Unravelling the Diversity of Microorganisms in Ticks from Australian Wildlife.

Ticks and tick-borne pathogens pose a significant threat to the health and welfare of humans and animals. Our knowledge about pathogens carried by ticks of Australian wildlife is limited. This study aimed to characterise ticks and tick-borne microorganisms from a range of wildlife species across six sites in Victoria, Australia. Following morphological and molecular characterisation (targeting 16S rRNA and cytochrome c oxidase I), tick DNA extracts (n = 140) were subjected to microfluidic real-time PCR-based screening for the detection of microorganisms and Rickettsia-specific real-time qPCRs. Five species of ixodid ticks were identified, including Aponomma auruginans, Ixodes (I.) antechini, I. kohlsi, I. tasmani and I. trichosuri. Phylogenetic analyses of 16S rRNA sequences of I. tasmani revealed two subclades, indicating a potential cryptic species. The microfluidic real-time PCR detected seven different microorganisms as a single (in 13/45 ticks) or multiple infections (27/45). The most common microorganisms detected were Apicomplexa (84.4%, 38/45) followed by Rickettsia sp. (55.6%, 25/45), Theileria sp. (22.2% 10/45), Bartonella sp. (17.8%, 8/45), Coxiella-like sp. (6.7%, 3/45), Hepatozoon sp. (2.2%, 1/45), and Ehrlichia sp. (2.2%, 1/45). Phylogenetic analyses of four Rickettsia loci showed that the Rickettsia isolates detected herein potentially belonged to a novel species of Rickettsia. This study demonstrated that ticks of Australian wildlife carry a diverse array of microorganisms. Given the direct and indirect human-wildlife-livestock interactions, there is a need to adopt a One Health approach for continuous surveillance of tick-associated pathogens/microorganisms to minimise the associated threats to animal and human health.

Environmental correlates of temporal variation in the prey species of Australian fur seals inferred from scat analysis.

Marine ecosystems in southeastern Australia are responding rapidly to climate change. We monitored the diet of the Australian fur seal (Arctocephalus pusillus doriferus), a key marine predator, over 17 years (1998-2014) to examine temporal changes. Frequency of occurrence (FO) of prey was used as a proxy for ecosystem change. Hard part analysis identified 71 prey taxa, with eight dominant taxa in greater than 70% of samples and predominantly included benthic and small pelagic fish. FO changed over time, e.g. redbait (Emmelichthys nitidus) reduced after 2005 when jack mackerel (Trachurus declivis) increased, and pilchard (Sardinops sajax) increased after 2009. Using generalized additive models, correlations between FO and environmental variables were evident at both the local (e.g. wind, sea surface temperature (SST)) and regional (e.g. El Niño-Southern Oscillation Index (SOI), Southern Annular Mode (SAM)) scales, with redbait and pilchard showing the best model fits (greater than 75% deviance explained). Positive SAM was correlated to FO for both species, and wind and season were important for redbait, while SOI and SST were important for pilchard. Both large-scale and regional processes influenced prey taxa in variable ways. We predict that the diverse and adaptable diet of the Australian fur seal will be advantageous in a rapidly changing ecosystem.

Genomic insights into the secondary aquatic transition of penguins.

Penguins lost the ability to fly more than 60 million years ago, subsequently evolving a hyper-specialized marine body plan. Within the framework of a genome-scale, fossil-inclusive phylogeny, we identify key geological events that shaped penguin diversification and genomic signatures consistent with widespread refugia/recolonization during major climate oscillations. We further identify a suite of genes potentially underpinning adaptations related to thermoregulation, oxygenation, diving, vision, diet, immunity and body size, which might have facilitated their remarkable secondary transition to an aquatic ecology. Our analyses indicate that penguins and their sister group (Procellariiformes) have the lowest evolutionary rates yet detected in birds. Together, these findings help improve our understanding of how penguins have transitioned to the marine environment, successfully colonizing some of the most extreme environments on Earth.

Sustained reduction in numbers of Australian fur seal pups: Implications for future population monitoring.

