Ecological Impacts of Invasive Carp in Australian Dryland Rivers

Marshall Jonathan (Orcid ID: 0000-0002-7177-4543) Negus Peter (Orcid ID: 0000-0003-2680-2573)

Ecological impacts of invasive carp in Australian dryland rivers

Jonathan C. Marshall1,2

Joanna J. Blessing1

Sara E. Clifford1

Kate M. Hodges1,3

Peter M. Negus1,4

Alisha L. Steward1,2

1Department of Environment and Science, Queensland Government, Brisbane, Australia

2Australian Rivers Institute, Griffith University, Nathan, Queensland, Australia

3Institute for Applied Ecology, University of Canberra, Canberra, ACT, Australia

4School of Earth and Environmental Sciences, University of Queensland, Brisbane, Queensland, Australia

Correspondence: Jonathan C. Marshall, Department of Environment and Science, Queensland Government, GPO Box 2454, Brisbane, Queensland 4001, Australia

Abstract

1. Invasive carp are widely reported to harm ecosystems. In Australia, carp are a serious pest, and consequently investigations of biocontrol options are under way. 2. Best practice biocontrol requires cost/risk:benefit evaluation. To assist this, the impacts of carp on aquatic ecosystems have been summarized. 3. To aid the evaluation of benefits, general predictions were tested by comparing dryland river ecosystems with and without carp, and ecosystem responses to a gradient in local carp density.

This is the author manuscript accepted for publication and has undergone full peer review but has not been through the copyediting, typesetting, pagination and proofreading process, which may lead to differences between this version and the Version of Record. Please cite this article as doi: 10.1002/aqc.3206

This article is protected by copyright. All rights reserved. 4. Expectations were that in the presence of carp, and with increasing density, there would be increasing turbidity, decreasing densities of macrophytes and macroinvertebrates and associated changes in assemblage composition, resulting in decreasing native fish density. 5. Not all expected responses were found, indicating that the general understanding of carp impact requires modification for dryland rivers. Notably, carp did not increase turbidity or reduce macroinvertebrate density or composition, probably because of key attributes of dryland rivers. In contrast, there were large impacts to native fish biomass, not from the mechanisms expected, but from food resource monopolization by carp. Macrophyte occurrence was reduced, but macrophytes are naturally rare in these rivers. It is likely that the extirpation of an endangered river snail resulted from carp predation. 6. Impacts on native fish may be reversible by carp control, but reversal of impacts on the snail may require carp elimination and snail reintroduction. Modelling is necessary to predict the probability of beneficial versus undesirable outcomes from carp control, and complementary measures to control other stressors may be needed. 7. Benefits of carp control on dryland river ecosystems are fewer than generally predicted. This reinforces the point that ecological understanding cannot always be transferred between diverse settings, and highlights the need to understand system characteristics relevant to causal impact pathways when applying generic carp impact models to specific settings. This has global relevance to future carp control efforts.

Key words

alien fish, biocontrol, macroinvertebrates, macrophytes, molluscivory, Murray-Darling Basin, National Carp Control Plan, native fish biomass, Notopala sublineata, turbidity

1. Introduction From their origin in Eurasia, carp (Cyprinus carpio) have been introduced to waterways in many parts of the world. This has often been deliberate, dating back at least to the era of the Roman Empire, as they are an important food and aquaculture species and are highly valued for recreational fishing (Vilizzi, 2012). Carp have also been widely reported to cause ecological harm where they have been introduced (Vilizzi, Tarkan, & Copp, 2015). This has generated tension between conservation

This article is protected by copyright. All rights reserved. interests, who lobby for carp control for ecological benefit (Weber & Brown, 2009), and fishing interests, who advocate maintaining or boosting carp stocks (Carpworld Magazine, 2019).

Contrasting attitudes towards carp in the United Kingdom (UK) exemplifies such tensions. Carp were introduced to the UK several hundred years ago, where they are recognized to cause environmental problems such as elevated nutrient concentrations, algal biomass and turbidity, and reduced abundances of native fish (Jackson, Quist, Downing, & Larscheid, 2010). These are impacts that should be managed under the requirements of the European Water Framework Directive (Council of the European Communities, 2000) in order to achieve ‘good ecological status’, yet many UK lakes are still heavily stocked with carp and managed for recreational carp fishing (Britton, Gozlan, & Copp, 2011; Hewlett, Snow, & Britton, 2009). Likewise, conservation efforts to increase populations of Eurasian otter (Lutra lutra) in the UK have created conflict, with carp fisherman believing otters feed on stocked carp and calling for culls of otters in important carp fishing regions (Almeida et al., 2012).

In Australia, carp are widespread and abundant in the waterways, impoundments, wetlands and lakes of the Murray-Darling Basin and in some other river systems (Koehn, 2004). First introduced in the mid-1800s, they only became invasive following the escape of the Boolarra carp strain from captivity in the 1960s. They have since spread widely following extensive river flooding in exceptionally wet years during the 1970s, to reach the current major extent of their distribution by the latter few decades of the twentieth century. Today, carp dominate many aquatic systems in the Murray-Darling Basin, constituting up to nearly 80% of the total fish biomass (Koehn, 2004). Although, as elsewhere, carp were introduced to Australian waters as a potential new human food source (Koehn, Brumley, & Gehrke, 2000), they are not generally accepted here as a desirable food or as a worthy angling species. Rather, they are broadly derided as an invasive pest requiring control or elimination (Koehn et al., 2000).

Native fishes in the Murray-Darling Basin have been subjected to numerous stressors since European settlement, including river regulation and water diversion, longitudinal and lateral fragmentation (Baumgartner, Zampatti, Jones, Stuart, & Mallen-Cooper, 2014), habitat degradation, and the introduction of alien species (Wilson, 2005). Many native species have undergone reductions in range and population size throughout the Basin, with the Yarra pygmy perch (Nannoperca obscura) recently identified as possibly the first fish extinction for the catchment (ABC News, 2019). Basin-

This article is protected by copyright. All rights reserved. scale restoration measures have been implemented or are planned, among other things, to restore native fish populations. These measures include environmental flow restoration by water recovery (Murray Darling Basin Authority, 2012) and measures to provide enhanced fish passage past dams and weirs (Barrett & Mallen-Cooper, 2006). Carp are considered to be a major remaining threat to native fish in the region (Koehn et al., 2000).

This perception led the Australian Government to develop a National Carp Control Plan (NCCP) to reduce carp abundance in order to help restore the health of Australian waterways and aquatic biodiversity, with a focus on native fish recovery (NCCP, 2018). The principal approach currently under consideration by the NCCP is biocontrol using cyprinid herpesvirus 3 (CyHV-3). Although various authors have raised doubts and uncertainties concerning the likely efficacy, safety and risks of unintended perverse outcomes from potential biocontrol of Australian carp with CyHV-3 (Becker, Ward, & Hick, 2018; Kopf et al., 2017; Lighten & Van Oosterhout, 2017; Marshall, Davison, Kopf, Boutier, & Vanderplasschen, 2018; Paton & McGinness, 2018; Thresher, Allman, & Stremick- Thompson, 2018), it remains important to this planning process to quantify the ecological benefits that successful carp control could achieve. Such information provides the denominator in a cost/risk: benefit evaluation at the core of best-practice biocontrol evaluation (Kopf et al., 2017) and is thus fundamental to the NCCP and its eventual recommendations to government.

