Western Range Ghost Bat Scat Analysis

Western Range Ghost Bat Scat Analysis Report

Biologic Environmental Survey Report to Rio Tinto

March 2020

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Document Status Revision Review / Approved for Approved for Issue to Author No. Issue Name Date

J. McShane, 1 T. Moyle, A. Hutchison C. Knuckey 07/02/2020 S. Reiffer

J. McShane, 2 H. Anderson C. Knuckey 20/03/2020 S. Reiffer

“IMPORTANT NOTE” Apart from fair dealing for the purposes of private study, research, criticism, or review as permitted under the Copyright Act, no part of this report, its attachments or appendices may be reproduced by any process without the written consent of Biologic Environmental Survey Pty Ltd (“Biologic”). All enquiries should be directed to Biologic.

We have prepared this report for the sole purposes of Rio Tinto Iron Ore (“Client”) for the specific purpose only for which it is supplied. This report is strictly limited to the Purpose and the facts and matters stated in it and does not apply directly or indirectly and will not be used for any other application, purpose, use or matter.

In preparing this report we have made certain assumptions. We have assumed that all information and documents provided to us by the Client or as a result of a specific request or enquiry were complete, accurate and up-to-date. Where we have obtained information from a government register or database, we have assumed that the information is accurate. Where an assumption has been made, we have not made any independent investigations with respect to the matters the subject of that assumption. We are not aware of any reason why any of the assumptions are incorrect.

This report is presented without the assumption of a duty of care to any other person (other than the Client) (“Third Party”). The report may not contain sufficient information for the purposes of a Third Party or for other uses. Without the prior written consent of Biologic:

a) This report may not be relied on by a Third Party; and

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In this note, a reference to loss and damage includes past and prospective economic loss, loss of profits, damage to property, injury to any person (including death) costs and expenses incurred in taking measures to prevent, mitigate or rectify any harm, loss of opportunity, legal costs, compensation, interest and any other direct, indirect, consequential or financial or other loss.

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TABLE OF CONTENTS

Executive Summary ...... 5

1 Introduction ...... 7 1.1 Project Background ...... 7 1.2 Objectives ...... 7 1.3 The Ghost Bat ...... 9

2 Methods ...... 10 2.1 Survey Timing and Locations ...... 10 2.2 Climate and Weather ...... 12 2.3 Personnel and Licences ...... 13 2.4 Cave Assessments ...... 13 2.5 Ghost Bat Observations ...... 14 2.6 Scat Counts and Collection ...... 14 2.7 Genetic Analysis ...... 15 2.8 Hormone Analysis ...... 15

3 Results and Discussion ...... 17 3.1 Cave Assessments ...... 17 3.2 Ghost Bat Observations ...... 17 3.3 Scat Monitoring ...... 18 3.4 Genetic Analysis ...... 19 3.5 Hormone Analysis ...... 25 3.6 Limitations ...... 28

4 Conclusion ...... 29

5 References ...... 31

6 Appendices ...... 33

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LIST OF FIGURES Figure 1.1: Project Area ...... 8

Figure 2.1: Survey Locations ...... 11

Figure 2.2: Recent and long term average (LTA) climate data for Paraburdoo Aero (BoM, 2020) ...... 12

Figure 3.1: Scat deposition rates ...... 19

Figure 3.2: Genotyped individuals recorded at each cave ...... 24

Figure 3.3: Genotyped individuals with elevated Progesterone (P4) levels recorded at each cave ...... 27

LIST OF TABLES Table 2.1: Sampling caves and survey timing ...... 10

Table 2.2: Survey Timing ...... 13

Table 2.3: Number of scats collected during each monitoring trip ...... 15

Table 3.1: Number observations of Ghost Bat made during the field survey ...... 17

Table 3.2: Scat number and deposition rate from each cave during the sampling period ...... 18

Table 3.3: Number of genotyped individuals detected by cave during each sampling period ...... 21

Table 3.4: Occurrence of genotyped individuals recorded at each cave...... 22

Table 3.5: Scats containing elevated Progesterone (P4) levels by cave ...... 26

Table 3.6: Study limitations and constraints ...... 28

APPENDICES Appendix A: Cave Assessments ...... 33

Appendix B: Genetic Analysis Report ...... 42

Appendix C: Detailed Hormone Analysis Results ...... 44

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EXECUTIVE SUMMARY

Hamersley Iron Pty Limited, a wholly owned subsidiary of the Rio Tinto Limited (Rio Tinto) owns and operates the Greater Paraburdoo mining operations in the Pilbara bioregion of Western Australia, approximately 6 kilometres (km) south of Paraburdoo. Rio Tinto, on behalf of the joint venture participants, is evaluating the potential development of a new iron ore mine at Western Range (deposits 36 West to 66 West) and the extension of existing operations at Paraburdoo (deposits 4EE, 14W-16W, 20W & 27W) and Eastern Range (deposits 42EE & 47E) within the Greater Paraburdoo locality (the Project Area).

Previous baseline and targeted biological surveys at Western Range have identified a year-round colony of Ghost Bat occurring within the Western Range locality – a Vulnerable listed species under both the Environment Protection and Biodiversity Conservation Act 1999 (EPBC Act) and the Biodiversity Conservation Act 2016 (BC Act). The most recent echolocation data suggests that Ghost Bats may be utilising a number of caves within Western Range, with four caves likely representing non-permanent diurnal roosts (Category 2; Bat Call, 2019). Continued investigation is required to confirm the relative importance of these caves, and of Western Range as a roosting habitat for Ghost Bats. Rio Tinto commissioned Biologic Environmental Survey (Biologic) to undertake a faecal scat study to determine the movement of individuals at previously identified caves and identify the relative significance of these. The key objectives of the study were to:

• determine the relative importance of caves within the area, specifically to conduct detailed cave assessments for all known caves occurring within the Project Area; • define the significance of caves, for Ghost Bats, occurring within the Project Area; • provide an estimate on the local population of Ghost Bats occurring within the Project Area; and • indicate how Ghost Bats use caves within the Project Area with respect to the surrounding region (i.e. permanently vs. transiently).

Monitoring was completed over five trips (surveys) between September 2018 and September 2019. Methods included counting and collection of scats, and genetic and hormone analysis of collected scats. Scats were collected and analysed from six of the seven caves sampled at Western Range, and four of five caves from Mount Truchanas, a disjunct range 35 km north of the Project Area. No scats were found in the two caves sampled at Turee. Tissue samples collected as part of another study completed within in the area, were also included in this study.

Over the monitoring period, eight Ghost Bat individuals were observed from 14 caves, seven within Western Range and one at Mount Truchanas. Each of the 14 caves was visited at least once during the survey period, with some caves visited up to five times, totalling 35 cave visitations. Black sheets were placed on midden piles within all caves to provide temporal resolution for scats deposited. If present, scats were collected during the 35 visitations, though 15 visitations recorded zero scats. Scats were collected from ten out of the 14 caves during the survey. The number of scats deposited within individual monitoring caves ranged from zero to ~500. Cave CPAN-12 at Mount Truchanas recorded the most scats and highest scat deposition rate of 500 scats and a deposition rate of 1.81 scats per day, respectively.

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There was a high success rate of genotyping with 96% of scat and tissue samples tested producing useable genotypes. A total of 54 unique genotypes (unique individuals) were identified from the 246 genotyped faecal and tissue samples, obtained from 11 caves. The highest abundance of individuals recorded at a single cave was 16 (CWRA-11) and 10 (CWRA-06), both caves within the Western Range area. However, this may be attributed to the greater sampling effort expended at these caves over the sampling period.

Six bats were detected using more than one cave. Where individuals were detected using multiple caves, dispersal was most frequent between closely located caves up to 1.5 km apart; however, one instance of an individual flying 11 km between caves at Western Range was recorded. The greatest distance travelled between caves was approximately 36 km, with an individual (583) travelling between CWRA-16 within Western Range to CPAN-18 at Mount Truchanas between February 2019 and September 2019.

A total of 230 Ghost Bat scats were collected during the 2018-19 breeding season and analysed for levels of progesterone faecal metabolite. Approximately one quarter (25%, n = 59) contained elevated progesterone levels (i.e. progesterone levels exceeding 970 ng/g) indicating to be from pregnant individuals.

Due to the possibility of contamination of non-pregnant bat scats by pregnant females, the specifics around individuals are not presented. Ten caves recorded elevated progesterone levels, with six of the caves from Western Range and four from Mount Truchanas. Cave CWRA-16 was likely to be used by a maternity group during the 2019 breeding period as indicated by the high portion of scats containing elevated progesterone levels (70%). However, the total number of scats recorded in the cave at this time was lower than would be expected if it were to be used as maternity cave, suggesting it was only being used for short visitations or temporary diurnal roosting. Four other caves (CPAN-18, CWRA-06, CWRA- 11 & CWRA-14) also contained scats with elevated progesterone, suggesting they were used by pregnant females very scarcely, and were not used as maternity caves during the course of the study.

The genetic effective population size at Western Range is estimated to be 11.3 and is consistent with other populations of a similar area within the Central Hamersley Ranges. However, it is likely that the caves monitored are a part of a larger network of caves in the surrounding region. Only one individual (of 43 bats, 2% chance) was recorded at both Western Range and Mt Truchanas, a distance of over 35 km (directly) and the furthest distance travelled known from within the region. While this record demonstrates the scale of movements which can be undertaken by the species, the scarcity of more such records makes it unclear where bats are originating from and/ or travelling to, though a location other than Mt Truchanas appears likely, such as the unsearched ranges located ~20 - 30 km to the east of Western Range, further along Paraburdoo Range, and the range ~20 km to the west of the Project Area.

Previous VHF tracking studies have demonstrated that the majority of individuals (though not all) forage in the plains to the north and south of Western Range, suggesting that movement of the bats is not restricted to the range itself. However, the associations between primary foraging grounds and general movement in somewhat still unknown. Due to this unknown, development should be mindful of restricting movements along the range from neighbouring areas.

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1 INTRODUCTION

1.1 Project Background Hamersley Iron Pty Limited, a wholly owned subsidiary of the Rio Tinto Limited (Rio Tinto) owns and operates the Greater Paraburdoo mining operations in the Pilbara bioregion of Western Australia, approximately 6 kilometres (km) south of Paraburdoo (Figure 1.1). Rio Tinto, on behalf of the joint venture participants, is evaluating the potential development of a new iron ore mine at Western Range (deposits 36 West to 66 West) and the extension of existing operations at Paraburdoo (deposits 4EE, 14W-16W, 20W & 27W) and Eastern Range (deposits 42EE & 47E) within the Greater Paraburdoo locality (the Project Area).

Previous baseline and targeted biological surveys at Western Range have identified a year-round colony of Ghost Bat occurring within the Western Range locality – a Vulnerable listed species under both the Environment Protection and Biodiversity Conservation Act 1999 (EPBC Act) and the Biodiversity Conservation Act 2016 (BC Act). The most recent echolocation data suggests that Ghost Bats may be utilising a number of caves within Western Range, with four caves likely representing non-permanent diurnal roosts (Category 2; Bat Call, 2019). Continued investigation is required to confirm the relative importance of these caves, and of Western Range as a roosting habitat for Ghost Bats.

Ghost Bats were identified at Western Range through one echolocation record in 2009 (Biota, 2010), followed by scat records in 2013 (Astron, 2013). Targeted surveys revealed two confirmed diurnal roosts and two potential diurnal roosts, along with two echolocation records (Astron, 2018b). Since then, monitoring of caves at Western Range has occurred with a total of 18 caves being monitored (Astron, 2018a; Bat Call, 2019, 2018). Biologic has also completed VHF studies over the area since September 2018, however sampling effort has focused on already identified caves (Biologic, 2020).

1.2 Objectives Rio Tinto commissioned Biologic Environmental Survey (Biologic) to undertake a faecal scat study to determine the movement of individuals at previously identified caves and identify the relative significance of these. The key objectives of the study were to:

• determine the relative importance of caves within the area, specifically to conduct detailed cave assessments for all known caves occurring within the Project Area; • define the significance of caves, for Ghost Bats, occurring within the Project Area; • provide an estimate on the local population of Ghost Bats occurring within the Project Area; and • indicate how Ghost Bats use caves within the Project Area with respect to the surrounding region (i.e. permanently vs. transiently).