Fur seal populations in the Southern Hemisphere were plundered in the late 1700s and early 1800s to provide fur for a clothing industry. Millions of seals were killed resulting in potentially major ecosystem changes across the Southern Hemisphere, the consequences of which are unknown today. Following more than a century of population suppression, partly through on-going harvesting, many of the fur seal populations started to recover in the late 1900s. Australian fur seals (Arctocephalus pusillus doriferus), one of the most geographically constrained fur seal species, followed this trend. From the 1940s to 1986, pup production remained at approximately 10,000 per year, then significant growth commenced. By 2007, live pup abundance had recovered to approximately 21,400 per year and recovery was expected to continue However, a species-wide survey in 2013 recorded a 20% decline, to approximately 16,500 live pups. It was not known if this decline was due to 2013 being a poor breeding year or a true population reduction. Here we report the results of a population-wide survey conducted in 2017 and annual monitoring at the most productive colony, Seal Rocks, Victoria that recorded a large decline in live pup abundance (-28%). Sustained lower pup numbers at Seal Rocks from annual counts between 2012-2017 (mean = 2908 ± 372 SD), as well as the population-wide estimate of 16,903 live pups in 2017, suggest that the pup numbers for the total population have remained at the lower level observed in 2013 and that the 5-yearly census results are not anomalies or representative of poor breeding seasons. Potential reasons for the decline, which did not occur range-wide but predominantly in the most populated and long-standing breeding sites, are discussed. To enhance adaptive management of this species, methods for future monitoring of the population are also presented. Australian fur seals occupy several distinct regions influenced by different currents and upwellings: range-wide pup abundance monitoring enables comparisons of ecosystem status across these regions. Forces driving change in Australian fur seal pup numbers are likely to play across other marine ecosystems, particularly in the Southern Hemisphere where most fur seals live.

Into the sea: Antimicrobial resistance determinants in the microbiota of little penguins (Eudyptula minor).

Terrestrial and aquatic birds have been proposed as sentinels for the spread of antimicrobial resistant bacteria, but few species have been investigated specifically in the context of AMR in the marine ecosystem. This study contrasts the occurrence of class 1 integrons and associated antimicrobial resistance genes in wild and captive little penguins (Eudyptula minor), an Australian seabird with local population declines. PCR screening of faecal samples (n = 448) revealed a significant difference in the prevalence of class 1 integrons in wild and captive groups, 3.2% and 44.7% respectively, with genes that confer resistance to streptomycin, spectinomycin, trimethoprim and multidrug efflux pumps detected. Class 1 integrons were not detected in two clinically relevant bacterial species, Klebsiella pneumoniae or Escherichia coli, isolated from penguin faeces. The presence of class 1 integrons in the little penguin supports the use of marine birds as sentinels of AMR in marine environments.

A physiological cost to behavioural tolerance.

Few studies of animal escape behaviour simultaneously investigate behavioural and physiological responses. Differences between these response types, however, have consequences for the way in which habituation or tolerance is interpreted - behavioural habituation may incur physiological costs. We simultaneously measured heart rate (HR) and behavioural responses during standardised approaches to incubating Masked Lapwings Vanellus miles, an urban-frequenting ground-nesting bird. We describe the existence of a distinct Physiological-Initiation Distance (PID) that precedes Flight-Initiation Distance (FID) but does not necessarily precede Alert Distance (AD). Two distinct response types occurred: 'startle', where a behavioural or physiological response coincided with the appearance of a person (always the investigator; 75.9 % of 58 birds) and 'non-startle' responses, where a behavioural or physiological response occurred after the appearance of, and commencement of the approach by, the person (24.1 % or 14 birds). For birds which were not startled, the interval between the initial heart rate increase and heart rate peak increased with clutch age. For birds which were startled, longer durations of post-peak HR elevation were associated with shorter FIDs and older clutches. Thus, reduced FIDs (generally interpreted as a sign of habituation or tolerance) are associated with greater physiological costs through longer durations of elevated HR. Additionally, the existence of, often long and undetectable, PIDs suggests: 1) that behavioural measures of response underestimate responses in general, and 2) that the methodological assumption when collecting FIDs, that starting distances exceed response distances, may often be incorrect yet are visually undetectable. Further studies of a variety of taxa are warranted to determine the associations between behavioural and physiological responses, and should these associations prove reliable, they would ideally generate general predictions of PID from readily measurable behavioural metrics (FID or AD), thus enabling prescriptions to manage the consequences of human interactions with wildlife.

Trophic niches of a seabird assemblage in Bass Strait, south-eastern Australia.