The benefits of successful carp biocontrol are considered here in two stages. First, aspects of the ecological harm carp cause at present are quantified. Second, the reversibility of this harm is considered if carp abundances were to be successfully reduced. This is addressed in the setting of the rivers of the northern part of Australia’s Murray-Darling Basin, that are often described as dryland rivers because much of their length flows through arid and semi-arid landscapes and they experience large-scale nett evaporative water losses (Balcombe et al., 2010). Such rivers offer several practical advantages for this study. They are characterized by highly intermittent flow regimes, with short duration flow pulses interspersed with long periods of no discharge, during which obligate aquatic organisms are restricted to remnant, persistent, in-channel waterholes. Isolation of the aquatic system into waterholes as discrete habitat patches during dry phases means that local impacts of carp are likely to predominate over regional impacts (Balcombe et al., 2006), making this an ideal environment for testing ecological responses to variable local carp abundance. Furthermore, adjacent to the carp-infested rivers of the Basin are two very similar rivers (in terms of

This article is protected by copyright. All rights reserved. hydrology, geomorphology and landscape setting) in which carp have not become established, providing the ideal setting for comparisons of similar ecosystems with and without carp at the catchment scale. Land use in the river catchments in this investigation is dominated by low-intensity grazing in relatively natural vegetation (QLUMP, 2013); otherwise, they are comparatively undeveloped, with no major population centres and little to no intensive agriculture or industry (Negus, Blessing, Clifford, & Steward, 2015). As such, they lack many of the additional stressors that compromise investigations of the impacts of a single stressor, such as carp impacts, in complex multi-stressor systems (Bajer et al., 2016). Dryland rivers of the type investigated here constitute approximately 40% of the catchment area of the Murray-Darling Basin (CSIRO, 2008), so they have wide relevance to Australian carp control.

The impacts of carp on aquatic ecosystems have been broadly reported and reviewed (Vilizzi et al., 2015; Weber & Brown, 2009) and are synthesized here into testable hypotheses. The main mechanisms by which carp perpetrate ecological impacts are by their feeding behaviour and by excreting nutrients. Excretion is hypothesized to result in increased turbidity from increased phytoplankton biomass. Feeding, and to a lesser extent spawning, are expected also to increase turbidity by mobilizing fine bed sediments leading to increased suspended sediment concentrations. Feeding is also predicted to reduce aquatic macrophyte and macroinvertebrate density – directly by consumption and disturbance, as well as indirectly by increased turbidity leading to increased incident light attenuation and less light availability for aquatic primary production. Although carp feeding is expected to have some direct adverse influence on native fish density, the indirect impacts of carp are predicted to be greater through reduced habitat availability (macrophytes), less food (macroinvertebrates and possibly juvenile native fish) and through impacts from elevated turbidity. Increased turbidity and reduced densities of aquatic macrophytes and macroinvertebrates are similarly predicted to trigger cascading impacts to other elements of the ecosystem including amphibians and birds. With regard to food availability for native fish, Kopf et al. (2019) predicted by modelling that food webs invaded by carp will have lower standing biomass of native species, when compared with systems without carp, but they also cautioned that carp impacts on native species via food web and ecosystem energetics remain poorly understood.

Thus, turbidity, macrophytes and macroinvertebrates are the key nodes in the conceptual impact pathways that carp are expected to generate, and native fish are a key endpoint for cascading

This article is protected by copyright. All rights reserved. ecosystem impacts from carp. These are the ecological attributes considered here in relation to carp presence, abundance and biomass in isolated, dryland, riverine waterholes. The objectives of this study were to test in a dryland river setting the expectations based on the above synthesis of carp impacts from the literature that in the presence of carp, and with increasing carp abundance and biomass, there would be: (a) increasing turbidity; (b) decreasing densities of macrophytes and macroinvertebrates with associated changes in their assemblage compositions; and (c) decreasing native fish density.

2. Methods

2.1 Study sites

Carp impacts on waterhole ecosystems were investigated at sites located within the Australian state of Queensland in the dryland river catchments of Ambathala Creek and the Bulloo, Paroo, Warrego and Nebine Rivers (Figure 1). The landscape in the region largely comprises sparsely vegetated, level plains with low gradients and often extensive floodplains (Balcombe et al., 2006; Power, Biggs, & Burton, 2007; Thoms, 2003). The soils are mostly ancient and highly weathered with high fine clay content and high sodium concentration, which increases their dispersability (Kirk, 1985; Power et al., 2007). This results in perpetually high turbidity and rivers that appear ‘muddy’. The climate is semi- arid with summer-dominated rainfall and high inter-annual variability. These river systems flow intermittently, (Kennard et al., 2010), often existing as a series of hydrologically disconnected pools known as waterholes (Sheldon et al., 2010).

The Paroo, Warrego and Nebine catchments are part of the northernmost drainages of Australia’s largest river system – the Murray-Darling Basin (Figure 1). Carp from the Boolarra strain, which escaped containment into the Murray River in the state of Victoria in the 1960s, reached these northern extremes of the Basin probably by the late 1970s and definitely by the 1990s, and carp were declared noxious in the Australian state of Queensland by 1994 (Koehn et al., 2000). The Ambathala and Bulloo catchments are isolated endorheic drainages that flow into terminal lakes. Because of their hydrological separation from the Murray-Darling Basin, carp have not become established in these two catchments. Although Ambathala Creek is often considered as part of the Warrego catchment for management purposes (e.g. DSITIA, 2013), it is hydrologically isolated from

This article is protected by copyright. All rights reserved. the Warrego River and its tributaries (Power et al., 2007), so it is not functionally part of the catchment or the broader Murray-Darling Basin.

Sample sites were selected from a data-frame of the 579 previously mapped permanent and near- permanent riverine waterholes in the region (Negus et al., 2015; Silcock, 2009) to maximize the spread of sites within and between catchments (Dobbie & Negus, 2013). Thirty-one sites were sampled: one in Ambathala Creek, six in the Bulloo catchment, eight in the Paroo, ten in the Warrego, and six in the Nebine catchment (Figures 2 and 3). Thus, there were seven sites in two catchments where carp were absent and twenty-four sites in three catchments with carp present (Figure 1).

Sites were sampled during austral spring between September and November 2012 with equivalent antecedent hydrological conditions at all sites – characterized by at least 6 months without any flow, other than very occasional and minor local events of magnitudes too small to instigate fish movement between waterholes (Marshall et al., 2016) (Supporting Information Figure S1). All the sampling protocols and procedures used were ethically reviewed and approved and all necessary permits were current.

2.2 Sampling methods

2.2.1 Carp and other fishes

The abundance and biomass of carp and other fish species were estimated following the fyke-net sampling method of Arthington, Balcombe, Wilson, Thoms, & Marshall (2005) with minor modifications. Three fyke nets were set parallel to the bank in 1–2 m water depth and with wings 10 m apart at each site: two facing downstream, and one facing upstream, with the two downstream-facing nets at least 100 m apart to avoid catch interference between nets. Waterholes at some sites were too small to fit three nets and only two nets were set, one facing in each direction. Cod-ends were secured out of the water to ensure that trapped air- breathing animals would not drown. Nets were set in the afternoon and retrieved the following morning. The number of nets set and the sampling duration of each net were recorded for subsequent calculation of catch per unit effort (CPUE). Each fish caught was identified to species and weighed to the nearest 0.1 g. Site abundance and site biomass of each species were expressed as CPUE on a per fyke-net basis, standardized by sampling

This article is protected by copyright. All rights reserved. duration to 19 hours (Arthington et al., 2005). At a few sites carp were observed but not caught in fyke nets, and for these sites carp abundance and biomass were assumed to be equivalent to the minimum values across all other sites at which carp were caught in the same catchment. If carp were neither caught nor observed, values of zero were assigned both for site abundance and site biomass. Native fish and total fish abundance and biomass were calculated as the summed CPUE values of all native fishes and all fishes at each site.

2.2.2 Aquatic macroinvertebrates

Two samples of aquatic macroinvertebrate assemblages were collected from each site from the littoral zone of waterholes using the Australian River Assessment System (AUSRIVAS) rapid assessment protocol (Queensland Department of Natural Resources and Mines, 2001), which involves kick samples with a 250 µm nylon mesh D-net over 10 m with live picking for 1 h by accredited operators and with application of a field collection quality assurance protocol (Negus et al., 2015). Assemblage data generated by the live-picking protocol emphasizes representativeness of composition over abundance estimation and is thus semi-quantitative with no more than 10 individuals of any one taxon collected by live picking for identification and enumeration. Samples were identified and counted in the laboratory under stereo dissecting microscopes to family level of taxonomic resolution in most instances, with the exceptions that Chironomidae were identified to subfamily, and Acarina, Cladocera, Copepoda, Oligochaeta and Ostracoda were not resolved further. The resulting data were quality assured for laboratory identification and enumeration accuracy (Negus et al., 2015). These levels of taxonomic resolution and sample quantification have been shown to accurately represent assemblage differences between sites based on species and abundance level data in Queensland dryland rivers (Marshall, Steward, & Harch, 2006).