Page | 7 495631 509631 523631 537631 551631 565631 579631

PARABURDOO

7445310 !

W e s t e r n R a n g e PARABURDOO ! CREEK 0# MILE 7431310

SEVEN P a r a b u r d o o R a n g e CREEK

PIRRABURDU SIX MILE CREEK

CREEK SEVEN MILE 7417310 7403310 Legend Rio Tinto Iron Ore Western Range PLNB VHF Study 0# Roost Figure 1.1: The Project Area GreaterMajor Paraburdoowaterline 1:300,000 Study Area Coordinate System: GDA 1994 MGA Zone 50 0 3.5 7 14 Projection: Transverse Mercator ¯ km Datum: GDA 1994 Size A4. Created 13/01/2020 Western Range Ghost Bat Scat Analysis

1.3 The Ghost Bat The Ghost Bat is listed as Vulnerable under the EPBC Act, the BC Act and by the International Union for Conservation of Nature (IUCN). In the Pilbara region, the species occurs in all four subregions (DBCA, 2018). The Pilbara population is estimated to be between 1,300 and 2,000 individuals (TSSC, 2016). The largest colonies occur within the Chichester subregion (total population estimated at approximately 1,500 individuals) where colonies are known to occur within disused mines (TSSC, 2016).

The distribution of Ghost Bats in the Pilbara is dependent on the presence of suitable roosting sites and foraging areas. Natural roosts generally comprise deep, complex caves beneath bluffs or low rounded hills (Armstrong & Anstee, 2000). There are few known maternity roosts in natural caves of the Pilbara (Armstrong & Anstee, 2000). Based on available data, breeding has been documented in natural caves at Mining Area C, Mount Brockman and West Angeles in the Hamersley subregion, and at Callawa and Tambrey Station in the Chichester subregion (Armstrong & Anstee, 2000). Ghost Bats move between a number of caves seasonally, or as dictated by weather conditions, and require a range of cave sites (Hutson et al., 2001). Outside of the breeding season, male bats are known to disperse widely, most likely during the wet season when conditions would allow bats to use caves that would otherwise not be suitable (Worthington-Wilmer et al., 1994). Genetic studies indicate that females are likely to stay close to the maternity roosts (Worthington-Wilmer et al., 1994).

Caves used by the species can be classified into five types: Night Roosts (caves that are only utilised during the night, mostly to feed on prey items or to rest, and are typically shallow caves and shelters/overhangs that can be well lit during the day); Night Roost/possible Diurnal Roosts (in addition to being utilised to feed on prey items during the night, these caves are more complex in structure and provide suitable temperature and humidity conditions suitable for roosting during the day, but no Ghost Bats have been recorded using them during the day); Diurnal Roosts (caves that are used for shelter during the day); Diurnal Roosts/possible Maternity Roosts (in addition to being used for shelter during the day, support breeding and Ghost Bats have been recorded in such caves during the breeding season); and Maternity Roosts (in addition to being used for shelter during the day and supporting breeding, these caves contain pregnant females or females with pups) (Biologic, 2015).

An average foraging area of 61 ha was recorded for Ghost Bats in the Northern Territory whereby Ghost Bats appear to have a short-range foraging strategy of up to 3 km (average 1.9 km), with vantage points changing approximately every 15 minutes. Moreover, the bats generally return to the same area each night (Tidemann et al., 1985). Although it has been suggested that Ghost Bats in the arid zone are semi- transient through most areas and will readily travel large distances (>4 km) (Biologic, 2019). Ghost Bats have a ‘sit and inspect’ foraging strategy; they hang on a perch where they visually inspect their surroundings for movement. Once their prey is detected it may be captured in the air, gleaned (taken from the surface of a substrate by a flying bat) from the ground or vegetation, or dropped on from a perch (Boles, 1999).

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2 METHODS The current project aimed to provide targeted analyses on Ghost Bats utilisation of caves in three focal cave systems: Western Range, Mount Truchanas and Turee (Figure 1.1). The objectives of the assessment were completed by undertaking standardised cave assessments at all sampling caves, calculating scat deposition rates at these caves to indicate usage patterns, completing genetic analysis to determine population dynamics and completing progesterone faecal analysis to identify visitations by pregnant individuals. Sampling for the study was completed over five separate trips between September 2018 and September 2019.

2.1 Survey Timing and Locations A total of 14 caves were monitored over the course of the study, equating to a total of 35 cave visitations (Table 2.1). The caves monitored are a subset of the known caves within the area and represent caves considered to be likely day roosts and/or maternity roosts as indicated by Bat Call WA and Astron (the status of each cave is detailed in the Appendix A). The selection of caves sampled was chosen by Rio Tinto personnel.

Sampling caves were grouped geographically: Western Range, a cluster of caves located upon the Paraburdoo Range and within the vicinity of the anticipated Project; Mount Truchanas, a disjunct range located approximately 35 km north of Western Range; and Turee which represents the eastern section of Paraburdoo Range and is located approximately 30 km east of Paraburdoo (Figure 1.1). All areas are west of Karijini National Park (Figure 2.1).

Table 2.1: Sampling caves and survey timing

Trip 1 Trip 2 Trip 3 Trip 4 Trip 5 Bat Total cave Cave ID Call/Astron 11/09/2018 20/02/2019 3/05/19 23/08/19 19/09/19 visitations Cave ID – – – – – per cave 12/09/2018 26/02/2019 6/05/19 27/08/19 30/09/19 Mt Truchanas caves CPAN-10 - x x - - x 3 CPAN-12 - x x - - x 3 CPAN-13 - x x - - x 3 CPAN-18 - x x - - x 3 CPAN-23 - x x - - x 3 Western Range caves CWRA-06 Cave 6 x x x x x 5 CWRA-11 Cave 11 - - x x x 3 CWRA-14 Cave 14 - x - x x 3 CWRA-15 Cave 15 - x - - x 2 CWRA-16 Cave 16 - - - - x 1 CWRA-18 Cave 18 - x - x x 3 Ratty Spring Ratty Spring x - - - - 1 Cave Roost Turee caves CTRE-03 - - - - - X 1 CTRE-05 - - - - - X 1 Total caves 7 9 2 4 13 35

Page | 10 518739 532739 546739 560739 574739 588739 602739 616739

7483253 !(CPAN-13

Mount Trunchanas

CPAN-10 CPAN-23!( CPAN-12!( 7469253 !(CPAN-18 7455253

W e s t e r n R a n g e T u r e e 7441253 CTRE-05!( !(CTRE-03 !(CWRA-11 CWRA-15 !(CWRA-16!(!(CWRA-06 CWRA-18!( PARABURDOO !( ! CWRA-14 !(Ratty Cave 7427253 Legend Rio Tinto Iron Ore Caves Western Range Ghost Bat Scat Analysis !( Mt Truchanas Figure 2.1: Survey Locations Greater Paraburdoo 1:350,000 !( TureeStudy Area Coordinate System: GDA 1994 MGA Zone 50 0 4 8 16 !( Western Range Projection: Transverse Mercator ¯ km Datum: GDA 1994 Size A4. Created 13/01/2020 Western Range Ghost Bat Scat Analysis

2.2 Climate and Weather During the sampling period (September 2018 to September 2019), Paraburdoo Aero Station (station number 007185, ~18 km northeast of Western Range) recorded 120.6 millimetres (mm) of rainfall, which is lower than the long-term annual average rainfall for the same period (313.1 mm) (BoM, 2020) (Figure 2.2). Observed minimum and maximum monthly temperatures were consistent with the long-term annual average temperatures for the same period. Ghost Bat activity and populations may vary with seasonal fluctuations (Johnston et al., 2015; Toop, 1985). Therefore, the below average annual rainfall experienced over the survey period may have influenced Ghost Bat activity across the monitoring sites.

50 80

45 70

40 60

35 C)

° 50 30

25 40 Rainfall (mm)

Temperature ( 20 30

15 20 10

10 5

0 0

Jul-19

Oct-18 Apr-19

Jan-19 Jun-19

Mar-19

Feb-19

Sep-18 Nov-18 Dec-18 Aug-19 Sep-19 May-19 Observed Rainfall (mm) LTA Rainfall (mm) LTA Min Temp (°C) LTA Max Temp (°C) Observed Min Temp (°C) Observed Max Temp (°C)

Figure 2.2: Recent and long term average (LTA) climate data for Paraburdoo Aero (BoM, 2020)

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2.3 Personnel and Licences The surveys were completed under Licence to Take Fauna for Scientific Purposes (Regulation 17) 08- 001704-4 (17/07/2018- 09/01/2019) issued to Mr. Chris Knuckey and (Regulation 17) 08-003289-1 (01/01/2019- 31/12/2019) issued to Mr. Chris Knuckey. Five surveys were undertaken by experienced ecologists between September 2018 and September 2019 (Table 2.2).

Table 2.2: Survey Timing

Trip Date Field team Tasks performed Thomas Rasmussen, 20 – 26 February Scat collection 1 Arnold Slabber, 2019 Talitha Moyle Ghost Bat tagging

Thomas Rasmussen, Scat collection 2 3 – 6 May 2019 Aidan Williams Ghost Bat tagging

23 – 27 August Thomas Rasmussen, 3 Scat collection 2019 Aidan Williams

Thomas Rasmussen, 19 – 30 September Scat collection 4 Aidan Williams, 2019 Syngeon Rodman Ghost Bat tagging

20 – 24 November Thomas Rasmussen, 5 Scat collection 2019 Aiden Williams

2.4 Cave Assessments Cave assessments were conducted for every cave sampled as part of the study. The key objective of the cave assessments was to identify the perceived and potential importance of each structure to the Ghost Bat. Each cave assessment recorded the following characteristics:

• Entrance location; • Entrance photograph; • Entrance: type, position, aspect, exposure, width, and height; • Floor slope; • Cave depth; • Chambers: main chamber height, and number of total chambers; • Water presence; • Indicative roost type; • Presence of target species: no. individuals, and/or secondary evidence.

Additionally, each cave was categorised as one of the following:

• Nil; • Possible Night Roost; • Night Roost / Possible Diurnal Roost; • Diurnal Roost;

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• Diurnal Roost / Possible Maternity Roost; • Maternity Roost.

2.5 Ghost Bat Observations Each of the monitoring caves was visited and inspected by two Biologic personnel. An ‘inspector’ entered the cave and the ‘spotter’ waited at the entrance of the cave to record any Ghost Bats flushed out of the cave and ensure the safety of the inspector. The Ghost Bat is distinctive from all other cave-dwelling bats within the Pilbara region, being both larger in size and lighter in colouration and was therefore easily distinguished if present. Any signs of reproduction (presence of pregnant females or pups) were noted.

2.6 Scat Counts and Collection The inspector searched the floor of each cave for scat material, typically in one or two piles in the darkest part of the cave (scats and middens are distinctive for this species, with scats being almost twice the size of other cave-dwelling species). Black sheets (measuring approximately 10 m2) were deployed on top of middens or large scat piles within each cave during previous surveys. The number of sheets within each cave varied according to the number and spatial spread of scat piles within each cave and was not dictated by usage at each cave. Up to 40 fresh scats were collected from the sheets during each time the cave was visited, although typically a nominal amount of 20 was collected (Table 3.2). Any scats not collected were cleared from the sheets to ensure an accurate scat count for the next sampling event. Searches were also conducted for scats off sheet to identify new scat piles. Scats were collected and stored in individual paper envelopes and were stored in a freezer until analysis. Three sampling events collected scats not on sheets, these are noted within Table 3.2.

The number of scats occurring both on-sheet and off-sheet were counted and recorded. It was not uncommon for sheets to sometimes be disturbed by other fauna, such as wallabies or goannas, and this was considered when determining the scat deposition rate. Total scats recorded on-sheet was later used to determine a scat deposition rate (the number of scats recorded divided by the number of days since sheets were last cleared). Owing to the variability of results, deposition rates were normalised using a square root transformation. The Mount Truchanas caves were previously sheeted by Biologic as part of another study (27-30/06/2017).