The foraging niches of seabirds are driven by a variety of factors, including competition for prey that promotes divergence in trophic niches. Bass Strait, south-eastern Australia, is a key region for seabirds, with little penguins Eudyptula minor, short-tailed shearwaters Ardenna tenuirostris, fairy prions Pachyptila turtur and common diving-petrels Pelecanoides urinatrix being particularly abundant in the region. The trophic niches of these species were investigated using isotopic values in whole blood and by identifying prey remains in stomach contents. The four species occupied different isotopic niches that varied among years, seasons and regions. Little penguins consumed mainly fish whereas the three procellariforms primarily consumed coastal krill Nyctiphanes australis. The dietary similarities between the procellariforms suggest that food resources are segregated in other ways, with interspecific differences in isotope niches possibly reflecting differential consumption of key prey, divergent foraging locations and depth, and differences in breeding phenology. Because oceanographic changes predicted to occur due to climate change may result in reduced coastal krill availability, adversely affecting these seabird predators, further information on foraging zones and feeding behaviour of small procellariform species is needed to elucidate more fully the segregation of foraging niches, the capacity of seabirds to adapt to climate change and the potential for interspecific competition in the region.

High-coverage genomes to elucidate the evolution of penguins.

BACKGROUND: Penguins (Sphenisciformes) are a remarkable order of flightless wing-propelled diving seabirds distributed widely across the southern hemisphere. They share a volant common ancestor with Procellariiformes close to the Cretaceous-Paleogene boundary (66 million years ago) and subsequently lost the ability to fly but enhanced their diving capabilities. With ∼20 species among 6 genera, penguins range from the tropical Galápagos Islands to the oceanic temperate forests of New Zealand, the rocky coastlines of the sub-Antarctic islands, and the sea ice around Antarctica. To inhabit such diverse and extreme environments, penguins evolved many physiological and morphological adaptations. However, they are also highly sensitive to climate change. Therefore, penguins provide an exciting target system for understanding the evolutionary processes of speciation, adaptation, and demography. Genomic data are an emerging resource for addressing questions about such processes. RESULTS: Here we present a novel dataset of 19 high-coverage genomes that, together with 2 previously published genomes, encompass all extant penguin species. We also present a well-supported phylogeny to clarify the relationships among penguins. In contrast to recent studies, our results demonstrate that the genus Aptenodytes is basal and sister to all other extant penguin genera, providing intriguing new insights into the adaptation of penguins to Antarctica. As such, our dataset provides a novel resource for understanding the evolutionary history of penguins as a clade, as well as the fine-scale relationships of individual penguin lineages. Against this background, we introduce a major consortium of international scientists dedicated to studying these genomes. Moreover, we highlight emerging issues regarding ensuring legal and respectful indigenous consultation, particularly for genomic data originating from New Zealand Taonga species. CONCLUSIONS: We believe that our dataset and project will be important for understanding evolution, increasing cultural heritage and guiding the conservation of this iconic southern hemisphere species assemblage.

Equitable Chick Survival in Three Species of the Non-Migratory Shorebird Despite Species-Specific Sexual Dimorphism of the Young.

Sex-biases in populations can have important implications for species' social biology, population demography and mating systems. It has recently been suggested that in some shorebirds, sex-specific bias in survival of precocial young may occur. This may be driven by variation in the brood sex-ratio and/or the sexual size dimorphism of young birds, which may influence predator escape capacity. Understanding the survival of young birds remains a significant knowledge gap for many taxa, especially when young birds are mobile and cryptic. Our aims were to estimate the sex-ratio variation in three species of Australian resident shorebird, specifically to determine: (1) whether seasonal brood sex-ratio variation at hatching is occurring, (2) the extent of any sex-biased chick survival, (3) if sex specific dimorphism at hatching or during growth occurs; and, (4) whether escape capacity differs between the sexes. We radio-tracked 50 Masked Lapwing Vanellus miles, 42 Red-capped Plover Charadrius ruficapillus and 27 Hooded Plover Thinornis cucullatus chicks from individual broods, examined the likelihood of hatchlings being male or female based on the hatching date within the breeding season, and compared size at hatching, growth and mortality of chicks of different sexes. There was no sex-bias with the hatching date across the breeding season, nor were there differences in survival or growth rates between sexes for any of the three species studied. In one species, male hatchlings had longer tarsi than females, but this did not result in differential escape propensity or improved survival. In conclusion, the hatching date, survival and growth of chicks from three species of resident shorebird was not influenced by their sex.