Counts for replicate samples at each site were averaged for analysis. Rare taxa, comprising less than 5% of the global total abundance across all sites (globally rare) and less than 5% of all individual site total abundances (locally rare), were removed because such rare taxa are inadequately sampled and thus add unwanted noise to subsequent analyses (Clarke & Warwick, 2001).

Following removal of rare taxa, macroinvertebrate richness was calculated as the count of all taxa recorded at a site. Macroinvertebrate abundance could not be calculated in the normal way owing to the truncation of taxon enumeration to a maximum of 10. An index of site abundance (henceforth

This article is protected by copyright. All rights reserved. referred to as ‘abundance’) was calculated to allow for the semi-quantitative nature of the data, as the count of taxa per site with truncated mean abundance greater than 9.5.

2.2.3 Notopala sublineata

Because of the benthic feeding habits of carp, certain benthic macroinvertebrate taxa are particularly vulnerable to impacts of direct consumption (Ip et al., 2014). In the study area, the distribution of the river snail Notopala sublineata (Gastropoda: Viviparidae), which was once common and widely distributed throughout the Murray-Darling Basin, has undergone a dramatic decline since the 1960s, such that by the 1980s populations were thought to remain in only a small number of locations. The species is at present considered extinct throughout its natural range in the state of New South Wales and it is listed as endangered on the IUCN red list of threatened species (New South Wales Department of Primary Industries, 2007; Ponder, 1996). The suggestion has been made that, at least in part, this decline has been due to carp predation (New South Wales Department of Primary Industries, 2007). Previous Queensland Government sampling found N. sublineata to be abundant still in the Bulloo River and Cooper Creek systems where carp are absent, but it was seldom detected at sites in the Murray-Darling Basin where carp are present (Queensland Government, unpublished data). Therefore, targeted sampling was undertaken to detect the presence of N. sublineata at each site in addition to sampling the general macroinvertebrate assemblage. This involved focused searching for aquatic snails by two people for 15 min in likely N. sublineata habitats (wood, leaf packs and root masses at the edges of waterholes) using hands, a sieve and D-net (Figure 3). Results were recorded as presence or absence of N. sublineata at each site.

2.2.4 Macrophytes

A standard set of digital photos was taken from a randomly chosen location on the edge of the waterhole at each site. Site photos covered views upstream, downstream and perpendicular to the bank from the photography point. Photos were systematically reviewed back in the laboratory for the presence and density of aquatic macrophytes of different growth forms (submerged, floating, and emergent). Presence was noted and density scored for each taxon present (Stephens & Dowling, 2002) using a categorical scale: absent (< 1% cover), isolated (1–5% cover), scattered (6–20% cover),

This article is protected by copyright. All rights reserved. beds/stands (21–50%), overgrown/filling channel (51–100%). Photos were independently evaluated by two operators with results standardized on a per site basis.

2.2.5 Turbidity and water quality

Turbidity (NTU), electrical conductivity (EC), pH and water temperature (oC) were recorded in-situ at each site using a multi-parameter probe (Horiba Water Quality Checker U-10) calibrated according to the manufacturer’s instructions.

2.3 Analysis methods

2.3.1 Ecosystem response to carp

The overall approach to testing hypotheses concerning ecosystem responses to carp was two-fold. First, tests were made for differences in ecosystem responses between sites from catchments with carp present (Paroo, Warrego, Nebine) or absent (Ambathala, Bulloo). Second, tests were conducted on the extent to which carp density measures explain variability in ecosystem response measures between sites in the three Murray-Darling Basin catchments with carp present. All analyses were undertaken in R (R Core Team, 2017), with details of the R packages used given below.

2.3.2 Aquatic macroinvertebrate responses to carp

To evaluate whether carp presence at the catchment scale reduced macroinvertebrate richness or abundance, one-sided t-tests were used to test for differences in these metrics between sites from catchments with carp present or absent.

Generalized linear models with Poisson distributions were used to test for variables explaining the variability of macroinvertebrate richness and abundance at sites in the three Murray-Darling Basin catchments with carp present. The variables included carp biomass and abundance, site location (latitude, longitude, catchment) and water quality (EC, pH, turbidity, water temperature). Variable deletion was undertaken to identify the most parsimonious (‘best’) of the significant (P < 0.05) models based on minimizing the Akaike Information Criterion (AIC). A visual assessment of the homogeneity of variance (residual vs fitted plot) and normality (normal Q-Q plot) showed that the assumptions of linear models were adequately met.

This article is protected by copyright. All rights reserved. To test whether macroinvertebrate assemblage composition differed between sites from catchments with carp present or absent, permutational multivariate analysis of variance was used applying distance matrices in the adonis2 routine in the vegan Package (Oksanen et al., 2018), with Bray-Curtis dissimilarity and 999 permutations. Non-metric multidimensional scaling ordination (nMDS) was conducted to illustrate differences in macroinvertebrate assemblage composition between sites from rivers with and without carp. This was plotted with sites categorized by river catchment and the presence or absence of carp in those catchments.

Macroinvertebrate taxa contributing to significant differences between sites from catchments with carp present or absent were identified by indicator species analysis (Dufrene & Legendre, 1997) using the indval routine in the labdsv Package (Roberts, 2016).

In order to better contextualize the magnitude and nature of any macroinvertebrate assemblage differences found between catchments with and without carp, analyses were undertaken of the macroinvertebrate data from only those catchments with carp present (Paroo, Warrego and Nebine). The permutational multivariate analysis of variance using distance matrices described above was repeated, but this time the test was for differences among the three catchments. Ordination by nMDS was repeated on this dataset to illustrate differences between rivers. Environmental variables were fitted as vectors or factors onto the ordination using the envfit routine in the vegan package (Oksanen et al., 2018), effectively assessing which environmental variables are most correlated with macroinvertebrate assemblage variability. The goodness of fit and significance of fitted vectors or factors was assessed using 1000 permutations of environmental variables. The nMDS was also displayed as a bubble plot with the size of symbols representing sites scaled first by the biomass and then by the abundance of carp at these sites.

A BIOENV analysis was conducted in the vegan Package (Oksanen et al., 2018), to identify the best sub-set of environmental variables to explain patterns in macroinvertebrate assemblages, as represented by the site Bray-Curtis distance matrix, from sites in the three catchments with carp present. The Gower distance measure was used for environmental variables so that both continuous and categorical variables could be included. The magnitude and significance of the best multivariate correlation from the BIOENV results was then tested with a Mantel’s test. The role of carp

This article is protected by copyright. All rights reserved. abundance and biomass in explaining macroinvertebrate assemblage patterns was evaluated by reviewing the role of these variables in adding explanatory power in the BIOENV results.

2.3.3. N. sublineata responses to carp

To evaluate whether carp presence at the catchment scale explained patterns in the occurrence of N. sublineata, one-sided t-tests were used to test for differences in these metrics between sites from catchments with carp present or absent.

2.3.4 Macrophyte responses to carp

A one-sided Fishers exact probability test was used to test whether the observed proportion of sites with macrophytes present was greater in catchments without carp than in catchments with carp.

2.3.5 Turbidity responses to carp

A two sample (one-sided) t-test was used to test the hypothesis that turbidity is higher at sites in catchments with carp than in those without carp. A generalized linear model with normal distribution was then used to test for variables explaining turbidity at sites in catchments with carp present. The variables included carp biomass and abundance, site location (latitude, longitude, catchment) and other water quality attributes (EC, pH, water temperature). Variable deletion was undertaken to identify the most parsimonious of the significant models (P < 0.05) based on minimizing AIC. A visual assessment of the homogeneity of variance (residual vs fitted plot) and normality (normal Q-Q plot) showed that the assumptions of linear models were adequately met.

2.3.6 Native fish responses to carp

Analyses of carp impacts on native fish density were undertaken on summary metrics of site assemblages, but not on assemblage composition differences between sites. The latter were not considered because, although there is considerable overlap in catchment species pools between the Murray-Darling Basin and the Bulloo, they are not identical (Marshall, Prior, Steward, & McGregor, 2006; Unmack, 2001).