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Table 2.3: Number of scats collected during each monitoring trip

Trip 1 Trip 2 Trip 3 Trip 4 Trip 5 Total scats Cave ID 11/09/2018 20/02/2019 3/05/19 23/08/19 19/09/19 - - - - - collected 12/09/2018 26/02/2019 6/05/19 27/08/19 30/09/19 Mount Truchanas caves CPAN-10 * 0 - - 26 26 CPAN-12 * 0 - - * 0 CPAN-13 * 0 - - 20 20 CPAN-18 * 0 - - 13 13 CPAN-23 * 0 - - 4 4 Western Range caves CWRA-06 20 * 20 6 2 48 CWRA-11 - - 21 15 0 36 CWRA-14 - 16 - 20 0 36 CWRA-15 - * - - 0 0 CWRA-16 - - - - 40 40 CWRA-18 - 20 - 0 0 20 Ratty Spring 6 - - - - 6 Cave Turee caves CTRE-03 - - - - 0 0 CTRE-05 - - - - 0 0 Note: * refers to caves that were visited during this period, but scat not recorded/collected. - Refers to caves not visited

2.7 Genetic Analysis Survey work undertaken throughout the Hamersley subregion has documented many known and potential Ghost Bat caves. However, how these caves are utilised and the extent and structure of the Ghost Bat population within the subregion is poorly known. Obtaining information on caves use by Ghost Bats is problematic due to the low number of bats present at any one time and the fact that Ghost Bats use multiple roosts and thus may not be present within a monitoring roost at the time of sampling. Genetic analyses of Ghost Bat tissue and scats is deemed the most appropriate approach for increasing knowledge of cave usage and movement of Ghost Bats (Biologic, 2016).

A total of 251 Ghost Bat scat samples and seven tissue samples from the 14 cave sites (11 caves had samples, three had zero samples) were genotyped by the genetics laboratory of the Department of Biodiversity Conservation and Attractions (DBCA) to determine the number of unique individuals and distinct genetic populations that had visited the caves between monitoring periods. The data was then analysed to determine dispersal patterns between caves (see Appendix B for a complete description of analysis techniques).

2.8 Hormone Analysis Scats collected between September 2018 - September 2019 were also analysed for faecal metabolites (hormones) to determine the occurrence of pregnant individuals within caves. Hormone analysis was undertaken by Dr Tamara Keeley from the University of Queensland to determine faecal progesterone levels within scats, indicative of pregnancy among individuals within the population (Appendix C). The

Page | 15 Western Range Ghost Bat Scat Analysis method has previously been used for numerous other bat species (Keeley et al., 2012a; Keeley et al., 2012b) and has been validated for Ghost Bat in a pilot study using the captive breeding colony from Perth Zoo (Keeley, 2018). Material from the Perth Zoo colony was also used to compared and validate analysis of samples collected from wild populations. Samples with progesterone levels above 970 nanograms per gram (ng/g) were considered likely to represent those originating from pregnant females (Keeley, 2018).

Only whole samples were analysed to avoid the possibility of hormone levels being artificially altered if pooled faecal sample pieces came from different individuals of differing reproductive status, resulting in a total of 230 samples being analysed. Faecal samples were analysed for progesterone metabolite concentrations by enzyme-immunoassay (EIA). Prior to analysis for hormone concentrations, each faecal sample was extracted using a basic hormone extraction procedure (Keeley et al., 2012a; Palme et al., 2013). Faecal samples were subsampled to a weight of either 0.1 ± 0.02 or 0.05 ± 0.002 grams (g) to which 5 ml of 80% methanol was added. Samples were rotated gently overnight, centrifuged at 1000 g for 10 min and then decanted and stored at -20°C until analysis. The supernatant was diluted 1:20 to 1:1000 (dependant on concentration) in assay buffer prior to analysis. Faecal progesterone metabolite concentrations were quantified by double antibody EIA using a goat anti-mouse IgG (Arbor Assays, USA), monoclonal progesterone antiserum (CL425), horseradish peroxidase conjugated label (both provided by C. Munro, University of California-Davis, Davis, USA) and progesterone (Sigma Aldrich Australia Ltd.) standards as previously described with minor modifications (Keeley et al., 2012b).

The antiserum (1:80,000) was incubated on a microtitre plate overnight, horseradish peroxidise conjugate (1:400,000), standards (0.016 - 4 nanograms per millimetre (ng/ml) and samples were loaded (50 µl/well) onto the plate and the EIA was performed as described elsewhere (Keeley et al., 2012b; Pollock et al., 2010). Intra and inter-assay coefficients of variation were both <10%. Cross-reactivities for the EIA antibodies were as previously described (Graham et al., 2001). Hormone concentrations were expressed as nanograms of hormone metabolites per gram of faeces (ng/g). Progesterone levels were considered to be elevated (i.e. progesterone levels indicating pregnancy or individual) for results of 970 ng/g or greater.

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3 RESULTS AND DISCUSSION

3.1 Cave Assessments Cave assessments were undertaken at all caves visited, fourteen caves in total, seven caves from Western Range, five caves from Mount Truchanas and two caves from Turee were monitored. Detailed cave assessments are available in Appendix A.

3.2 Ghost Bat Observations Overall, eight Ghost Bat individuals were observed from four of the 14 caves monitored during the sampling period (Table 3.1). The locations comprised seven individuals in the Western Range caves, one in the Mount Truchanas caves and zero at the Turee caves. No cave recorded individuals on more than one visitation.

Five Ghost Bats were recorded in cave CWRA-11 during Trip 3 (May 2019), one single Ghost Bat was recorded in caves CWRA-06 and CWRA-14 during Trip 2 (February 2019) and one single Ghost Bat was recorded from cave CPAN-12 during Trip 5 (September 2019).

As only eight individuals were recorded over the 35 visitations in the area, it is likely that these areas form part of a larger home range. As this is the first year this monitoring has been undertaken in the area, and the species is renowned for their sporadic and nomadic cave use (Armstrong & Anstee, 2000), it is difficult to ascertain trends from this.

Table 3.1: Number observations of Ghost Bat made during the field survey

Trip 1 Trip 2 Trip 3 Trip 4 Trip 5 Cave ID 11/09/2018 20/02/2019 3/05/19 23/08/19 19/09/19 - - - - - 12/09/2018 26/02/2019 6/05/19 27/08/19 30/09/19 Mount Truchanas caves CPAN-10 0 0 - - 0 CPAN-12 0 0 - - 1 CPAN-13 0 0 - - 0 CPAN-18 0 0 - - 0 CPAN-23 0 0 - - 0 Western Range caves CWRA-06 0 1 0 0 0 CWRA-11 - - 5 0 0 CWRA-14 - 1 - 0 0 CWRA-15 - 0 - - 0 CWRA-16 - - - - 0 CWRA-18 - 0 - 0 0 Ratty Spring Cave 0 - - - - Turee caves CTRE-03 - - - - 0

CTRE-05 - - - - 0

Page | 17 Western Range Ghost Bat Scat Analysis

3.3 Scat Monitoring The number of scats deposited on the black sheets, and hence during a determined period, allows for an assessment of individual cave usage by Ghost Bats and a comparison of usage between the caves. Table 3.2 shows the scat number and deposition rate (number of scats on the sheets divided by the number of days since sheets were last cleared) during the current monitoring survey.

The number of scats deposited within individual monitoring caves ranged from zero to ~500 (Table 3.2). One cave CPAN-12 recorded the most scats in the monitoring with 500 scats recorded during Trip 1 (September 2018), with a deposition rate of 1.81 scats per day. The next highest cave CWRA-14 recorded 200 scats between trip 2 and trip 4, recording a deposition rate of 1.09 scats per day.

Table 3.2: Scat number and deposition rate from each cave during the sampling period

Trip 1 Trip 2 Trip 3 Trip 4 Trip 5 Cave ID 11/09/2018 20/02/2019 3/05/19 23/08/19 19/09/19 - - - - - 12/09/2018 26/02/2019 6/05/19 27/08/19 30/09/19 Mount Truchanas caves CPAN-10 50 (0.18)* 0 (0.00) - - 30 (0.13) CPAN-12 500 (1.81)* 0 (0.00) - - * CPAN-13 40 (0.14)* 0 (0.00) - - 40 (0.17) CPAN-18 0.00* 0 (0.00) - - 13 (0.05) CPAN-23 4 (0.014)* 0 (0.00) - - 4 (0.01) Western Range caves CWRA-06 * * 40 (0.168) 6 (0.05) 2 (0.05) CWRA-11 - - * 20 (0.17) 0 (0.00) CWRA-14 - 16# - 200 (1.09) 0 (0.00) CWRA-15 - 0 (0.00) - - * CWRA-16 - - - - 40 # CWRA-18 - 0 (0.00) - 0 (0.00) 0 (0.00) Ratty Spring Cave 6# - - - - Turee caves CTRE-03 - - - - 0 (0.00) CTRE-05 - - - - 0 (0.00) Note: ‘#’ refers to caves where scats where collected (not on sheets), ‘*’ refers to caves that were visited during the monitoring survey, but no scat collected. Further, this table only includes the caves that were monitored during the 2018-2019 monitoring assessment

Page | 18 Western Range Ghost Bat Scat Analysis

2 1.8 1.6 1.4 1.2 1 0.8 0.6 0.4 0.2 0 Scat deposition Scat deposition rate (scats per day) 11/09/2018 - 20/02/2019 - 3/05/2019 - 23/08/2019 - 19/09/2019 - 12/09/2019 26/02/2019 6/05/2019 27/08/2019 30/09/2019 Trip 1 Trip 2 Trip 3 Trip 4 Trip 5

CPAN-10 CPAN-12 CPAN-13 CPAN-18 CPAN-23 CWRA-06 CWRA-11 CWRA-14 CWRA-18 CWRA-15 CTRE-03 CTRE-05

Figure 3.1: Scat deposition rates

Overall, 35 cave visitations were conducted at 14 caves over the five surveys. The sampling effort between Mount Truchanas and Western Range was substantially different, making it difficult to make direct comparison between the two areas. However, cave CPAN-12 at Mount Truchanas recorded the most scat deposited, and equally highest scat deposition rate recorded during the study, 500 scats and a deposition rate of 1.81 scats, respectively. Based on this Mount Truchanas likely represents an area of equal, or greater, significance for the Ghost Bat within the central Hamersley Ranges.

Ghost Bats move between a number of caves seasonally, or as dictated by weather conditions, and require a range of cave sites (Hutson et al., 2001), with presence ranging from occasional visitation to persistence over a long period. Previous studies have suggested that Ghost Bats in the Hamersley Range occur in small family groups that move from cave to cave (Armstrong & Anstee, 2000; Biologic, 2014).

3.4 Genetic Analysis A total of 246 of 257 (96%) of scat samples and tissue samples tested produced useable genotypes, which is considered extremely high for genetic analysis. An additional eight tissue samples were collected from live bats in the area. Detailed results of the genetic analysis can be found in Appendix B.

A total of 54 unique genotypes (unique individuals) were identified from 249 successfully genotyped scat and tissue samples, including 43 unique genotypes identified within Western Range and 12 unique genotypes identified from Mount Truchanas (Table 3.3; Appendix B). Six of eight tissue samples collected from live-captured bats were matched to scats at the study site (individuals 566, 567, 569, 600, 602, 604). Individuals 617 and 618 were not matched to any scats. Individual 618 was captured from cave CPAN- 12, however, no scats were recorded from this cave.

Page | 19 Western Range Ghost Bat Scat Analysis

The number of individuals recorded from each cave ranged from one to 16 (Table 3.3); however, this is likely to have been influenced by the varied sampling effort between caves over the monitoring period. The highest abundance of individuals was recorded at caves CWRA-11 (n = 16) and CWRA-06 (n = 10). Caves CPAN-18, CWRA-11 and Ratty Spring Cave had higher numbers of individuals detected for the numbers of scats analysed (ratio 0.41-0.50), potentially indicating different activity or residency patterns in these locations. In particular, cave CWRA-11 had 16 unique individuals detected from 39 scats, with most individuals detected in the May 2019 sample. Conversely, despite a larger sampling effort from cave CWRA-06, only 10 unique individuals were detected from 44 scats (ratio 0.23; Appendix B).

For CWRA-11 there was a peak in genotyped individuals in May 2019 (14 unique genotypes detected) and then an additional two in August 2019. Whereas cave CRWA-06 had ten unique genotypes detected in September 18, then in May 19 and September 19, two of those previously detected individuals were again detected. Of the 54 unique genotypes identified within Western Range and Mount Truchanas, six individuals were detected at two different caves (genotyped individuals 566, 569, 572, 573, 583 and 608), the remaining 48 individuals were only detected visiting one cave (Table 3.4).