Understanding meta-population trends of the Australian fur seal, with insights for adaptive monitoring.

Effective ecosystem-based management requires estimates of abundance and population trends of species of interest. Trend analyses are often limited due to sparse or short-term abundance estimates for populations that can be logistically difficult to monitor over time. Therefore it is critical to assess regularly the quality of the metrics in long-term monitoring programs. For a monitoring program to provide meaningful data and remain relevant, it needs to incorporate technological improvements and the changing requirements of stakeholders, while maintaining the integrity of the data. In this paper we critically examine the monitoring program for the Australian fur seal (AFS) Arctocephalus pusillus doriferus as an example of an ad-hoc monitoring program that was co-ordinated across multiple stakeholders as a range-wide census of live pups in the Austral summers of 2002, 2007 and 2013. This 5-yearly census, combined with historic counts at individual sites, successfully tracked increasing population trends as signs of population recovery up to 2007. The 2013 census identified the first reduction in AFS pup numbers (14,248 live pups, -4.2% change per annum since 2007), however we have limited information to understand this change. We analyse the trends at breeding colonies and perform a power analysis to critically examine the reliability of those trends. We then assess the gaps in the monitoring program and discuss how we may transition this surveillance style program to an adaptive monitoring program than can evolve over time and achieve its goals. The census results are used for ecosystem-based modelling for fisheries management and emergency response planning. The ultimate goal for this program is to obtain the data we need with minimal cost, effort and impact on the fur seals. In conclusion we identify the importance of power analyses for interpreting trends, the value of regularly assessing long-term monitoring programs and proper design so that adaptive monitoring principles can be applied.

Penguin colony attendance under artificial lights for ecotourism.

Wildlife watching is an emerging ecotourism activity around the world. In Australia and New Zealand, night viewing of little penguins attracts hundreds of thousands of visitors per year. As penguins start coming ashore after sunset, artificial lighting is essential to allow visitors to view them in the dark. This alteration of the nightscape warrants investigation for any potential effects of artificial lighting on penguin behavior. We experimentally tested how penguins respond to different light wavelengths (colors) and intensities to examine effects on the colony attendance behavior at two sites on Phillip Island, Australia. At one site, nocturnal artificial illumination has been used for penguin viewing for decades, whereas at the other site, the only light is from the natural night sky. Light intensity did not affect colony attendance behaviors of penguins at the artificially lit site, probably due to penguin habituation to lights. At the not previously lit site, penguins preferred lit paths over dark paths to reach their nests. Thus, artificial light might enhance penguin vision at night and consequently it might reduce predation risk and energetic costs of locomotion through obstacle and path detection. Although penguins are faithful to their path, they can be drawn to artificial lights at small spatial scale, so light pollution could attract penguins to undesirable lit areas. When artificial lighting is required, we recommend keeping lighting as dim and time-restricted as possible to mitigate any negative effects on the behavior of penguins and their natural habitat.

Microbiota of little penguins and short-tailed shearwaters during development.

The establishment and early colonisation of the gastrointestinal (GI) tract has been recognised as a crucial stage in chick development, with pioneering microbial species responsible for influencing the development of the GI tract and influencing host health, fitness and disease status throughout life. Development of the microbiota in long lived seabirds is poorly understood. This study characterised the microbial composition of little penguin and short-tailed shearwater chicks throughout development, using Quantitative Real Time PCR (qPCR) and 16S rRNA sequencing. The results indicated that microbial development differed between the two seabird species with the short-tailed shearwater microbiota being relatively stable throughout development whilst significant fluctuations in the microbial composition and an upward trend in the abundance of Firmicutes and Bacteroidetes were observed in the little penguin. When the microbial composition of adults and chicks was compared, both species showed low similarity in microbial composition, indicating that the adult microbiota may have a negligible influence over the chick's microbiota.

Towards a quantitative indicator of feather disruption following the cleansing of oiled birds.