This article is protected by copyright. All rights reserved. To evaluate whether carp presence at the catchment scale reduced native fish biomass or abundance, one-sided t-tests were used to test for differences in these metrics between sites from catchments with carp present or absent. Preliminary investigation suggested that there was also a difference associated with carp presence-absence in the variability between sites in native fish biomass and abundance. This was evaluated using one-sided F tests of variances.

Generalized linear models with Poisson distributions were used to test for variables explaining the variability in native fish biomass and abundance at sites in the three Murray-Darling Basin catchments with carp present. Variables included carp biomass and abundance, site location (latitude, longitude, catchment) and water quality (EC, pH, turbidity, water temperature). Variable deletion was undertaken to identify the most parsimonious of the significant (P < 0.05) models based on minimizing AIC. A visual assessment of the homogeneity of variance (residual vs fitted plot) and normality (normal Q-Q plot) showed that the assumptions of linear models were adequately met. Partial correlation plots (term plots) of the relationships between variables included in the best models and native fish metrics were generated to aid interpretation of significant regression results.

3. Results

3.1 Carp CPUE

As expected, carp were absent from all samples at sites in the Ambathala Creek and Bulloo River catchments, and neither were carp observed to be present at any of the sites in these two catchments. Within the three Murray-Darling Basin catchments, carp were caught at 19 of the 24 sites, and were observed to be present at three of the remaining five sites (Table 1). Thus, the planned comparisons of ecosystem response to the presence or absence of carp at catchment scales were vindicated. CPUE and biomass of carp caught across Murray-Darling Basin sites ranged from 0– 100% of the total site fish assemblages in the standardized fyke net catches. Carp averaged 60%, 51% and 37% of the total fish biomass in samples from the Paroo, Nebine and Warrego catchments respectively, and overall within these three catchments there were strong gradients of both carp abundance (range 0–33 individuals CPUE, mean four individuals CPUE) and biomass (range 0–16,200 g CPUE, mean 2004 g CPUE) against which to consider ecosystem responses to carp density (Table 1).

The waters of the sites were generally near neutral to weakly alkaline, with a mean pH of 7.5 (Table 1). Two sites were exceptions with acidic water (Yowah Creek at Thandy Waterhole North, pH 4.6; Ward River at Bayrick Fish Hole, pH 6.4). The electrical conductivity was generally low, with a mean of 371 µS cm-1, but Cuttaburra Creek at Tinnenburra Waterhole was an exception having a high EC of 5760 µS cm-1, suggestive of possible saline groundwater input. Water temperatures at the time of sampling ranged from 13–29oC, with a mean of 22oC.

3.3 Responses to carp

3.3.1 Aquatic macroinvertebrates

Following removal of rare taxa, the macroinvertebrate fauna of the study sites comprised 38 taxa dominated by insects and crustaceans (Supporting Information site-by-taxa data, Supporting Information Table S1), with site richness varying from 18–31 taxa and the index of site abundance (henceforth referred to as ‘abundance’) ranging from 0–9 taxa (Table 1).

Contrary to expectations, neither macroinvertebrate richness (t = -1.03, df = 8.49, P = 0.83), nor macroinvertebrate abundance (t = 0.45, df = 9.32, P = 0.33) were significantly higher in the absence of carp at the catchment scale. Likewise, neither the abundance nor the richness of macroinvertebrates at sites in catchments where carp were present could be significantly explained by local carp biomass or abundance, nor by any of the other environmental variables representing site location or water quality. In both cases, generalized linear models were not significant, with P values much greater than 0.05 (P = 0.76 for the macroinvertebrate richness model and P = 0.81 for the macroinvertebrate abundance model).

Multivariate analyses found a small but significant difference in macroinvertebrate assemblage composition between sites depending on carp presence at the catchment scale (F = 2.4864, P = 0.004). The small magnitude of this effect is further indicated by the broad overlap of sites from catchments with and without carp across two-dimensional nMDS ordination space (Figure 4).

Indicator species analysis identified eight taxa making significant (P < 0.05) contributions to the small but significant differences in macroinvertebrate assemblage composition between sites in

This article is protected by copyright. All rights reserved. catchments with and without carp. Of these, three taxa (Gomphidae, Libellulidae and Glossiphoniidae) had lower abundances, and five taxa (Staphylinidae, Hydrochidae, Ecnomidae, Leptoceridae and Temnocephalidea) had higher abundances in the presence of carp. Those macroinvertebrates with higher abundances had larger indicator values, meaning they made more contribution to the difference than did the taxa indicative of carp absence (mean values of indicators of carp presence 0.68 and carp absence 0.40, respectively) (Table 2, Figure 5).

There was also a small but significant difference in macroinvertebrate assemblage composition between the Paroo, Warrego and Nebine catchments with carp present (F = 2.4894, p = 0.001) (Figure 6). The magnitude of the difference in macroinvertebrate assemblages among these three rivers with carp present (F = 2.489) was almost identical to that among sites from catchments with and without carp (F = 2.486), suggesting that catchment biogeography is the true driver of the observed differences. The nMDS bubble plot of sites in the three catchments with carp present, with symbols scaled by carp biomass (Figure 7), further illustrates the minor influence of carp biomass upon macroinvertebrate composition. A near identical result was obtained for carp abundance, which is therefore not presented. Fitting environmental vectors to this ordination further highlighted the lack of correlations between macroinvertebrate assemblage variability and both carp biomass and abundance, neither of which were significant (P > 0.05). Turbidity (r2 = 0.40, P = 0.003), water temperature (r2 = 0.30, P = 0.029) and catchment (r2 = 0.25, P = 0.01) were the only significant environmental vectors (P < 0.05).

BIOENV identified five environmental variables as the best subset matching patterns in site macroinvertebrate assemblages in the three catchments with carp present (Table 3). The Mantels test found a distance matrix based upon the best subset to have a significant but weak correlation with the macroinvertebrate distance matrix (r = 0.303, P = 0.002). The most important environmental variables in this relationship were EC and spatial descriptors, with carp abundance and biomass adding little explanatory advantage (Table 3).

3.3.2 N. sublineata

In contrast to the results for macroinvertebrate assemblage composition, there were stark differences in the occurrence of the river snail N. sublineata at sites from catchments with carp present and absent, with the species absent from every site in the catchments with carp, and

This article is protected by copyright. All rights reserved. present in all sites in the catchments without carp (Table 1). Because this distribution was a perfect fit to the model, ANOVA could not be undertaken, and because the snails were absent from all sites in the three catchments where carp are established, linear modelling could not be applied.

3.3.3 Turbidity

As expected, turbidity was high at most sites varying from 7–800 NTU, with site averages for the catchments Ambathala 83 NTU, Bulloo 352 NTU, Nebine 211 NTU, Paroo 421 NTU and Warrego 116 NTU (Table 1). Contrary to expectation, turbidity was not higher at sites in catchments with carp than in those without carp (t = 0.98, df = 15.18, P = 0.82). Mean site turbidity was actually lower, but not significantly so, at sites with carp (242 NTU) than those without carp (313 NTU). The generalized linear regression model of turbidity at sites in the three catchments with carp present was not significant (P = 0.14), indicating that variability in turbidity between sites could not be explained by carp biomass or abundance, site location, or other aspects of water quality.

3.3.4 Macrophytes

Macrophytes were absent from 27 of the 31 study sites. Where present, they were floating or emergent in growth and had low density (isolated, 1–5%) at all sites except one. The exception was Fifteen Mile Creek at Hay Paddock Hut Waterhole in the Bulloo catchment, which had higher density (beds/stands, 21–50%) of floating and emergent Nymphoides crenata (Figure 2). Other macrophyte taxa recorded at sites were Azolla sp., Ludwigia sp. and Persicaria sp. (see Supporting Information site by taxa data).

Macrophytes were present at three sites in the catchments without carp and one site in the catchments with carp. As expected, the proportion of sites with macrophytes was significantly greater (P = 0.03) in catchments without carp (43% of sites) than with carp (4% of sites).

