 2. Jane Eales, Bellingen Shire Council, recording Quanta Hydrolab readings at estuarine sites (12 July 2016).

Plate 3. John Schmidt, NSW Office of Environment & Heritage and Jane Eales, Bellingen Shire Council completed the 2015-16 water sampling program after training in Round 1 (24 November 2015).

Plate 4. Water sampling team training by Dr Sarah Mika, University of New England, Aquatic Ecology and Restoration Research Group. Left to right: Dr Sarah Mika, Piers Harper, North Coast Local Land Services, Sandy Eager, Bellingen Shire Council, John Schmidt, NSW Office of Environment & Heritage (24 November 2015, J. Eales, BSC).

Plate 5. BELL4, Bellinger River at Marx Hill, Fernmount (24 November 2015, J. Eales, BSC).

Plate 6. BELL4, Bellinger River at Marx Hill, Fernmount (24 November 2015, J. Eales, BSC).

Plate 7. BELL6, Bellinger River at Lean’s Bridge (first bridge along Darkwood Road from Thora) (12 July 2016, J. Eales, BSC).

Plate 7. BELL6, Bellinger River at Lean’s Bridge (12 July 2016, J. Eales, BSC).

Plate 8. ROSR1, Rosewood River at Summerville’s Road bridge crossing (12 July 2016, J. Eales, BSC).

Plate 9. ROSR1 (12 July 2016, J. Eales, BSC).

Plate 10. NNCK1, Never Never River at bridge crossing at Arthur Keough Reserve. John Schmidt, NSW Office of Environment & Heritage, taking water samples (12 July 2016, J. Eales, BSC).

Plate 11. John Schmidt, Office of Environment & Heritage, filtering water samples for nitrogen and phosphorus analysis, Never Never River (12 July 2016, J. Eales, BSC).

Plate 11. KALR6, Kalang River at bridge crossing, Sunny Corner Road (12 July 2016, J. Eales, BSC).

Plate 12. SPKC1, Spicketts Creek bridge crossing on Bowraville Road (12 July 2016, J. Eales, BSC).

Appendix 4 – Detailed Microbial Source Tracking results for KALR1 and BELL1

E. coli Strain Typing

Sampling Point Description Bellingen Shire Council BELL1 Sampling Point 99380 Sample Identification 4438358 Sampling Date 23/06/2016

E. coli Result 180 CFU/100ml No. of isolates analysed 5

SOURCE EVALUATION* MAMMALIAN OMNIVORE, AVIAN (3) and ENVIRONMENTAL / HERBIVORE types detected

Approved Signatory Lisa TEAKLE (Senior Technical Officer, Method Development)

END OF REPORT

* The source discrimination is not definitive. The determination(s) have been based on the likelihood (%) of E. coli animal excretion rates and gut colonisation data for specific genetic marker groups and serotypes of animal sources (1- 13). Geographical, Ecological, Environmental and Historical epidemiology of possible sources need to be recognised including any unique occurrences of particular strains.

1. Berthe et al. 2013. Applied and Environmental Microbiology. 79(15) 4684-4693 2. Clermont et al. 2013. Environmental Microbiology Reports. 5(1) 58-65 3. Clermont et al. 2011. Environmental Microbiology. 13(9) 2468-2477 4. Clermont et al. 2000. Applied and Environmental Microbiology. 66(10) 4555-4558 5. Dombek et al. 2000. Applied and Environmental Microbiology. 66(6) 2572-2577 6. Duriez et al. 2001. Microbiology. 147(6) 1671-1676 7. Fluit et al. 2010. Journal of Clinical Microbiology. 48(11) 3979-3989 8. Gordon et al. 2010. Journal of AOAC International. 93(3) 974-984 9. Gordon et al. 2003. Microbiology. 149(12) 3575-3586 10. Luo et al. 2011. P.N.A.S. 108: 7200-7205 11. Orsi et al. 2007. Journal of Water and Health. 5(2) 323-327 12. Power et al. 2005. Environmental Microbiology. 7(5) 631-640 13. Tenaillon et al. 2010. Nature Reviews Microbiology. 8: 207-217

E. coli Strain Typing

Sampling Point Description Bellingen Shire Council KALR1 Sampling Point 99380 Sample Identification 4438361 Sampling Date 23/06/2016

E. coli Result 260 CFU/100ml No. of isolates analysed 5

SOURCE EVALUATION* MAMMALIAN OMNIVORE and ENVIRONMENTAL / HERBIVORE (4) types detected

Approved Signatory Lisa TEAKLE (Senior Technical Officer, Method Development)

END OF REPORT