Two individuals (572 and 573) were both detected moving between caves CWRA-06 and CWRA-18, approximately 1.15 km apart. These individuals were not detected using cave CWRA-16 that is only 240 m from CWRA-06 (Ottewell et al., 2020). Six bats were detected using more than one cave. Where individuals were detected using multiple caves, dispersal was most frequent between closely located caves up to 1.5 km apart; however, one instance of an individual flying 11 km between caves at Western Range was recorded. The greatest distance travelled between caves was approximately 36 km, with an individual (583) travelling between CWRA-16 within Western Range to CPAN-18 at Mount Truchanas between February 19 and September 19. Overall, the mean dispersal distance was 8.6 ± 5.7 km.

Three caves (CWRA-06, CWAR-11 and CWRA-14) were sampled from multiple trips during the monitoring, (Table 3.4). In each of these caves, there appeared to be one to two resident individuals based on the relative abundance of scats and the number of ‘recaptures’: individuals 565 (6 scats total), 566 (25 scats, 1 tissue), 567 (16 scats, 1 tissue), 569 (22 scats, 1 tissue), 570 (10 scats). All bats, apart from 570 (detected in the last sampling period), were detected at multiple sampling times during the study period.

This is the first Ghost Bat study to use DBCAs custom-designed SNP array, so it is not possible to directly compare genetic diversity patterns with previous studies, at this point in time. However, the genetic effective population (genetic diversity) size at Western Range/ Mount Truchanas (Ne = 11.3) is similar to other previously studied populations, e.g. West Angelas Ne = 13.9 (Ottewell et al., 2019a), Robe River Ne = 13.2 (Ottewell et al., 2019b).

Page | 20 Western Range Ghost Bat Scat Analysis

Table 3.3: Number of genotyped individuals detected by cave during each sampling period

Number of Genotyped Individuals from each

sampling period

Cave Trip 1 Trip 2 Trip 3 Trip 4 Trip 5

(Sept (Feb (May (Aug (Sept umber n

18) 19) 19) 19) 19)

Total Individuals Total Individual genotype genotype Individual

Western Range 565, 566, 572, 573, 608, 612, 613, 614, 615, CWRA-06 10 2 2 10 616 567, 568, 599, 600, 601, 602, 603, 604, 605, CWRA-11 14 2 16 606, 607, 608, 609, 610, 611, 617 CWRA-14 3 2 4 569, 570, 578, 579 CWRA-15 0 N/A 566, 582, 583, 584, 585, 586 CWRA-16 8 8 587, 588 571, 572, 573, 574, 575, 576, CWRA-18 7 7 577 Ratty Spring 3 3 569, 580, 581 Cave Mount Truchanas

CPAN-10 3 3 594, 595, 596 CPAN-12 1 1 618 CPAN-13 2 2 597, 598 CPAN-18 5 5 583, 589, 590, 591, 592 CPAN-23 1 1 593 Turee CTRE-03 0 N/A CTRE-05 0 N/A

Page | 21 Western Range Ghost Bat Scat Analysis

Table 3.4: Occurrence of genotyped individuals recorded at each cave.

Western Range caves Mount Truchanas caves Genotyped Caves Ratty individual CWRA-06 CWRA-11 CWRA-14 CWRA-16 CWRA-18 CPAN-10 CPAN-12 CPAN-13 CPAN-18 CPAN-23 visited Spring Cave

565 ● 1

566 ● ● 2

567 ● 1

568 ● 1

569 ● ● 2

570 ● 1

571 ● 1

572 ● ● 2

573 ● ● 2

574 ● 1

575 ● 1

576 ● 1

577 ● 1

578 ● 1

579 ● 1

580 ● 1

581 ● 1

582 ● 1

583 ● ● 2

584 ● 1

585 ● 1

586 ● 1

587 ● 1

588 ● 1

589 ● 0

590 ● 0

591 ● 0

592 ● 0

Page | 22 Western Range Ghost Bat Scat Analysis

Western Range caves Mount Truchanas caves Genotyped Caves Ratty individual CWRA-06 CWRA-11 CWRA-14 CWRA-16 CWRA-18 CPAN-10 CPAN-12 CPAN-13 CPAN-18 CPAN-23 visited Spring Cave

593 ● 0

594 ● 0

595 ● 0

596 ● 0

597 ● 0

598 ● 0

599 ● 1

600 ● 1

601 ● 1

602 ● 1

603 ● 1

604 ● 1

605 ● 1

606 ● 1

607 ● 1

608 ● ● 2

609 ● 1

610 ● 1

611 ● 1

612 ● 1

613 ● 1

614 ● 1

615 ● 1

616 ● 1

617 ● 1

618 ● 1 Total 10 16 4 8 7 3 3 1 2 5 1 54

Page | 23 518739 532739 546739 560739 574739 588739 602739 616739

CPAN-13 7483253

Mount Trunchanas

CPAN-23 CPAN-10 CPAN-12

7469253 CPAN-18 7455253

W e s t e r n R a n g e T u r e e

7441253 CTRE-05 Legend CWRA-11 CTRE-03 Genotyped indivduals CWRA-06 0 CWRA-15 CWRA-16 1 - 4 CWRA-14 PARABURDOO CWRA-18 ! Ratty Cave 5 - 9

10 - 16 7427253 Rio Tinto Iron Ore Western Range Ghost Bat Scat Analysis Figure 3.2: Number of genotyped individuals Greater Paraburdoo recorded at caves 1:350,000 Study Area Coordinate System: GDA 1994 MGA Zone 50 0 4 8 16 Projection: Transverse Mercator ¯ km Datum: GDA 1994 Size A4. Created 13/01/2020 Western Range Ghost Bat Scat Analysis

3.5 Hormone Analysis Of the 230 scat samples analysed, 59 (25.65%) scats contained elevated progesterone levels (i.e. samples with progesterone levels exceeding 970 ng/g) (Table 3.5; Appendix C), from 10 caves, six from Western Range (CWRA-06, CWRA-11, CRWA-14, CWRA-16, CWRA-18 and Ratty Spring Cave) whereas four from Mount Truchanas had elevated progesterone levels (CPAN-10, CPAN-13, CPAN-18 and CPAN-23).

Two samples recorded intermediate levels of progesterone (700-950ng/g) levels, one from cave CPAN- 18 and the other CWRA-14, suggesting the hormone was either decaying due to the age of the scat, or the individual may have recently been/ at early stages of pregnancy. All remaining samples (169) recorded <700 ng/g. Presumed non-pregnant individuals recorded an average of 252.8 ± 116.2 ng/g, whereas presumed pregnant individuals recorded an average of 7831.1 ± 10,542.8 ng/g. Elevated progesterone levels ranged from 1,011.11 ng/g (in September 2019 from CPAN-18) to 46,643.48 ng/g (in August 2019 from CRWA-14). High levels (39,673.46 ng/g) were found at cave CWRA-11 during the May survey (scats collected off ground, not sheets). Due to the potential for females to contaminate male scats with progesterone, we do not present the number of individuals (unique genotypes) that were detected with elevated progesterone.

Cave CWRA-16 was likely to be used by a maternity group during the 2019 breeding period as indicated by the high portion of scats containing elevated progesterone levels (70%). However, the total number of scats recorded in the cave at this time was lower than would be expected if it were to be used as maternity cave, suggesting it was only being used for short visitations or temporary diurnal roosting. Four other caves (CPAN-18, CWRA-06, CWRA-11 and CWRA-14) also contained scats with elevated progesterone, suggesting they were used by pregnant females very scarcely, and were not used as maternity caves during the course of the study.

Page | 25 Western Range Ghost Bat Scat Analysis

Table 3.5: Scats containing elevated Progesterone (P4) levels by cave

Trip 1 Trip 2 Trip 3 Trip 4 Trip 5 Percentage Total scats of scats Cave ID 11/09/2018 20/02/2019 3/05/19 23/08/19 19/09/19 - - - - - collected with 12/09/2018 26/02/2019 6/05/19 27/08/19 30/09/19 elevated P4 Mount Truchanas caves CPAN-10 - - - - 2 26 7.692 CPAN-12 - - - - - 0 0 CPAN-13 - - - - 2 20 10.00 CPAN-18 - - - - 4 13 30.769 CPAN-23 - - - - 1 4 25.00 Western Range caves CWRA-06 6 - 0 0 0 48 12.500 CWRA-11 - - 3 1 - 36 11.111 CWRA-14 - 2 - 2 - 36 11.111 CWRA-15 - - - - - 0 0 CWRA-16 - - -- - 28 40 70.000 CWRA-18 - 7 - - 20 35.000 Ratty Spring 1 - - - - 6 16.667 Cave Turee caves CTRE-03 - - - - - 0 0 CTRE-05 - - - - - 0 0 Note: ‘–‘ refers to caves where scat was not analysed from that visitation.

Page | 26 518739 532739 546739 560739 574739 588739 602739 616739

CPAN-13 7483253

Mount Trunchanas

CPAN-23 CPAN-10 CPAN-12

7469253 CPAN-18 7455253

W e s t e r n R a n g e T u r e e

7441253 Legend CTRE-05 Indivduals With High CWRA-11 CTRE-03 Progestrogene CWRA-06 0 CWRA-15 CWRA-18 PARABURDOO 1 - 2 CWRA-14 ! CWRA-16 Ratty Cave 3 - 4

5 - 7 7427253 Rio Tinto Iron Ore Western Range Ghost Bat Scat Analysis Figure 3.3: Genotyped individuals with elevated Greater Paraburdoo Progesterone (P4) levels recorded at each cave 1:350,000 Study Area Coordinate System: GDA 1994 MGA Zone 50 0 4 8 16 Projection: Transverse Mercator ¯ km Datum: GDA 1994 Size A4. Created 13/01/2020 Western Range Ghost Bat Scat Analysis

3.6 Limitations Whilst the results from the scat counts are encouraging and are currently considered the most effective method to sample and monitor Ghost Bats, the technique does have its limitations. It is unlikely bats only defecate on the sheets, and scats deposited outside of the cave or in areas inaccessible within the cave are not included in the analysis. Scats may get eaten by insects or degrade quickly (depending on diet) or get kicked onto or off sheets by other vertebrate species, such as wallabies or goannas. The varying levels of sampling effort throughout the monitoring period, did impact on the ability to compare the relative importance of caves.

Table 3.6: Study limitations and constraints

Potential limitation or Constraint Applicability to this study constraint The field personnel involved in the study have an excess of 15 years’ experience undertaking fauna surveys in the Pilbara. The team leader Experience of personnel No (Thomas Rasmussen) has specific experience surveying for Ghost Bats and scat monitoring. Proportion of fauna The study was focussed on a single species (Ghost Bat) – the species No identified was readily identified upon observation, as was their scat. Sources of information Previous Ghost Bat surveys have been conducted within the Western (recent or historic) and Partial Range area, and previous reports and resources required to complete availability of contextual the study were made available. information The study was adequately completed to meet the requirements of the Completeness No scope of works for this species. Other studies are currently being undertaken to meet similar requirements for other species. Some eastern sections of the Project Area have been cleared for drilling activities, resulting in a loss of vegetation and thus potential foraging habitat. However, these sections are comparatively small, Disturbance No indicating that such disturbances are not a significant barrier to species roosting across most of the Project Area. No recent fires occurred at Greater Paraburdoo and therefore impacts of fire on bat roosting can be excluded. The study was conducted across multiple seasons to determine cave Timing of survey, usage and movement of individuals across the Project Area. For these No weather, seasonality reasons timing was well suited to understanding the most important caves within the area. Remoteness or access The majority of the Project Area was accessible either by vehicle or on No issues foot.