3.3.5 Native fish

Eleven native fish species were collected, with the most widespread species, occurring at more than 10 sites, being golden perch (Macquaria ambigua), bony bream (Nematalosa erebi) and Hyrtl’s catfish (Neosilurus hyrtlii). These species also contributed the most to overall native fish biomass and substantially to total native fish abundance. Goldfish (Carassius auratus) was the only other non- native fish species apart from carp in the samples, and it occurred at 13 sites and in all catchments

This article is protected by copyright. All rights reserved. except the Bulloo (abundance 0–4 CPUE, biomass 0–206 g CPUE). In total, the biomass of carp from all sites (47 kg) was greater than the biomass of any native species (highest M. ambigua 31 kg), but three native species had higher total abundances (N. erebi 374, Ambassis agassizii 144 and N. hyrtlii 107) than did carp (96) (see Supporting Information site by taxa data).

Sites from catchments without carp had significantly higher native fish biomass, which on average was nearly four times greater than at sites in catchments with carp (mean without carp = 4713 g, mean with carp = 1221 g, one-sided t = 3.7, P = 0.004) and near-significantly higher native fish abundance (mean without carp = 72, mean with carp = 14, one-sided t = 1.9, P = 0.054) than catchments with carp present. The variances of both native fish biomass and native fish abundance were significantly greater in catchments without carp than with carp (one-sided F tests of variances: biomass F = 4.0, P = 0.007, abundance F = 72.8, P < 0.001).

Additional analyses were undertaken to test whether the significantly lower native fish biomass in catchments with carp represented displacement of native biomass by carp biomass, using both total fish biomass and total fish biomass without goldfish. Results were almost identical owing to the low biomass contribution of goldfish, so only total fish biomass is presented. The total fish biomass in catchments without carp was not significantly greater than in catchments with carp (mean without carp = 4713 g, mean with carp = 3235 g, one-sided t = 1.3, P = 0.11). Likewise, the variance was not significantly greater either (one-sided F test of variance F = 0.48, P = 0.82).

Site carp density had only a minor influence on native fish biomass and abundance at sites within the Murray-Darling basin catchments with carp present (Figure 8). The generalized linear regression model of native fish abundance at sites in the three catchments with carp present was significant (pseudo R2 = 0.38, P < 0.001), but the best model did not include metrics of carp density (Table 4). Rather, variability in native fish abundance was explained by spatial variables of catchment separated along a gradient in longitude and by upstream to downstream differences structured by latitude, with EC contributing because of the above average native fish abundance at the one site with very high EC (Supporting Information Figure S2). The equivalent generalized linear regression model of native fish biomass was also significant (pseudo R2 = 0.35, P < 0.001). The best-fit model included carp biomass but not carp abundance, and also included spatial and water quality attributes of sites (Table 4). As with abundance, the strongest influence on native fish biomass was

This article is protected by copyright. All rights reserved. spatial, with catchment separated along a gradient in longitude and by upstream to downstream differences structured by latitude. Carp biomass had a negative influence on native fish biomass as expected. In addition, turbidity, pH and water temperature had positive influences on native fish biomass and EC a negative influence (Supporting Information Figure S3)

4. Discussion

4.1 Ecological effects of carp

4.1.1 Aquatic ecosystems

This study has demonstrated that in the setting of the dryland rivers of the northern catchments of Australia’s Murray-Darling Basin, alien invasive carp generated some large and important ecosystem impacts – notably the reduction in native fish biomass, and the extirpation of the river snail N. sublineata at catchment scales. However, carp did not generate some of the other key ecosystem changes expected based on the synthesis of studies of carp impacts in other aquatic systems (Vilizzi et al., 2015; Weber & Brown, 2009). Furthermore, it is likely that the observed reduction in native fish biomass was not generated by the principal mechanisms expected.

Of the three key nodes in the expected carp impact pathways, two – macroinvertebrate assemblages and turbidity – were not responsive here to carp presence or carp density. This is despite the study system of isolated waterholes in dryland rivers offering several apparent advantages for detecting such impacts if they were present: suitable spatial control rivers with similar environmental settings but without introduced carp, expected dominance of proximal over distal influences on local ecosystems during long hydrological isolation of sites, minimal influence of compromising human stressors to ecosystems, and strong similarity in antecedent hydrological conditions at all sites. The third key node in the expected carp impact pathways – macrophyte occurrence – was significantly reduced by carp presence, but even without carp, macrophytes are naturally rare, at very low densities, and restricted in growth in the isolated riverine waterholes of this study. The only important exception to the conclusion that carp had no meaningful impacts via these three nodes was the strong finding that carp extirpate the endangered river snail N. sublineata. There was also a strong and significant reduction in native fish density in relation to carp presence, and less so to carp

This article is protected by copyright. All rights reserved. density, but this appears perpetrated via a different impact pathway from the hypothesised effects via carp modifications to turbidity, macroinvertebrate and macrophyte density.

It is important to recognize that this study was a ‘natural experiment’, relying on the un-manipulated distributions of carp as experimental treatments. As such, inferences drawn cannot be as strong as those from controlled manipulative experimentation, and this caveat has been highlighted previously for ecological studies of invasive fish impacts (Tricarico, Junqueira, & Dudgeon, 2016). By their nature, natural experiments cannot definitively attribute causation to patterns, since variation in extrinsic environmental factors is uncontrolled and one or more such factors may be responsible for the observed patterns (see Cooper and Dudley, 1988). Despite the observations of equivalence between treatments in many potential confounding factors, this remains the case here. Further confirmation of the carp impacts in dryland rivers observed here awaits future experimental manipulation of carp density in these systems.

4.1.2 N. sublineata

The results reinforce the documented loss of N. sublineata from throughout much of the Murray- Darling Basin, here for the first time for the northern extremes of the Basin. There has been uncertainty as to the principal causes of this decline, with one of the best-supported hypotheses being that it stems from changes in the nature of the periphyton community on which the snails feed as a result of altered flow regimes. Other possible explanations for the species decline include removal of large wood from rivers, increased sedimentation, and the invasion of carp (Sheldon & Walker, 1993). The results here refute altered flow regimes as the main mechanism, as the catchments in this study were subject only to minor water resource development and many of the individual sites with N. sublineata absent had little or no flow modification (State of Queensland; Department of Science, Information Technology, Innovation and the Arts, 2013) (Supporting Information Table S2). It is also unlikely that differences in removal of large wood or sedimentation are notable in these little-developed catchments (Negus et al., 2015). Results of this study clearly implicate carp as a likely mechanism of N. sublineata decline, as the presence and absence of carp at the catchment scale directly corresponds to patterns found in N. sublineata occurrence (Supporting Information Table S2).

This article is protected by copyright. All rights reserved. Elsewhere, carp have been shown to severely deplete the abundance of a number of gastropod species by predation. For example, in experimental wetland enclosures, all snails of several species were consumed in only 8 weeks except for the few individuals that were too large to fit into the carp’s mouths, prompting the description of carp as molluscivores, and the suggestion that they could be used for biocontrol of invasive gastropods (Ip et al., 2014; Wong, Kwong, & Qiu, 2009). Although these attributes were not measured, the mouths of most carp collected in this study were easily large enough for them to eat all of the N. sublineata individuals collected. Thus, the documented molluscivorous habits of carp, coupled with the diametrically-opposed distributions of carp and N. sublineata at the catchment scale, provide compelling evidence that carp predation is a major driver of the decline of N. sublineata in the Murray-Darling Basin. Thus, carp are implicated in their listing as endangered on both the IUCN red list of threatened species and in New South Wales (New South Wales Department of Primary Industries, 2007; Ponder, 1996).