A computer-based imaging method for determining feather microstructure coherency following a cleansing treatment, was developed, calibrated and trialled on Mallard Duck (Anas platyrhyhchos) feathers. The feathers were initially contaminated with a light crude oil and then cleansed by either detergent (Deacon 90) treatment or, alternatively, by magnetic particle technology (MPT) using iron powder. The imaging method provides a single quantitative parameter for the coherence of feather microstructure and the results confirm that MPT treatment imparts less disruption to the feather microstructure than detergent treatment. It is proposed that this imaging method can be developed and implemented for the assessment of feather disruption and possibly damage, either for the trialling of different treatment protocols, or as a tool during the rehabilitation process, along with other such indicators, to give a more comprehensive assessment of feather condition than is currently available.

Seabird mortality induced by land-based artificial lights.

Artificial lights at night cause high mortality of seabirds, one of the most endangered groups of birds globally. Fledglings of burrow-nesting seabirds, and to a lesser extent adults, are attracted to and then grounded (i.e., forced to land) by lights when they fly at night. We reviewed the current state of knowledge of seabird attraction to light to identify information gaps and propose measures to address the problem. Although species in families such as Alcidae and Anatidae can be grounded by artificial light, the most affected seabirds are petrels and shearwaters (Procellariiformes). At least 56 species of Procellariiformes, more than one-third of them (24) threatened, are subject to grounding by lights. Seabirds grounded by lights have been found worldwide, mainly on oceanic islands but also at some continental locations. Petrel breeding grounds confined to formerly uninhabited islands are particularly at risk from light pollution due to tourism and urban sprawl. Where it is impractical to ban external lights, rescue programs of grounded birds offer the most immediate and employed mitigation to reduce the rate of light-induced mortality and save thousands of birds every year. These programs also provide useful information for seabird management. However, these data are typically fragmentary, biased, and uncertain and can lead to inaccurate impact estimates and poor understanding of the phenomenon of seabird attraction to lights. We believe the most urgently needed actions to mitigate and understand light-induced mortality of seabirds are estimation of mortality and effects on populations; determination of threshold light levels and safe distances from light sources; documentation of the fate of rescued birds; improvement of rescue campaigns, particularly in terms of increasing recovery rates and level of care; and research on seabird-friendly lights to reduce attraction.

Metals and metalloids in Little Penguin (Eudyptula minor) prey, blood and faeces.

Piscivorous species like the Little Penguin (Eudyptula minor) are particularly at risk of being negatively impacted by pollution due to their heightened exposure through aquatic food chains. Therefore, determining the concentration of heavy metals in the fish prey of seabirds is an essential component of assessing such risk. In this study, we report on arsenic, cadmium, mercury, lead and selenium concentrations in three fish species, which are known to comprise a substantial part of the diet of Little Penguins at the urban colony of St Kilda, Melbourne, Australia. Metal concentrations in the fish sampled were generally within the expected limits, however, arsenic and mercury were higher than reported elsewhere. Anchovy (Engraulis australis) and sandy sprat (Hyperlophus vittatus) contained higher Hg concentrations than pilchard (Sardinops sagax), while sandy sprat and pilchard contained more selenium. We present these findings together with metal concentrations in Little Penguin blood and faeces, sampled within weeks of the fish collection. Mercury concentrations were highest in the blood, while faeces and fish prey species contained similar concentrations of arsenic and lead, suggesting faeces as a primary route of detoxification for these elements. We also investigated paired blood - faecal samples and found a correlation for selenium only. Preliminary data from stable isotope ratios in penguin blood indicate that changes in penguin blood mercury concentrations cannot be explained by trophic changes in their diet alone, suggesting a variation of bioavailable Hg within this semi-enclosed bay.

Seasonal variation and annual trends of metals and metalloids in the blood of the Little Penguin (Eudyptula minor).

Little Penguins (Eudyptula minor) are high-trophic coastal feeders and are effective indicators of bioavailable pollutants in their foraging zones. Here, we present concentrations of metals and metalloids in blood of 157 Little Penguins, collected over three years and during three distinct seasons (breeding, moulting and non-breeding) at two locations: the urban St Kilda colony and the semi-rural colony at Phillip Island, Victoria, Australia. Penguin metal concentrations were foremostly influenced by location (St Kilda>Phillip Island for non-essential elements) and differed among years and seasons at both locations, reflecting differences in seasonal metal bioaccumulation or seasonal exposure through prey. Mean blood mercury concentrations showed an increasing annual trend and a negative correlation with flipper length at St Kilda. Notably, this study is the first to report on blood metal concentrations during the different stages of moult, showing the mechanism of non-essential metal mobilisation and detoxification.