Page | 28 Western Range Ghost Bat Scat Analysis

4 CONCLUSION

This survey has provided insight into the colony of Ghost Bats occurring at Western Range. Four caves were confirmed as diurnal roosts, three within Western Range (CWRA-11, CWRA-06 & CWRA-14) and one at Mount Truchanas (CPAN-12) via observation of Ghost Bats roosting. Caves CRWA-11 and CWRA-06 were of elevated significance during the study because they were used by a relatively large number of individuals (16 and 10). Additionally, CWRA-14 recorded the highest number of scats over the course of the study. These findings correspond with their previously perceived, though unproven, status as potential maternity caves. Cave CWRA-16 was likely to be used by a maternity group during the 2019 breeding period as indicated by the high portion of scats containing elevated progesterone levels (70%). However, the total number of scats recorded in the cave at this time was lower than would be expected if it were to be used as maternity cave, suggesting it was only being used for short visitations or temporary diurnal roosting. Four other caves (CPAN-18, CWRA-06, CWRA-11 and CWRA-14) also contained scats with elevated progesterone, suggesting they were used by pregnant females very scarcely, and were not used as maternity caves during the course of the study. Whilst the proportion of scats with elevated progesterone levels in these caves was low during the study (31%, 12.5%, 11% and 11%), their cave characteristics suggest they provide the appropriate conditions for a maternity roost. Based on results of this study as well as previous work completed across the area, three caves at Western Range represent possible maternity caves (CWRA-06, CWRA-11, CWRA-14), three represent diurnal roosts (CWRA-15, CWRA-16, CWRA-18) and one represents a possible diurnal roost (Ratty Spring cave). As Ghost Bats are known to move between caves, it is possible that these caves were not used as maternity roosts during the current monitoring but have been used previously.

Based on the genotyping from scat and tissue samples, 54 unique individuals were identified, with 43 of those identified from within Western Range. The genetic effective population size was estimated at 11 individuals (Ne = 11.3). This is similar to other previously colonies within the central Hamersley Ranges, e.g. West Angelas Ne = 13.9. Usage of the caves within Western Range was low with only 8 individuals observed over 35 visitations. High scat abundance at CWRA-06, CWRA-11 and CWRA-14 suggests they have one to two resident individuals, but otherwise use at the remaining caves within the Project Area is transient. Ghost Bats are known to have a number of caves that they frequently move between (Armstrong & Anstee, 2000). It is likely that the caves monitored are a part of a larger network of caves in the surrounding region. Only one individual (of 43 bats, 2% chance) was recorded at both Western Range and Mt Truchanas, a distance of over 35 km (directly) and the furthest distance travelled known from within the region. While this record demonstrates the scale of movements which can be undertaken by the species, the scarcity of more such records makes it unclear where bats are originating from and/ or travelling to, though a location other than Mt Truchanas appears likely. Given the targeted cave searching work completed in the area to date (Astron, 2013, 2018b), it appears unlikely that there remains undocumented caves within Western Range itself. A far more likely, although unproven, scenario is the unsearched ranges located ~20 - 30 km to the east of Western Range, further along Paraburdoo Range, and the range ~20 km to the west of the Project Area.

Page | 29 Western Range Ghost Bat Scat Analysis

Previous VHF tracking studies have demonstrated that the majority of individuals (though not all) forage in the plains to the north and south of Western Range, suggesting that movement of the bats is not restricted to the range itself. However, the associations between primary foraging grounds and general movement in somewhat still unknown. For example, whether long-range dispersal movements (such as that recorded between Mt Truchanas and Western Range) are undertaken more directly across foraging plains or over longer periods utilising ranges (the former comprising overnight high energy expenditure movements). Due to this unknown, development should be mindful of restricting movements along the range from neighbouring areas.

Page | 30 Western Range Ghost Bat Scat Analysis

5 REFERENCES

Armstrong, K. N., & Anstee, S. D. (2000). The Ghost Bat in the Pilbara: 100 years on. Australian Mammalogy, 22, 93-101.

Astron, Environmental Services. (2013). Western Range level 1 vegetation, flora and fauna survey. Unpublished report prepared for Rio Tinto Iron Ore.

Astron, Environmental Services. (2018a). Greater Paraburdoo Ghost Bat, Macroderma gigas - Contextual Study. December 2018. Unpublished report prepared for Rio Tinto Iron Ore.

Astron, Environmental Services. (2018b). Western Range EPA level 1 targeted conservation significant fauna assessment. Unpublished report prepared for Rio Tinto Iron Ore.

Bat Call, W.A. (2019). Paraburdoo Western Range. Acoustic Survey of Ghost Bat Activity. July 2018 to March 2019.

Bat Call, WA. (2018). Paraburdoo Western Range - Acoustic Survey of Bat and Night Parrot Activity. June 2018. Unpublished report prepared for Rio Tinto Iron Ore.

Biologic, Environmental Survey. (2014). Pilbara Regional Ghost Bat Review. Unpublished report prepared for BHP Billiton Iron Ore.

Biologic, Environmental Survey. (2015). Mining Area C - Life of Project Environmental Management Plan Revision 6: Environmental Impact Assessment of Vertebrate Fauna. Unpublished report prepared for BHP Billiton Iron Ore.

Biologic, Environmental Survey. (2016). Mining Area C - Southern Flank Environmental Impact Assessment for Ghost Bat (Macroderma gigas). Unpublished report for BHP Biliton Iron Ore.

Biologic, Environmental Survey. (2019). Warrawoona Gold Project: VHF Bat Foraging Studies. March 2019. Unpublished report for Calidus Resources Ltd.

Biologic, Environmental Survey. (2020). Memo: Western Range Ghost Bat VHF Study. Unpublished report prepared for Rio Tinto.

Biota, Environmental Sciences. (2010). Western Range Phase 1 Fauna Survey. Unpublished report prepared for Rio Tinto Iron Ore.

Boles, W. (1999). Avian prey of the Australian Ghost Bat Macroderma gigas (Microchiroptera: Megadermatidae): prey characteristics and damage from predation. Australian Zoologist, 31, 82-91.

BoM, Bureau of Meteorology. (2020). Climate Data Online. Retrieved 2020 http://www.bom.gov.au./climate/data/index.shtml

DBCA, Department of Biodiversity, Conservation and Attractions. (2018). NatureMap: Mapping Western Australia's Biodiversity (custom search). Retrieved 2018 http://naturemap.dec.wa.gov.au./default.aspx

Graham, L., Schwarzenberger, F., Möstl, E., Galama, W., & Savage, A. (2001). A versatile enzyme immunoassay for the determination of progestogens in feces and serum. Zoo Biology, 20(3), 227-236. doi:10.1002/zoo.1022

Hutson, A. M., Mickelburgh, S. P., & Racet, P. A. (2001). Microchiropteran Bats: Global Status Survey and Conservation Action Plan.

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Johnston, M., Herrod, A., Little, N., Bould, L., & Gigliotti, F. (2015). An opportunistic observation of Ghost Bat (Macroderma gigas) predation on six bird species within Karijini National Park. Journal of the Royal Society of Western Australia, 98, 89-91.

Keeley, T. (2018). West Angelas Ghost Bat Research. Unpublished report prepared for Biologic Environmental Survey.

Keeley, T., Goodrowe, K. L., Graham, L., Howell, C., & MacDonald, S. E. (2012a). The reproductive endocrinology and behavior of Vancouver Island marmot (Marmota vancouverensis). Zoo Biology, 31(3), 275-290. doi:10.1002/zoo.20384

Keeley, T., O’Brien, J. K., Fanson, B. G., Masters, K., & McGreevy, P. D. (2012b). The reproductive cycle of the Tasmanian devil (Sarcophilus harrisii) and factors associated with reproductive success in captivity. General and Comparative Endocrinology, 176(2), 182-191. doi:https://doi.org/10.1016/j.ygcen.2012.01.011

Ottewell, K., Thavornkanlapachai, R., McArthur, S., & Byrne, M. (2020). Cave use by the Ghost Bat (Macroderma gigas) in the Western Range mining precinct.

Palme, R., Touma, C., Arias, N., Dominchin, M. F., & Lepschy, M. (2013). Steroid extraction: Get the best out of faecal samples. Veterinary Medicine Austria, 100, 238-246.

Pollock, K., Boot, R., Wilson, R., Keeley, T., Grogan, K., Kennerley, P., & Johnston, S. D. (2010). Oestrus in the Julia Creek dunnart (Sminthopsis douglasi) is associated with wheel running behaviour but not necessarily changes in body weight, food consumption or pouch morphology. Animal Reproduction Science, 117, 135-146.

Tidemann, C. R., Priddel, D. M., Nelson, J. E., & Pettigrew, J. D. (1985). Foraging behaviour of the Australian Ghost Bat, Macroderma gigas (Microchiroptera: Megadermatidae). Australian Journal of Zoology, 33(5), 705-713. doi:http://dx.doi.org/10.1071/ZO9850705

Toop, J. (1985). Habitat requirements, survival strategies and ecology of the Ghost Bat Macroderma gigas dobson, (Microchiroptera, Megadermatidae) in central coastal Queensland. QPWS Rockhampton.

TSSC, Threatened Species Scientific Commitee. (2016). Conservation Advice: Macroderma gigas, Ghost Bat. Canberra, Australian Capital Territory:

Worthington-Wilmer, J., Moritz, C., Hall, L., & Toop, J. (1994). Extreme population structuring in the threatened ghost bat, Macroderma gigas: evidence from mitochondrial DNA. Proceeding of the Royal Society of London B, 257, 193-198.

Page | 32 Western Range Ghost Bat Scat Analysis

6 APPENDICES

Appendix A: Cave Assessments

Page | 33 Western Range Ghost Bat Scat Analysis

(m) (m)

Date Date

Type

Cave Cave Cave

Water Water

Shape

Aspect

or Scat or Scat

Cave ID Cave Present

Position

Estimate

Entrance Entrance Entrance Entrance Entrance Age Scat

Category

No Cham No

Exposure Scats Bat

Width Width (m)

Assessed

Height (m) Height

Number of Number

Scat Count Count Scat

Floor Slope Floor Depth Cave

Initial Ghost Ghost Initial

Coordinates in cave Bats Cham Height Cham Height

CWRA- Possible -23.190509, Mid Sheltere Round/ Fresh Taphozous 11/09/2018 Incline SW Cavity 3 1.5 25 1 8 None 3500 Estimate 06 Maternity 117.490191 Slope d Oval (<1mth) georgianus

Macroderma gigas, CWRA- Possible -23.169918, Mid Semi Horizont Fresh Taphozous 5/05/2019 Incline S Cavity 10 1 30 1 4 None 3000 Estimate 11 Maternity 117.383671 Slope Exposed al (<1mth) georgianus, Vespadelus finlaysoni

Taphozous CWRA- Possible -23.204898, Upper Sheltere Round/ Fresh georgianus, 26/08/2019 Incline S Overhang 8 5.5 50 2 3.43 None 0 Count 14 Maternity 117.526619 Slope d Oval (<1mth) Vespadelus finlaysoni

CWRA- Diurnal -23.1872, Mid Round/ Recent (1 to Taphozous N/A 21/02/2019 Incline SE Exposed Cavity 3 1.5 40 4 2.5 None 4500 Estimate 15 Roost 117.437 Slope Oval 6mths) georgianus

Page | 34

Western Range Ghost Bat Scat Analysis

(m) (m)

Date Date

Type

Cave Cave Cave

Water Water

Shape

Aspect

or Scat or Scat

Cave ID Cave Present

Position

Estimate

Entrance Entrance Entrance Entrance Entrance Age Scat

Category

No Cham No

Exposure Scats Bat

Width Width (m)

Assessed

Height (m) Height

Number of Number

Scat Count Count Scat

Floor Slope Floor Depth Cave

Initial Ghost Ghost Initial

Coordinates in cave Bats Cham Height Cham Height

Taphozous CWRA- Diurnal -23.19241, Mid Semi Round/ Fresh georgianus, 30/09/2019 Incline SE Cavity 2 3 40 1 4.5 None 500 Estimate 16 Roost 117.489115 Slope Exposed Oval (<1mth) Vespadelus finlaysoni

Taphozous CWRA- Diurnal -23.189496, Mid Round/ Fresh georgianus, 22/02/2019 Incline SE Exposed Cavern 3 2 21 2 3 None 4500 Estimate 18 Roost 117.479007 Slope Oval (<1mth) Vespadelus finlaysoni

Taphozous Ratty Possible -23.211718, Mid Sheltere Horizont Recent (1 to georgianus, Spring Diurnal 12/09/2018 Incline W Overhang 6 0.5 20 1 3 None 6 Count 117.541512 Slope d al 6mths) Vespadelus Cave Roost finlaysoni

Page | 35

Western Range Ghost Bat Scat Analysis

(m) (m)

Date Date

Type

Cave Cave Cave

Water Water

Shape

Aspect

or Scat or Scat

Cave ID Cave Present

Position

Estimate

Entrance Entrance Entrance Entrance Entrance Age Scat

Category

No Cham No

Exposure Scats Bat

Width Width (m)