4.1.3 Turbidity

The mechanism by which carp increase turbidity as identified in other studies is largely via their omnivorous benthic feeding behaviour where they repeatedly suck and filter sediment (termed roiling), with the resulting resuspension of fine bed material increasing turbidity (Vilizzi et al., 2015). To a lesser extent, their spawning behaviour can also resuspend fine sediment. Previous studies directly examining the relationship between carp and turbidity have used in situ and ex situ (artificial) experimental enclosures, or direct carp removal. All focus on off-channel and still-water habitats such as wetlands, billabongs, reservoirs, or experimental ponds (Supporting Information Table S3). In these studies, increased carp biomass typically contributed to increased turbidity, and exclusion experiments resulted in reduced turbidity. However, the restricted focus of these studies to lentic environments neglects the vast riverine environments where carp are present in very high densities. Extrapolating from lentic to lotic habitats is likely to be inappropriate, as sediment composition, dynamics and disturbance profiles may be very different. The interaction of intermittent flow, carp and turbidity requires independent examination.

The dryland rivers studied here are characterized by a natural propensity for high turbidity, which has been realized as a result of more than 100 years of catchment disturbance from activities such as livestock grazing and native vegetation clearing. Their catchments have a high content of clays

This article is protected by copyright. All rights reserved. associated with high sodium concentrations, which increases their dispersability (Kirk, 1985; Power et al., 2007), and because these particles are typically very small (Perret, Newman, Nègre, Chen, & Buffle, 1994) they are colloidal and do not readily settle (Buffle & Leppard, 1995), resulting in perpetually high turbidity (Boulton & Brock, 1999). Despite this propensity for high turbidity, when first encountered by early explorers in Australia, some of these now turbid rivers were described as being clear (Scott, 2005), and the Darling River in the northern Murray-Darling Basin has been found to have suspended sediment yields nearly 20 times greater than before European settlement (Dosseto, Turner, & Douglas, 2006). Such changes to optical water properties occurred well before carp invasions (Gell et al., 2009), probably as a result of post-European settlement agricultural and grazing practices (Prosser et al., 2001) and the influence of bed and bank disturbances by feral mammals such as pigs and goats (Negus et al., 2015; Steward, Negus, Marshall, Clifford, & Dent, 2018). Even in other studies where carp have been shown to increase turbidity, factors other than carp have been identified as having a greater influence (King, Robertson, & Healey, 1997). Such observed changes suggest that carp have not added significantly to this past increase in turbidity in these dryland river systems.

4.1.4 Macrophytes

Although there was a carp effect, with a significantly greater proportion of sites in catchments without carp having some macrophytes present than in catchments with carp, macrophytes were either absent or at very low density almost everywhere sampled. Low macrophyte density and the species recorded are consistent with previous surveys in the dryland rivers of western Queensland where macrophytes were rare (Mcgregor, Marshall, & Thoms, 2006; Silcock, 2009). This was attributed to light limitation owing to perpetually high turbidity resulting in limited euphotic areas for macrophyte growth (Bunn, Thoms, Hamilton, & Capon, 2006). Constant high turbidity can attenuate light to such an extent that submerged macrophytes are excluded (Jones, Collins, Naden, & Sear, 2012). Although their absence in the current study may be also in part because the photographic sampling method used may not detect submerged macrophytes as readily as those at or above the water surface, submerged macrophytes are seldom if ever encountered in these catchments. Light limitation also potentially explains the result that all macrophytes detected were emergent or floating (Middelboe & Markager, 1997), and the low density even of these forms. Although the impact of turbidity and light attenuation is less severe for emergent and floating

This article is protected by copyright. All rights reserved. macrophytes, their submerged parts also contribute substantially to photosynthesis, particularly early in their growth, so that they are still affected by constant high turbidity (Jones et al., 2012). There is no obvious explanation why one site, Fifteen Mile Creek at Hay Paddock Hut Waterhole, had conspicuous emergent and floating macrophyte cover (Figure 2), as its turbidity of 352 NTU was high and unexceptional; however, these plants here were restricted to shallow littoral regions of the waterhole as reported elsewhere for emergent and floating macrophytes in perpetually turbid rivers (Jones et al., 2012).

As turbidity was not significantly increased by carp, their impacts on macrophytes observed here were most likely from direct rather than indirect carp influences. Direct carp impacts reported elsewhere include uprooting of macrophytes while feeding, especially in soft substrates, and their occasional consumption, which together can lead to reduced macrophyte diversity and abundance or even total elimination (Weber & Brown, 2009). Indeed, the macrophyte taxa recorded in this study have traits reported to reduce vulnerability to carp, such as rapid growth, vegetative propagation and roots resistant to physical damage (Okada, Grewell, & Jasieniuk, 2009; Partridge, 2001; Stephens & Dowling, 2002; Wagner, 1997).

4.1.5 Aquatic macroinvertebrate assemblages

In contrast to the results for N. sublineata, general macroinvertebrate assemblages, sampled using the standard approach adopted for river health monitoring in Australia for the past 25 years (Davies, 2000), showed little to no response to the presence of carp at the catchment scale. Contrary to expectations, both macroinvertebrate richness and abundance were on average slightly higher in the presence of carp than in their absence, although these differences were not significant. The multivariate composition of assemblages was slightly different depending on carp presence or absence from catchments, but these differences were of the same magnitude as differences among the three individual catchment assemblages where carp are present, suggesting the effect of carp to be very minor to insignificant. Several individual taxa had significantly lower abundances in catchments with carp present, but there were more taxa and with greater effect sizes that had significantly higher abundances in the catchments with carp present. Although this suggests little or no true influence of carp on assemblage composition, it remains possible that the three taxa

This article is protected by copyright. All rights reserved. identified, Gomphidae, Libellulidae (both Odonata) and Glossiphoniidae (Hirudinea), may have been adversely affected by the presence of carp. Elsewhere, Odonata have been shown to be affected by carp predation (Weber & Brown, 2009). Likewise, other taxa identified may have possibly benefited from the presence of carp: Staphylinidae, Hydrochidae (both Coleoptera), Ecnomidae, Leptoceridae (both Trichoptera), and Temnocephalidea (Platyhelminthes). There is no obvious reason to explain why these particular eight taxa would be affected by carp presence compared with the 30 other taxa not significantly influenced in either way. Similarly, in the three catchments with carp present, macroinvertebrate assemblages showed little to no response to the density of carp at the site scale, with carp biomass and abundance explaining little of the variability in macroinvertebrate assemblages between sites.

These findings are in contrast to other studies that have found impacts from carp on macroinvertebrates (Vilizzi et al., 2015). The nature of the dryland river environment provides some possible explanation for this. Populations of macroinvertebrates that inhabit these rivers are resilient to frequent and widespread local extirpations from drying as a result of flow intermittency and subsequent habitat drying (Marshall, Sheldon, Thoms, & Choy, 2006). This resilience via dispersal may predispose them to unresponsiveness to carp impacts by permitting rapid repopulation of depleted habitat patches following carp predation. Furthermore, the harsh environments of dryland rivers result in tolerant, generalist macroinvertebrate assemblages (Marshall, Sheldon, et al., 2006; Marshall, Steward, et al., 2006) more resistant to indirect carp impacts, such as habitat alteration or increases in nutrient concentrations (Weber & Brown, 2009), than more sensitive macroinvertebrates in other settings. Indirect carp impacts to macroinvertebrates, from modifications to turbidity and macrophyte density are not likely here, given the results on macrophytes and turbidity.

4.1.6 Native fish

The mean biomass of native fish in catchments with carp present was approximately 400% lower than in the catchments with no carp. This difference is an order of magnitude greater than the model-based predictions of Kopf et al. (2019), who predicted a reduction of 47–68% in native fish biomass due to carp across the rivers of the Murray-Darling Basin, probably as a result of carp feeding at low trophic levels, thereby converting food and energy previously available to native

This article is protected by copyright. All rights reserved. fishes into carp biomass. This, they argued, in concert with their long lifespan and a lack of predators for mature carp, results in monopolization and retention of energy by carp to the detriment of native fish species. Their conclusion is reinforced by the results of this study.