Prior exposure to capture heightens the corticosterone and behavioural responses of little penguins (Eudyptula minor) to acute stress.

Studies of physiology can provide important insight into how animals are coping with challenges in their environment and can signal the potential effects of exposure to human activity in both the short and long term. In this study, we measured the physiological and behavioural response of little penguins (Eudyptula minor) that were naïve to human activity over 30 min of capture and handling. We assessed relationships between corticosterone secretion, behaviour, sex and time of day in order to characterize the determinants of the natural stress response. We then compared the response of these naïve penguins with the responses of female little penguins that had been exposed to research activity (bimonthly nest check and weighing) and to both research activity (monthly nest check and weighing) and evening viewing by tourists. We found that corticosterone concentrations increased significantly over 30 min of capture, with naïve penguins demonstrating a more acute stress response during the day than at night. Penguins that had previously been exposed to handling at the research and research/visitor sites showed elevated corticosterone concentrations and consistently more aggressive behaviour after 30 min compared with naïve birds, although there were no significant differences in baseline corticosterone concentrations. Our findings demonstrate that these little penguins have not habituated to routine capture, but rather mount a heightened physiological and behavioural response to handling by humans. Less invasive research monitoring techniques, such as individual identification with PIT tags and automatic recording and weighing, and a reduction in handling during the day should be considered to mitigate some of the potentially negative effects of disturbance. Given the paucity of data on the long-term consequences of heightened stress on animal physiology, our study highlights the need for further investigation of the relationship between the corticosterone stress response and fitness outcomes, such as breeding success and survival.

Associations between Resting, Activity, and Daily Metabolic Rate in Free-Living Endotherms: No Universal Rule in Birds and Mammals.

Energy management models provide theories and predictions for how animals manage their energy budgets within their energetic constraints, in terms of their resting metabolic rate (RMR) and daily energy expenditure (DEE). Thus, uncovering what associations exist between DEE and RMR is key to testing these models. Accordingly, there is considerable interest in the relationship between DEE and RMR at both inter- and intraspecific levels. Interpretation of the evidence for particular energy management models is enhanced by also considering the energy spent specifically on costly activities (activity energy expenditure [AEE] = DEE - RMR). However, to date there have been few intraspecific studies investigating such patterns. Our aim was to determine whether there is a generality of intraspecific relationships among RMR, DEE, and AEE using long-term data sets for bird and mammal species. For mammals, we use minimum heart rate (fH), mean fH, and activity fH as qualitative proxies for RMR, DEE, and AEE, respectively. For the birds, we take advantage of calibration equations to convert fH into rate of oxygen consumption in order to provide quantitative proxies for RMR, DEE, and AEE. For all 11 species, the DEE proxy was significantly positively correlated with the RMR proxy. There was also evidence of a significant positive correlation between AEE and RMR in all four mammal species but only in some of the bird species. Our results indicate there is no universal rule for birds and mammals governing the relationships among RMR, AEE, and DEE. Furthermore, they suggest that birds tend to have a different strategy for managing their energy budgets from those of mammals and that there are also differences in strategy between bird species. Future work in laboratory settings or highly controlled field settings can tease out the environmental and physiological processes contributing to variation in energy management strategies exhibited by different species.

Human residential status and habitat quality affect the likelihood but not the success of lapwing breeding in an urban matrix.

Wildlife living in the suburbs faces the challenge of dealing with human presence and yard management (including the occurrence of pets) which vary at the scale of the house block. This study examined the influence of ecological factors (e.g. extent of grass and food availability) and anthropogenic factors (e.g. human activity and garden usage) on breeding site choice and reproductive success of the ground-nesting masked lapwing Vanellus miles on Phillip Island, Australia. Lapwings nested less frequently in residential properties (high levels of human usage) compared with vacant blocks and holiday houses. They were also more likely to breed on properties with high food availability and larger areas of grass. None of these variables influenced clutch size or the probability of eggs hatching, although larger clutches and higher hatching rates tended to be associated with more food. This study shows that, for an urban exploiting species, habitat quality is not homogenous at the scale of the house block, and that human activity is avoided by a species generally considered highly tolerant of people.