Assessed

Height (m) Height

Number of Number

Scat Count Count Scat

Floor Slope Floor Depth Cave

Initial Ghost Ghost Initial

Coordinates in cave Bats Cham Height Cham Height

CTRE- Possible -23.159257, Upper Semi Horizont Intern Recent (1 to 16/05/2019 Incline SE Cavity 3.5 1.5 35 5 8 2500 Estimate None 03 Maternity 117.945688 Slope Exposed al al 6mths)

CTRE- Possible -23.150999, Upper Round/ Intern Fresh 16/05/2019 Incline SE Exposed Cavern 3.5 1.5 30 3 5 2000 Estimate None 05 Maternity 117.937757 Slope Oval al (<1mth)

Diurnal Vespadelus CPAN- Roost, -22.882782, Upper Sheltere Horizont Recent (1 to finlaysoni, 27/06/2017 Flat E Cavity 2.5 1.2 15 1 4 None 250 Estimate 10 Possible 117.694796 Slope d al 6mths) Taphozous Maternity georgianus

Page | 36

Western Range Ghost Bat Scat Analysis

(m) (m)

Date Date

Type

Cave Cave Cave

Water Water

Shape

Aspect

or Scat or Scat

Cave ID Cave Present

Position

Estimate

Entrance Entrance Entrance Entrance Entrance Age Scat

Category

No Cham No

Exposure Scats Bat

Width Width (m)

Assessed

Height (m) Height

Number of Number

Scat Count Count Scat

Floor Slope Floor Depth Cave

Initial Ghost Ghost Initial

Coordinates in cave Bats Cham Height Cham Height

Macroderma gigas, CPAN- Possible -22.882791, Upper Semi Round/ Fresh Taphozous 27/06/2017 Incline E Cavern 4 3.2 70 3 6 None 1500 Estimate 12 Maternity 117.694481 Slope Exposed Oval (<1mth) georgianus, Vespadelus finlaysoni

Taphozous CPAN- Possible -22.768701, Upper Horizont Fresh georgianus, 28/06/2017 Incline N Exposed Cavity 5 2 40 1 4 None 3000 Estimate 13 Maternity 117.57025 Slope al (<1mth) Vespadelus finlaysoni

Diurnal Taphozous CPAN- Roost, -22.900935, Mid Sheltere Horizont Fresh georgianus, 29/06/2017 Incline NW Cavity 7 3 35 1 4 None 5000 Estimate 18 Possible 117.639889 Slope d al (<1mth) Vespadelus Maternity finlaysoni

Page | 37

Western Range Ghost Bat Scat Analysis

(m) (m)

Date Date

Type

Cave Cave Cave

Water Water

Shape

Aspect

or Scat or Scat

Cave ID Cave Present

Position

Estimate

Entrance Entrance Entrance Entrance Entrance Age Scat

Category

No Cham No

Exposure Scats Bat

Width Width (m)

Assessed

Height (m) Height

Number of Number

Scat Count Count Scat

Floor Slope Floor Depth Cave

Initial Ghost Ghost Initial

Coordinates in cave Bats Cham Height Cham Height

Taphozous CPAN- Diurnal -22.886522, Mid Semi- Cavity, Round/ Recent (1 to georgianus, 30/06/2017 Flat NE 2 1 20 4 2 None 50 Estimate 23 Roost 117.620005 Slope exposed Cavern Oval 6mths) Vespadelus finlaysoni

CPAN- Night -22.827409, Lower Sheltere Horizont Taphozous 27/06/2017 Flat E Cavity 6 1.5 16 1 1.5 None 3 Count - 02 Roost 117.597228 Slope d al georgianus

CPAN- Night -22.883369, Upper Sheltere Round/ Vespadelus 27/06/2017 Incline NW Overhang 6 4 25 1 6 None 2 Count - 07 Roost 117.696068 Slope d Oval finlaysoni

Page | 38

Western Range Ghost Bat Scat Analysis

(m) (m)

Date Date

Type

Cave Cave Cave

Water Water

Shape

Aspect

or Scat or Scat

Cave ID Cave Present

Position

Estimate

Entrance Entrance Entrance Entrance Entrance Age Scat

Category

No Cham No

Exposure Scats Bat

Width Width (m)

Assessed

Height (m) Height

Number of Number

Scat Count Count Scat

Floor Slope Floor Depth Cave

Initial Ghost Ghost Initial

Coordinates in cave Bats Cham Height Cham Height

Taphozous Possible CWRA- -23.170062, Upper Sheltere Round/ georgianus, Diurnal 2/10/2018 Incline S Cavity 3 1 18 2 3 None 0 Count - 02 117.383671 Slope d Oval Vespadelus Roost finlaysoni

Taphozous Possible CWRA- -23.192695, Upper Semi Round/ georgianus, Diurnal 22/02/2019 Incline SE Cavity 1.5 1 15 1 2.5 None 0 Estimate - 17 117.488930 Slope Exposed Oval Vespadelus Roost finlaysoni

Taphozous CTRE- Night -23.151037, Upper Semi Horizont georgianus, 16/05/2019 Flat S Cavity 4 1 15 2 2 None 0 Count - 08 Roost 117.939283 Slope Exposed al Vespadelus finlaysoni

Page | 39

Western Range Ghost Bat Scat Analysis

(m) (m)

Date Date

Type

Cave Cave Cave

Water Water

Shape

Aspect

or Scat or Scat

Cave ID Cave Present

Position

Estimate

Entrance Entrance Entrance Entrance Entrance Age Scat

Category

No Cham No

Exposure Scats Bat

Width Width (m)

Assessed

Height (m) Height

Number of Number

Scat Count Count Scat

Floor Slope Floor Depth Cave

Initial Ghost Ghost Initial

Coordinates in cave Bats Cham Height Cham Height

CTRE- Night -23.150911, Upper Semi Horizont Vespadelus 16/05/2019 Flat S Cavity 3 1.2 10 2 2 None 0 Count - 07 Roost 117.938252 Slope Exposed al finlaysoni

CTRE- -23.151229, Upper Semi Horizont Nil 16/05/2019 Flat S Cavity 5 1 18 2 3 None 0 Estimate - Unknown 06 117.993746 Slope Exposed al

CTRE- Night -23.157139, Mid Sheltere Intern Old (6mths Taphozous 16/05/2019 Incline E Cavity Vertical 3 8 16 2 8 3 Count 09 Roost 117.936783 Slope d al to 3yrs) georgianus

Page | 40

Western Range Ghost Bat Scat Analysis

(m) (m)

Date Date

Type

Cave Cave Cave

Water Water

Shape

Aspect

or Scat or Scat

Cave ID Cave Present

Position

Estimate

Entrance Entrance Entrance Entrance Entrance Age Scat

Category

No Cham No

Exposure Scats Bat

Width Width (m)

Assessed

Height (m) Height

Number of Number

Scat Count Count Scat

Floor Slope Floor Depth Cave

Initial Ghost Ghost Initial

Coordinates in cave Bats Cham Height Cham Height

CTRE- Night -23.156997, Upper Declin Round/ Taphozous 16/05/2019 N Exposed Overhang 30 17 18 3 2 None 0 Count - 04 Roost 117.937827 Slope e Oval georgianus

Intern CTRE- -23.157846, Mid Sheltere Round/ al and Vespadelus Nil 14/05/2019 Incline E Cavity 1 1 10 1 3 1 Count - 02 117.936233 Slope d Oval Extern finlaysoni al

Taphozous CTRE- Night -23.150730, Upper Sheltere Round/ Intern Old (6mths georgianus, 14/05/2019 Incline S Cavity 1.2 0.8 17 2 2 20 Estimate 01 Roost 117.92670 Slope d Oval al to 3yrs) Vespadelus finlaysoni

Page | 41 Western Range Ghost Bat Scat Analysis

Appendix B: Genetic Analysis Report

Page | 42 Western Range Ghost Bat Scat Analysis

Page | 43 Western Range Ghost Bat Scat Analysis

Appendix C: Detailed Hormone Analysis Results

Page | 44 Western Range Ghost Bat Scat Analysis

Lab Extraction Progesterone Sample ID Site ID Cave Latitude Longitude sample # Weight (g) ng/g 19-0424 CWRA06.20190826.01 CWRA06 -23.19050965 117.4901912 2546 0.0255 305.88

19-0425 CWRA06.20190826.02 CWRA06 -23.19050965 117.4901912 2547 0.0097 352.99

19-0426 CWRA06.20190826.03 CWRA06 -23.19050965 117.4901912 2548 0.0230 387.83

19-0427 CWRA06.20190826.04 CWRA06 -23.19050965 117.4901912 2549 0.0242 371.90

19-0428 CWRA06.20190826.05 CWRA06 -23.19050965 117.4901912 2550 0.0216 568.52

19-0429 CWRA06.20190826.06 CWRA06 -23.19050965 117.4901912 2551 0.0265 182.64

19-0430 CWRA11.20190827.01 CWRA11 -23.16991852 117.3836713 2552 0.0114 106.67

19-0431 CWRA11.20190827.02 CWRA11 -23.16991852 117.3836713 2553 0.0202 291.09

19-0432 CWRA11.20190827.03 CWRA11 -23.16991852 117.3836713 2554 0.0257 164.98

19-0433 CWRA11.20190827.04 CWRA11 -23.16991852 117.3836713 2555 0.0097 135.26

19-0434 CWRA11.20190827.05 CWRA11 -23.16991852 117.3836713 2556 0.0241 116.18

19-0435 CWRA11.20190827.06 CWRA11 -23.16991852 117.3836713 2557 0.0104 201.54

19-0436 CWRA11.20190827.07 CWRA11 -23.16991852 117.3836713 2558 0.0088 141.82

19-0437 CWRA11.20190827.08 CWRA11 -23.16991852 117.3836713 2559 0.0244 114.75

19-0438 CWRA11.20190827.09 CWRA11 -23.16991852 117.3836713 2560 0.0121 91.24

19-0439 CWRA11.20190827.10 CWRA11 -23.16991852 117.3836713 2561 0.0227 119.82

19-0440 CWRA11.20190827.11 CWRA11 -23.16991852 117.3836713 2562 0.0118 111.19

19-0441 CWRA11.20190827.12 CWRA11 -23.16991852 117.3836713 2563 0.0242 32727.27**

19-0442 CWRA11.20190827.13 CWRA11 -23.16991852 117.3836713 2564 0.0232 415.52

19-0443 CWRA11.20190827.14 CWRA11 -23.16991852 117.3836713 2565 0.0235 178.72

19-0444 CWRA11.20190827.15 CWRA11 -23.16991852 117.3836713 2566 0.0237 99.58

19-0445 CWRA14.20190826.01 CWRA14 -23.20489875 117.5266193 2567 0.0237 133.33

19-0446 CWRA14.20190826.02 CWRA14 -23.20489875 117.5266193 2568 0.0242 310.74

19-0447 CWRA14.20190826.03 CWRA14 -23.20489875 117.5266193 2569 0.0223 278.03

19-0448 CWRA14.20190826.04 CWRA14 -23.20489875 117.5266193 2570 0.0241 222.41

19-0449 CWRA14.20190826.05 CWRA14 -23.20489875 117.5266193 2571 0.0247 262.35

19-0450 CWRA14.20190826.06 CWRA14 -23.20489875 117.5266193 2572 0.0088 456.36

19-0451 CWRA14.20190826.07 CWRA14 -23.20489875 117.5266193 2573 0.0250 180.80

19-0452 CWRA14.20190826.08 CWRA14 -23.20489875 117.5266193 2574 0.0247 192.71

19-0453 CWRA14.20190826.09 CWRA14 -23.20489875 117.5266193 2575 0.0250 212.80

19-0454 CWRA14.20190826.10 CWRA14 -23.20489875 117.5266193 2576 0.0259 154.44

19-0455 CWRA14.20190826.11 CWRA14 -23.20489875 117.5266193 2577 0.0235 302.98

19-0456 CWRA14.20190826.12 CWRA14 -23.20489875 117.5266193 2578 0.0120 282.67

19-0457 CWRA14.20190826.13 CWRA14 -23.20489875 117.5266193 2579 0.0238 250.42

19-0458 CWRA14.20190826.14 CWRA14 -23.20489875 117.5266193 2580 0.0256 26812.5**

19-0459 CWRA14.20190826.15 CWRA14 -23.20489875 117.5266193 2581 0.0230 46643.48**

19-0460 CWRA14.20190826.16 CWRA14 -23.20489875 117.5266193 2582 0.0252 344.44

19-0461 CWRA14.20190826.17 CWRA14 -23.20489875 117.5266193 2583 0.0243 164.61

Page | 45 Western Range Ghost Bat Scat Analysis

Lab Extraction Progesterone Sample ID Site ID Cave Latitude Longitude sample # Weight (g) ng/g 19-0462 CWRA14.20190826.18 CWRA14 -23.20489875 117.5266193 2584 0.0224 266.07