River food-webs are supported at their bases by high quality algal production (Brett et al., 2017; Guo, Kainz, Sheldon, & Bunn, 2016), which in turbid dryland rivers is light-limited (Bunn, Balcombe, Davies, Fellows, & McKenzie-Smith, 2006). Food limitation in waterholes has been implicated for mass fish mortality (Arthington & Balcombe, 2011; Arthington et al., 2005; Balcombe, Bunn, McKenzie-Smith, & Davies, 2005) and access to high quality food enhances fish survivorship (Sternberg, Balcombe, Marshall, Lobegeiger, & Arthington, 2012). Given these processes, it is reasonable to assume that utilization of basal resources by carp reduces their availability to native fishes, reducing native fish biomass. However, the trophic relationships of carp in these rivers are poorly understood (Jardine et al., 2015) and confirmation by studies on food webs and energetics in systems with and without carp are clearly needed, with the rivers used in this study being an ideal setting for further investigations.

Carp can also feed on the early life stages of native fishes, thus reducing their recruitment and biomass (Weber & Brown, 2011), but in the dryland rivers of the northern Murray-Darling Basin there is no indication of this occurring, even shortly after floodplain inundation when many native fishes spawn in these systems (Woods, Lobegeiger, Fawcett, & Marshall, 2012). Although it seems unlikely that piscivory is a major mechanism by which carp reduced native fish biomass in this study, it may exert some influence, through impacts on eggs, juveniles and disturbance of freshwater catfish nests.

Interpretation of native biomass differences here needs to consider also that the two carp-less catchments are at the west of an east–west geographical temperature gradient, with westerly regions experiencing higher temperatures, which may in turn result in greater rates of primary production and hence more food to support higher fish biomass (Balcombe et al., 2010). This may have contributed to the Bulloo and Ambathala catchment sites having higher native fish biomass than the other catchments. However, their total fish biomass was not significantly greater than that of the other catchments (including both native fish and carp), which reinforces our conclusion that

4.2 Reversing the impacts of carp

The matter of reversibility of impacts should carp control succeed is mainly relevant in these rivers to N. sublineata and native fish biomass where carp impacts were greatest.

Notopala sublineata differs from most other macroinvertebrates in these waterholes in its life history strategy to survive flow intermittency. Whereas most macroinvertebrate taxa rely on high resilience via dispersal to repopulate, N. sublineata relies upon high desiccation resistance to allow local populations to survive in situ during drying events. This is achieved by snails tightly closing the shell aperture with a thick operculum that seals them against water loss (Carini, Hughes, & Bunn, 2006). The limited vagility of the species and its viviparous life cycle with no larval stage suggest low dispersal capacity, confirmed by high levels of genetic structuring and restricted gene flow among populations both within and among dryland river catchments (Carini & Hughes, 2006). This implies that recolonization following local extirpations may not occur or at least would take a very long time, which may explain why N. sublineata distribution here was determined by carp presence at the catchment scale rather than local carp density, and why the species has disappeared throughout its range overlap with carp, except in isolated pockets where carp cannot feed on them, such as in irrigation pipes (Sheldon & Walker, 1993). This suggests that the benefits of successful carp biocontrol for this snail would be realized only if it were both entirely effective, which is unlikely (Becker et al., 2018; Marshall et al., 2018), and followed up with dedicated complementary reintroduction actions for most locations.

Reversing carp impacts on native fish biodiversity in dryland rivers may be more achievable following successful biocontrol. Given the results of this study, the main mechanism by which carp control would be likely to benefit native fish biomass would be by lessening carp monopolization of high- quality, basal food resources, thereby making more available for conversion into native fish biomass. The generally low levels of other human stressors on fish biomass in these study rivers (Negus et al., 2015) may also expedite the potential reversibility of carp impacts. Non-linearity between carp biomass reduction and the potential recovery of native fish biomass (Kopf, Humphries, Bond, Sims, & Watts, 2019) make it difficult to forecast biocontrol response, but we suggest that in dryland rivers

This article is protected by copyright. All rights reserved. it has the potential to be greater than the 17–47% biomass recovery Kopf et al. (2019) modelled for these rivers. However, it is also important to recognize that the responses of native species to successful invasive species control are poorly understood (Lurgi, Ritchie, & Fordham, 2018), difficult to predict and can even be negative (Ballari, Kuebbing, & Nuñez, 2016), with the potential for increased abundance of alternative invasive or nuisance species (Ballari et al., 2016; Kopf et al., 2017). In the systems studied here, successful control of carp could potentially result in proliferation of other exotic fish species already present such as goldfish, or the accidental or deliberate introduction and establishment of invasive Oreochromis mossambicus, the Mozambique tilapia, which is present in catchments to the north and east of the study region (Hutchison, Sarac, & Norris, 2011), to the detriment of native species recovery. Given the trophic position of carp, it is also possible that their successful control could result in nuisance algal blooms to the detriment of native species (Carman & Tomevska, 2019). Application of tools such as multispecies ecological network models to aid in predictions of ecosystem outcomes from control of invasive species are warranted to reduce these uncertainties (Kopf et al., 2017; Lurgi et al., 2018).

Influences from other threats and stressors can also prevent native species recovery following successful control of an invasive species (Arthington, Dulvy, Gladstone, & Winfield, 2016; Kopf et al., 2017). In the rivers of Australia’s Murray-Darling Basin there are many interacting threats and stressors in addition to carp, including altered flow regimes, riparian habitat loss and degradation, river and floodplain disconnection, barriers to fish passage, poor water quality, and cold water pollution. In fact it has been suggested that in the more developed, southern parts of the Basin, carp proliferation may be a symptom rather than a cause of declines in river health (Driver, Harris, Norris, & Closs, 1997; Koehn et al., 2000). Carp control would represent a critical and significant step toward native species recovery and overall restoration of the Murray-Darling Basin; however, carp control in isolation is unlikely to achieve total recovery in most locations. Native species recovery requires a wider restoration programme on decadal timescales with complementary actions and technical expertise from all facets of applied aquatic ecology.

4.3 Conclusions Several key aspects of the general understanding of how carp affect aquatic ecosystems (Vilizzi et al., 2015; Weber & Brown, 2009) were shown here not to apply to Australian dryland rivers (Figure 9). These differences are driven largely by system setting. Characteristics of Australian dryland rivers –

This article is protected by copyright. All rights reserved. their hydrology, topography, geology and geomorphology – combine to render inappropriate many ecological concepts that were derived elsewhere (Boulton & Brock, 1999), and the same applies to concepts of carp impacts on aquatic ecosystems. This reinforces the general need for careful consideration when transferring ecological understanding between systems that have differences in critical characteristics and function differently. Specifically, this highlights the need to understand system characteristics relevant to causal impact pathways when applying generic carp impact models to specific settings. This has global relevance to future carp control efforts.

5. Acknowledgements Fish and macroinvertebrate samples were collected under the authority of Queensland General Fisheries Permit 153301, and animal ethics approval for the work was granted by Queensland Ethics Permit DSITIA/2012/07/03. The work was funded and undertaken as part of the Queensland Government Q-catchments programme of river condition assessment. Assistance with data collection in the field was provided by James Fawcett, Bill Senior, John Bowlen, Jaye Lobegeiger, Delwyn Hansen, Glenn McGregor, Tess Mullins, Dean Holloway and Ric Newson. The Australian Wildlife Conservancy and many landholders, including Traditional Owners, gave their permissions and assisted with site access. Thanks to Shane Brooks, Jarrod Lyon and Ivor Stewart for permission to use their conceptual model of carp impacts that was modified here as Figure 9. Philip Boon, Przemek Bajer, Stephen Balcombe, Bill Senior, Dean Holloway and an anonymous referee reviewed and provided fabulous suggestions to improve earlier versions of this manuscript, for which the authors are most grateful. Author Marshall is a member of the Scientific Advisory Group for the NCCP and declares no conflict of interest in the preparation of this manuscript.

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)

b (g) (g) pH biomass EC (°C) Site Carp CPUE (NTU) CPUE sublineata present number carp (%) carp (%) richness

(µS cm Turbidity Site name Native fish Catchment abundance Water temp. N. Carp Total fish CPUE Total fish biomass Macroinvertebrate Macroinvertebrate Native fish biomass

1 Ambathala Creek at Ambathala Ambathala 0 0 0 0 84 4036 24 8 YES 83 7.6 104 23 2 Blackwater Creek at Adavale Bulloo 0 0 0 0 31 8579 28 2 YES 453 7.5 161 21 3 Bulloo River at Norley Station Bulloo 0 0 0 0 162 3897 22 8 YES 230 7.5 116 24 4 Bulloo River at Pinkenetta Station Bulloo 0 0 0 0 4 1388 19 4 YES 534 7.4 101 27 5 Fifteen Mile Creek at Hay Paddock Hut Waterhole Bulloo 0 0 0 0 203 7079 18 2 YES 352 7.3 99 26 6 Bulloo River at Thargomindah Bulloo 0 0 0 0 7 4408 22 2 YES 278 7.5 117 27 7 Bulloo River at Quilpie Bulloo 0 0 0 0 11 3604 30 3 YES 263 7.2 107 29 8 Nebine Creek at Murra Murra Nebine 3 1028 24 53 9 811 26 1 NO 510 7.3 62 17 9 Wallam Creek at Bollon Reserve Nebine 2 350 6 7 29 4810 26 2 NO 39 7.7 174 21 10 Nebine Creek at Aqua Downs Nebine 1 297 100 100 0 0 23 3 NO 102 7.4 80 22 11 Wallam Creek at Homeboin Waterhole Nebine 3 2700 12 51 18 2630 28 5 NO 100 7.4 137 22 12 Wallam Creek at One Mile Waterhole Nebine 1† 297† 0 0 14 1024 27 4 NO 16 7.7 196 24 13 Nebine Creek at Roseleigh Crossing Nebine 33 16200 63 97 20 516 20 3 NO 500 8.3 441 24 14 Paroo River at Wimmera Waterhole Paroo 2 933 15 39 8 1384 28 3 NO 298 7.4 84 18 15 Moonjaree Creek at Moonjaree Paroo 4 1452 21 47 14 1590 19 5 NO 489 7.4 110 17 16 Gumholes Creek at Bowra 1 Paroo 2 3700 5 79 40 989 28 8 NO 298 7.6 122 19

This article is protected by copyright. All rights reserved. 17 Beechal Creek at Boolbury Waterhole Paroo 2 801 6 34 25 1530 27 4 NO 360 7.7 65 14 18 Yowah Creek at Thandy Waterhole North Paroo 22 6631 53 96 16 190 22 6 NO 477 4.6 290 18 19 Paroo River at 14 Mile Waterhole Paroo 5 2688 52 64 4 1393 31 4 NO 514 7.5 81 13 20 Pingine Waterhole at Adavale Cheepie Rd Paroo 2 1035 12 49 10 861 24 9 NO 800 7.6 62 18 21 Rolwegan Creek at Unnamed Waterhole Paroo 3 1572 19 74 11 539 29 4 NO 133 8.0 660 21 22 Cuttaburra Creek at Tinnenburra Waterhole Warrego 2 685 6 59 27 458 22 0 NO 110 8.6 5760 23 23 Warrego River at Merwah waterhole Warrego 1† 398† 0 0 9 771 21 0 NO 7 8.0 263 25 24 Langlo River at Rylestone Waterhole Warrego 4 2039 25 50 10 1896 30 9 NO 502 7.3 69 19 25 Ward River at Bayrick Fish Hole Warrego 5 1828 31 66 10 833 22 0 NO 111 6.4 139 23 26 Warrego River at Charleville Warrego 1 1200 30 71 2 495 27 4 NO 28 7.7 454 23 27 Warrego River at Cunnamulla Weir Warrego 0 0 0 0 8 156 18 3 NO 18 7.8 298 19 28 Warrego River at Wyandra Warrego 1 † 398 † 0 0 14 963 28 2 NO 29 7.6 271 22 29 Warrego River at Augathella Warrego 0 0 0 0 22 1213 29 2 NO 35 7.6 540 21 30 Ward River at Binnowee Warrego 1 398 100 100 0 0 24 1 NO 310 7.4 81 20 31 Warrego River at Wallen Warrego 1 1468 7 25 13 4247 25 5 NO 14 8.1 273 25 aCPUE – catch per unit effort bEC – electrical conductivity

† Carp observed at the site but not captured in sample – lowest CPUE and biomass measured from other sites in the catchment were adopted for analyses

This article is protected by copyright. All rights reserved. Table 2. Taxa identified by indicator species analysis as making significant (P < 0.05) contributions to the small but significant differences in macroinvertebrate assemblage composition among sites in catchments with and without carp present. Macroinvertebrate indicators of carp presence or absence had higher abundance in catchments with carp present or absent, respectively.

Taxon Indicator of carp Indicator value Probability

Gomphidae absence 0.53 0.003

Libellulidae absence 0.41 0.019

Glossiphoniidae absence 0.28 0.042

Staphylinidae presence 0.71 0.007

Ecnomidae presence 0.70 0.03

Leptoceridae presence 0.67 0.012

Hydrochidae presence 0.66 0.048

Temnocephalidea presence 0.64 0.025

This article is protected by copyright. All rights reserved. Table 3. Results of BIOENV analysis to identify sub-sets of environmental variables to generate a distance matrix that best matches the macroinvertebrate distance matrix from samples in three river catchments with carp present. The five-variable sub-set was the best fit.

Number of variables Best set Spearman r 1 ECa 0.207 2 Catchment, ECa 0.265 3 Latitude, Longitude, ECa 0.273 4 Catchment, Latitude, ECa, Carp abundance 0.283 5 Catchment, Latitude, ECa, Turbidity, Carp biomass 0.303 aEC – electrical conductivity

This article is protected by copyright. All rights reserved. Table 4. Best generalized linear regression models of native fish biomass and abundance. In each case, all variables with listed coefficients were significant (P < 0.05).

(Intercept) Latitude Longitude Catchment Catchment Carp Turbidity pH ECa Water Paroo Warrego biomass temperature Native fish biomass -216.7587 -0.5039 1.3926 2.6093 1.7515 -0.0001 0.0009 0.5517 -0.0004 0.0483 Native fish abundance -201.9840 -0.3728 1.3194 2.8446 1.5167 0.0002 aEC is electrical conductivity

Figure 1. Map of the study region indicating the location of study catchments and sites and the biomass of carp sampled from each site. Catchments shaded grey have carp present and white have carp absent. Inset map shows the location of the study region in Australia and in relation to the Murray-Darling Basin (MDB). Site numbers refer to sites as listed in Table 1.

Figure 2. Photos of selected sampling sites in each of the catchments. Site 5 was the only one of the 31 sampling sites with conspicuous macrophytes present.

Figure 3. Targeted sampling for the endangered river snail Notopala sublineata in a Bulloo River waterhole.

Figure 4. Two-dimensional nMDS plot illustrating differences in macroinvertebrate assemblage composition between sites, coded to indicate river catchment and the presence (solid symbols) or absence (open symbols) of carp from those catchments. Polygons surrounding points illustrate the large overlap in macroinvertebrate assemblages between sites from catchments with and without carp.

Figure 5. Box plots of the abundance of macroinvertebrate taxa identified by indicator species analysis as making significant (P < 0.05) contributions to differences in assemblage composition among sites in catchments with carp absent and present. Bold bars indicate medians, boxes the interquartile range (IQR), the whiskers the minimum and maximum values that do not exceed 1.5 times the IQR, and outlying points any values beyond these.

This article is protected by copyright. All rights reserved. Figure 6. Ordination plot of macroinvertebrate assemblages at sites in the three river catchments with carp present. Symbols indicate catchment and polygons surrounding points illustrate the large overlap in macroinvertebrate assemblages between these three catchments.

Figure 7. Ordination bubble plot of macroinvertebrate assemblage composition at sites in the three river catchments with carp present. Symbols indicate catchment, and the size of the symbols are scaled to indicate the relative biomass of carp at the sites.

Figure 8. Biplots of carp density and native fish metrics at the sites in catchments with carp present (a) carp biomass vs native fish biomass, (b) carp biomass vs native fish abundance, (c) carp abundance vs native fish biomass, (d) carp abundance vs native fish abundance.

Figure 9. Refined conceptual model of carp impacts on aquatic ecosystems of dryland river waterholes, based on findings from this study. Modified from Brooks (2018) based on Weber & Brown (2009) and Vilizzi et al. (2015) and used with permission.