19-0463 CWRA14.20190826.19 CWRA14 -23.20489875 117.5266193 2585 0.0231 259.74

19-0464 CWRA14.20190826.20 CWRA14 -23.20489875 117.5266193 2586 0.0240 128.33

19-0465 CWRA18.20190222.01 CWRA18 -23.18949638 117.4790067 2587 0.0237 452.32

19-0466 CWRA18.20190222.02 CWRA18 -23.18949638 117.4790067 2588 0.0244 467.21

19-0467 CWRA18.20190222.03 CWRA18 -23.18949638 117.4790067 2589 0.0255 6274.51**

19-0468 CWRA18.20190222.04 CWRA18 -23.18949638 117.4790067 2590 0.0244 1257.38**

19-0469 CWRA18.20190222.05 CWRA18 -23.18949638 117.4790067 2591 0.0225 4933.33**

19-0470 CWRA18.20190222.06 CWRA18 -23.18949638 117.4790067 2592 0.0225 549.33

19-0471 CWRA18.20190222.07 CWRA18 -23.18949638 117.4790067 2593 0.0234 3589.74**

19-0472 CWRA18.20190222.08 CWRA18 -23.18949638 117.4790067 2594 0.0247 247.77

19-0473 CWRA18.20190222.09 CWRA18 -23.18949638 117.4790067 2595 0.0250 366.40

19-0474 CWRA18.20190222.10 CWRA18 -23.18949638 117.4790067 2596 0.0248 2862.90**

19-0475 CWRA18.20190222.11 CWRA18 -23.18949638 117.4790067 2597 0.0247 1136.84**

19-0476 CWRA18.20190222.12 CWRA18 -23.18949638 117.4790067 2598 0.0258 1289.92**

19-0477 CWRA18.20190222.13 CWRA18 -23.18949638 117.4790067 2599 0.0255 346.67

19-0478 CWRA18.20190222.14 CWRA18 -23.18949638 117.4790067 2600 0.0258 244.96

19-0479 CWRA18.20190222.15 CWRA18 -23.18949638 117.4790067 2601 0.0246 380.49

19-0480 CWRA18.20190222.16 CWRA18 -23.18949638 117.4790067 2602 0.0265 246.04

19-0481 CWRA18.20190222.17 CWRA18 -23.18949638 117.4790067 2603 0.0257 199.22

19-0482 CWRA18.20190222.18 CWRA18 -23.18949638 117.4790067 2604 0.0260 312.31

19-0483 CWRA18.20190222.19 CWRA18 -23.18949638 117.4790067 2605 0.0253 374.70

19-0484 CWRA18.20190222.20 CWRA18 -23.18949638 117.4790067 2606 0.0230 502.61

19-0485 CWRA14.20190226.01 CWRA14 -23.20489875 117.5266193 2607 0.0229 1226.20**

19-0486 CWRA14.20190226.02 CWRA14 -23.20489875 117.5266193 2608 0.0265 341.13

19-0487 CWRA14.20190226.03 CWRA14 -23.20489875 117.5266193 2609 0.0262 459.54

19-0488 CWRA14.20190226.04 CWRA14 -23.20489875 117.5266193 2610 0.0185 1236.76**

19-0489 CWRA14.20190226.05 CWRA14 -23.20489875 117.5266193 2611 0.0090 129.78

19-0490 CWRA14.20190226.06 CWRA14 -23.20489875 117.5266193 2612 0.0246 265.04

19-0492 CWRA14.20190226.08 CWRA14 -23.20489875 117.5266193 2613 0.0109 127.71

19-0493 CWRA14.20190226.09 CWRA14 -23.20489875 117.5266193 2614 0.0236 300.00

19-0494 CWRA14.20190226.10 CWRA14 -23.20489875 117.5266193 2615 0.0238 252.10

19-0495 CWRA14.20190226.11 CWRA14 -23.20489875 117.5266193 2616 0.0230 384.35

19-0496 CWRA14.20190226.12 CWRA14 -23.20489875 117.5266193 2617 0.0103 180.19

19-0497 CWRA14.20190226.13 CWRA14 -23.20489875 117.5266193 2618 0.0109 123.30

19-0498 CWRA14.20190226.14 CWRA14 -23.20489875 117.5266193 2619 0.0120 241.33

19-0499 CWRA14.20190226.15 CWRA14 -23.20489875 117.5266193 2620 0.0193 393.78

19-0500 CWRA14.20190226.16 CWRA14 -23.20489875 117.5266193 2621 0.0243 813.17*

Page | 46 Western Range Ghost Bat Scat Analysis

Lab Extraction Progesterone Sample ID Site ID Cave Latitude Longitude sample # Weight (g) ng/g 19-0501 CWRA06.20190506.01 CWRA06 -23.19050965 117.4901912 2622 0.0247 111.74

19-0502 CWRA06.20190506.02 CWRA06 -23.19050965 117.4901912 2623 0.0224 314.29

19-0503 CWRA06.20190506.03 CWRA06 -23.19050965 117.4901912 2624 0.0196 179.59

19-0504 CWRA06.20190506.04 CWRA06 -23.19050965 117.4901912 2625 0.0227 197.36

19-0505 CWRA06.20190506.05 CWRA06 -23.19050965 117.4901912 2626 0.0248 237.10

19-0506 CWRA06.20190506.06 CWRA06 -23.19050965 117.4901912 2627 0.0247 97.17

19-0507 CWRA06.20190506.07 CWRA06 -23.19050965 117.4901912 2628 0.0264 159.09

19-0508 CWRA06.20190506.08 CWRA06 -23.19050965 117.4901912 2629 0.0095 200.42

19-0509 CWRA06.20190506.09 CWRA06 -23.19050965 117.4901912 2630 0.0257 239.69

19-0510 CWRA06.20190506.10 CWRA06 -23.19050965 117.4901912 2631 0.0252 220.63

19-0511 CWRA06.20190506.11 CWRA06 -23.19050965 117.4901912 2632 0.0241 215.77

19-0512 CWRA06.20190506.12 CWRA06 -23.19050965 117.4901912 2633 0.0243 271.60

19-0513 CWRA06.20190506.13 CWRA06 -23.19050965 117.4901912 2634 0.0237 448.95

19-0514 CWRA06.20190506.14 CWRA06 -23.19050965 117.4901912 2635 0.0209 206.70

19-0515 CWRA06.20190506.15 CWRA06 -23.19050965 117.4901912 2636 0.0254 157.48

19-0516 CWRA06.20190506.16 CWRA06 -23.19050965 117.4901912 2637 0.0225 131.56

19-0517 CWRA06.20190506.17 CWRA06 -23.19050965 117.4901912 2638 0.0243 288.07

19-0518 CWRA06.20190506.18 CWRA06 -23.19050965 117.4901912 2639 0.0257 147.86

19-0519 CWRA06.20190506.19 CWRA06 -23.19050965 117.4901912 2640 0.0091 182.86

19-0520 CWRA06.20190506.20 CWRA06 -23.19050965 117.4901912 2641 0.0248 167.74

19-0521 RattyCave.20190912.1 RattyCave -23.211718 117.541512 2642 0.0240 205.00

19-0522 RattyCave.20190912.2 RattyCave -23.211718 117.541512 2643 0.0231 199.13

19-0523 RattyCave.20190912.3 RattyCave -23.211718 117.541512 2644 0.0252 1011.11**

19-0524 RattyCave.20190912.4 RattyCave -23.211718 117.541512 2645 0.0084 339.05

19-0525 RattyCave.20190912.5 RattyCave -23.211718 117.541512 2646 0.0256 201.56

19-0526 RattyCave.20190912.6 RattyCave -23.211718 117.541512 2647 0.0232 186.21

19-0527 CWRA16.20190930.01 CWRA16 -23.19241059 117.4891159 2648 0.0252 319.05

19-0528 CWRA16.20190930.02 CWRA16 -23.19241059 117.4891159 2649 0.0178 564.04

19-0529 CWRA16.20190930.03 CWRA16 -23.19241059 117.4891159 2650 0.0239 1871.13**

19-0530 CWRA16.20190930.04 CWRA16 -23.19241059 117.4891159 2651 0.0213 1318.31**

19-0531 CWRA16.20190930.05 CWRA16 -23.19241059 117.4891159 2652 0.0236 5169.49**

19-0532 CWRA16.20190930.06 CWRA16 -23.19241059 117.4891159 2653 0.0200 6850.00**

19-0533 CWRA16.20190930.07 CWRA16 -23.19241059 117.4891159 2654 0.0244 1832.79**

19-0534 CWRA16.20190930.08 CWRA16 -23.19241059 117.4891159 2655 0.0263 249.43

19-0535 CWRA16.20190930.09 CWRA16 -23.19241059 117.4891159 2656 0.0235 177.02

19-0536 CWRA16.20190930.10 CWRA16 -23.19241059 117.4891159 2657 0.0092 4782.61**

19-0537 CWRA16.20190930.11 CWRA16 -23.19241059 117.4891159 2658 0.0094 1615.32**

19-0538 CWRA16.20190930.12 CWRA16 -23.19241059 117.4891159 2659 0.0254 11181.10**

Page | 47 Western Range Ghost Bat Scat Analysis

Lab Extraction Progesterone Sample ID Site ID Cave Latitude Longitude sample # Weight (g) ng/g 19-0539 CWRA16.20190930.13 CWRA16 -23.19241059 117.4891159 2660 0.0253 18782.61**

19-0540 CWRA16.20190930.14 CWRA16 -23.19241059 117.4891159 2661 0.0108 6933.33**

19-0541 CWRA16.20190930.15 CWRA16 -23.19241059 117.4891159 2662 0.0244 19278.69**

19-0543 CWRA16.20190930.17 CWRA16 -23.19241059 117.4891159 2663 0.0252 22857.14**

19-0544 CWRA16.20190930.18 CWRA16 -23.19241059 117.4891159 2664 0.0100 17856**

19-0545 CWRA16.20190930.19 CWRA16 -23.19241059 117.4891159 2665 0.0102 429.80

19-0546 CWRA16.20190930.20 CWRA16 -23.19241059 117.4891159 2666 0.0219 1947.03**

19-0547 CWRA16.20190930.21 CWRA16 -23.19241059 117.4891159 2667 0.0094 2035.74**

19-0548 CWRA16.20190930.22 CWRA16 -23.19241059 117.4891159 2668 0.0121 309.42

19-0549 CWRA16.20190930.23 CWRA16 -23.19241059 117.4891159 2669 0.0103 1090.49**

19-0550 CWRA16.20190930.24 CWRA16 -23.19241059 117.4891159 2670 0.0118 3525.42**

19-0552 CWRA16.20190930.26 CWRA16 -23.19241059 117.4891159 2671 0.0113 532.39

19-0553 CWRA16.20190930.27 CWRA16 -23.19241059 117.4891159 2672 0.0099 9309.09**

19-0554 CWRA16.20190930.28 CWRA16 -23.19241059 117.4891159 2673 0.0114 30821.05**

19-0555 CWRA16.20190930.29 CWRA16 -23.19241059 117.4891159 2674 0.0118 9193.22**

19-0557 CWRA16.20190930.31 CWRA16 -23.19241059 117.4891159 2675 0.0250 13344.00**

19-0559 CWRA16.20190930.33 CWRA16 -23.19241059 117.4891159 2676 0.0091 3956.04**

19-0560 CWRA16.20190930.34 CWRA16 -23.19241059 117.4891159 2677 0.0225 120.89

19-0561 CWRA16.20190930.35 CWRA16 -23.19241059 117.4891159 2678 0.0240 4958.33**

19-0562 CWRA16.20190930.36 CWRA16 -23.19241059 117.4891159 2679 0.0200 1820.00**

19-0563 CWRA16.20190930.37 CWRA16 -23.19241059 117.4891159 2680 0.0231 4502.16**

19-0564 CWRA16.20190930.38 CWRA16 -23.19241059 117.4891159 2681 0.0106 1923.02**

19-0565 CWRA16.20190930.39 CWRA16 -23.19241059 117.4891159 2682 0.0255 10823.53**

19-0566 CWRA16.20190930.40 CWRA16 -23.19241059 117.4891159 2683 0.0100 4080.00**

19-0567 CPAN18.20190928.01 CPAN18 -22.90093512 117.639889 2684 0.0125 1015.04**

19-0568 CPAN18.20190928.02 CPAN18 -22.90093512 117.639889 2685 0.0244 370.49

19-0569 CPAN18.20190928.03 CPAN18 -22.90093512 117.639889 2686 0.0108 1097.78**

19-0570 CPAN18.20190928.04 CPAN18 -22.90093512 117.639889 2687 0.0233 406.87

19-0572 CPAN18.20190928.06 CPAN18 -22.90093512 117.639889 2688 0.0102 1101.18**

19-0576 CPAN18.20190928.10 CPAN18 -22.90093512 117.639889 2689 0.0109 915.96*

19-0577 CPAN18.20190928.11 CPAN18 -22.90093512 117.639889 2690 0.0113 1251.68**

19-0578 CPAN18.20190928.12 CPAN18 -22.90093512 117.639889 2691 0.0115 264.35

19-0579 CPAN18.20190928.13 CPAN18 -22.90093512 117.639889 2692 0.0093 471.40

19-0580 CPAN23.20190928.01 CPAN23 -22.88652235 117.6200053 2693 0.0093 223.66

19-0581 CPAN23.20190928.02 CPAN23 -22.88652235 117.6200053 2694 0.0114 1404.91**

19-0582 CPAN23.20190928.03 CPAN23 -22.88652235 117.6200053 2695 0.0247 98.79

19-0584 CPAN10.20190927.01 CPAN10 -22.88278253 117.6947968 2696 0.0251 364.94

19-0585 CPAN10.20190927.02 CPAN10 -22.88278253 117.6947968 2697 0.0246 447.15

Page | 48 Western Range Ghost Bat Scat Analysis

Lab Extraction Progesterone Sample ID Site ID Cave Latitude Longitude sample # Weight (g) ng/g 19-0586 CPAN10.20190927.03 CPAN10 -22.88278253 117.6947968 2698 0.0089 203.15

19-0587 CPAN10.20190927.04 CPAN10 -22.88278253 117.6947968 2699 0.0232 346.55

19-0588 CPAN10.20190927.05 CPAN10 -22.88278253 117.6947968 2700 0.0091 202.20

19-0589 CPAN10.20190927.06 CPAN10 -22.88278253 117.6947968 2701 0.0093 375.05

19-0590 CPAN10.20190927.07 CPAN10 -22.88278253 117.6947968 2702 0.0199 130.65

19-0592 CPAN10.20190927.09 CPAN10 -22.88278253 117.6947968 2703 0.0094 224.68

19-0593 CPAN10.20190927.10 CPAN10 -22.88278253 117.6947968 2704 0.0096 191.67

19-0594 CPAN10.20190927.11 CPAN10 -22.88278253 117.6947968 2705 0.0093 220.22

19-0595 CPAN10.20190927.12 CPAN10 -22.88278253 117.6947968 2706 0.0189 232.80

19-0596 CPAN10.20190927.13 CPAN10 -22.88278253 117.6947968 2707 0.0223 197.31

19-0597 CPAN10.20190927.14 CPAN10 -22.88278253 117.6947968 2708 0.0121 216.86

19-0598 CPAN10.20190927.15 CPAN10 -22.88278253 117.6947968 2709 0.0124 183.23

19-0599 CPAN10.20190927.16 CPAN10 -22.88278253 117.6947968 2710 0.0085 1370.35**

19-0600 CPAN10.20190927.17 CPAN10 -22.88278253 117.6947968 2711 0.0100 369.60

19-0601 CPAN10.20190927.18 CPAN10 -22.88278253 117.6947968 2712 Not enough

19-0602 CPAN10.20190927.19 CPAN10 -22.88278253 117.6947968 2713 0.0095 192.00

19-0603 CPAN10.20190927.20 CPAN10 -22.88278253 117.6947968 2714 0.0098 210.61

19-0604 CPAN10.20190927.21 CPAN10 -22.88278253 117.6947968 2715 0.0247 234.82

19-0605 CPAN10.20190927.22 CPAN10 -22.88278253 117.6947968 2716 0.0084 1683.81**

19-0606 CPAN10.20190927.23 CPAN10 -22.88278253 117.6947968 2717 0.0126 248.89

19-0607 CPAN10.20190927.24 CPAN10 -22.88278253 117.6947968 2718 0.0180 93.33

19-0608 CPAN10.20190927.25 CPAN10 -22.88278253 117.6947968 2719 Not enough

19-0609 CPAN10.20190927.26 CPAN10 -22.88278253 117.6947968 2720 0.0105 178.29

19-0610 CPAN13.20190928.01 CPAN13 -22.7687006 117.5702501 2721 0.0089 188.76

19-0612 CPAN13.20190928.03 CPAN13 -22.7687006 117.5702501 2722 0.0105 269.71

19-0613 CPAN13.20190928.04 CPAN13 -22.7687006 117.5702501 2723 0.0202 247.52

19-0615 CPAN13.20190928.06 CPAN13 -22.7687006 117.5702501 2724 0.0095 631.58

19-0616 CPAN13.20190928.07 CPAN13 -22.7687006 117.5702501 2725 0.0218 258.72

19-0617 CPAN13.20190928.08 CPAN13 -22.7687006 117.5702501 2726 0.0075 177.07

19-0618 CPAN13.20190928.09 CPAN13 -22.7687006 117.5702501 2727 0.0118 115.25

19-0620 CPAN13.20190928.11 CPAN13 -22.7687006 117.5702501 2728 0.0112 6514.29**

19-0621 CPAN13.20190928.12 CPAN13 -22.7687006 117.5702501 2729 0.0124 249.03

19-0622 CPAN13.20190928.13 CPAN13 -22.7687006 117.5702501 2730 0.0214 185.05

19-0623 CPAN13.20190928.14 CPAN13 -22.7687006 117.5702501 2731 0.0108 115.56

19-0624 CPAN13.20190928.15 CPAN13 -22.7687006 117.5702501 2732 0.0097 192.99

19-0625 CPAN13.20190928.16 CPAN13 -22.7687006 117.5702501 2733 0.0191 190.58

19-0626 CPAN13.20190928.17 CPAN13 -22.7687006 117.5702501 2734 0.0090 8960.00**

19-0627 CPAN13.20190928.18 CPAN13 -22.7687006 117.5702501 2735 0.0109 171.74

Page | 49 Western Range Ghost Bat Scat Analysis

Lab Extraction Progesterone Sample ID Site ID Cave Latitude Longitude sample # Weight (g) ng/g 19-0628 CPAN13.20190928.19 CPAN13 -22.7687006 117.5702501 2736 0.0109 233.39

19-0631 CWRA11.20190505.02 CWRA11 -23.16991852 117.3836713 2737 0.0238 142.86

19-0632 CWRA11.20190505.03 CWRA11 -23.16991852 117.3836713 2738 0.0246 169.11

19-0633 CWRA11.20190505.04 CWRA11 -23.16991852 117.3836713 2739 0.0260 169.23

19-0634 CWRA11.20190505.05 CWRA11 -23.16991852 117.3836713 2740 0.0250 182.40

19-0635 CWRA11.20190505.06 CWRA11 -23.16991852 117.3836713 2741 0.0091 530.99

19-0636 CWRA11.20190505.07 CWRA11 -23.16991852 117.3836713 2742 0.0240 240.00

19-0637 CWRA11.20190505.08 CWRA11 -23.16991852 117.3836713 2743 0.0231 93.51

19-0638 CWRA11.20190505.09 CWRA11 -23.16991852 117.3836713 2744 0.0232 351.72

19-0639 CWRA11.20190505.10 CWRA11 -23.16991852 117.3836713 2745 0.0240 366.67

19-0640 CWRA11.20190505.11 CWRA11 -23.16991852 117.3836713 2746 0.0217 173.27

19-0641 CWRA11.20190505.12 CWRA11 -23.16991852 117.3836713 2747 0.0249 131.73

19-0642 CWRA11.20190505.13 CWRA11 -23.16991852 117.3836713 2748 0.0238 104.20

19-0643 CWRA11.20190505.14 CWRA11 -23.16991852 117.3836713 2749 0.0245 39673.47**

19-0644 CWRA11.20190505.15 CWRA11 -23.16991852 117.3836713 2750 0.0244 245.90

19-0645 CWRA11.20190505.16 CWRA11 -23.16991852 117.3836713 2751 0.0246 131.71

19-0646 CWRA11.20190505.17 CWRA11 -23.16991852 117.3836713 2752 0.0260 1980.00**

19-0647 CWRA11.20190505.18 CWRA11 -23.16991852 117.3836713 2753 0.0225 4666.67**

19-0648 CWRA11.20190505.19 CWRA11 -23.16991852 117.3836713 2754 0.0105 396.19

19-0649 CWRA11.20190505.20 CWRA11 -23.16991852 117.3836713 2755 0.0190 210.53

19-0650 CWRA11.20190505.21 CWRA11 -23.16991852 117.3836713 2756 0.0105 99.05

19-0651 CWRA06.20180911.01 CWRA06 -23.19050965 117.4901912 2757 0.0241 1661.41**

19-0652 CWRA06.20180911.02 CWRA06 -23.19050965 117.4901912 2758 0.0252 315.87

19-0653 CWRA06.20180911.03 CWRA06 -23.19050965 117.4901912 2759 0.0116 29875.86**

19-0654 CWRA06.20180911.04 CWRA06 -23.19050965 117.4901912 2760 0.0262 1131.30**

19-0655 CWRA06.20180911.05 CWRA06 -23.19050965 117.4901912 2761 0.0238 1157.98**

19-0656 CWRA06.20180911.06 CWRA06 -23.19050965 117.4901912 2762 0.0258 1209.30**

19-0657 CWRA06.20180911.07 CWRA06 -23.19050965 117.4901912 2763 0.0258 139.53

19-0658 CWRA06.20180911.08 CWRA06 -23.19050965 117.4901912 2764 0.0247 217.00

19-0659 CWRA06.20180911.09 CWRA06 -23.19050965 117.4901912 2765 0.0268 197.01

19-0660 CWRA06.20180911.10 CWRA06 -23.19050965 117.4901912 2766 0.0243 348.97

19-0661 CWRA06.20180911.11 CWRA06 -23.19050965 117.4901912 2767 0.0268 380.60

19-0662 CWRA06.20180911.12 CWRA06 -23.19050965 117.4901912 2768 0.0121 317.36

19-0663 CWRA06.20180911.13 CWRA06 -23.19050965 117.4901912 2769 0.0235 418.72

19-0664 CWRA06.20180911.14 CWRA06 -23.19050965 117.4901912 2770 0.0247 1619.43**

19-0665 CWRA06.20180911.15 CWRA06 -23.19050965 117.4901912 2771 0.0114 329.82

19-0666 CWRA06.20180911.16 CWRA06 -23.19050965 117.4901912 2772 0.0259 176.06

19-0667 CWRA06.20180911.17 CWRA06 -23.19050965 117.4901912 2773 0.0256 146.88

Page | 50 Western Range Ghost Bat Scat Analysis

Lab Extraction Progesterone Sample ID Site ID Cave Latitude Longitude sample # Weight (g) ng/g 19-0668 CWRA06.20180911.18 CWRA06 -23.19050965 117.4901912 2774 0.0238 235.29

19-0669 CWRA06.20180911.19 CWRA06 -23.19050965 117.4901912 2775 0.0206 221.36

19-0670 CWRA06.20180911.20 CWRA06 -23.19050965 117.4901912 2776 0.0235 125.96

19-0672 CWRA06.20190929.02 CWRA06 -23.19050965 117.4901912 2777 0.0251 521.12 1 ** – elevated progesterone levels (>970 ng/g); * – intermediate progesterone levels (700–950 ng/g)

Page | 51