Liasis Fuscus Research Articles

Gut microbiome-immune system interaction in reptiles.

Reptiles are ectothermic amniotes in a world dominated by endotherms. Reptiles originated more than 300million years ago and they often dwell in polluted environments which may expose them to pathogenic micro-organisms, radiation and/or heavy metals. Reptiles also possess greater longevity and may live much longer than similar-sized land mammals, for example, turtles, tortoises, crocodiles and tuatara are long-lived reptiles living up to 100years or more. Many recent studies have emphasized the pivotal role of the gut microbiome on its host; thus, we postulated that reptilian gut microbiome and/or its metabolites and the interplay with their robust immune system may contribute to their longevity and overall hardiness. Herein, we discuss the composition of the reptilian gut microbiome, immune system-gut microbiome cross-talk, antimicrobial peptides, reptilian resistance to infectious diseases and cancer, ageing, as well the current knowledge of the genome and epigenome of these remarkable species. Preliminary studies have demonstrated that microbial gut flora of reptiles such as crocodiles, tortoises, water monitor lizard and python exhibit remarkable anticancer and antibacterial properties, as well as comprise novel gut bacterial metabolites and antimicrobial peptides. The underlying mechanisms between the gut microbiome and the immune system may hold clues to developing new therapies overall for health, and possible extrapolation to exploit the ancient defence systems of reptiles for Homo sapiens benefit.

Anticancer Properties of Asian Water Monitor Lizard (Varanus salvator), Python (Malayopython reticulatus) and Tortoise (Cuora kamaroma amboinensis).

Cancer contributes to significant morbidity and mortality despite advances in treatment and supportive care. There is a need for the identification of effective anticancer agents. Reptiles such as tortoise, python, and water monitor lizards are exposed to heavy metals, tolerate high levels of radiation, feed on rotten/germ-infested feed, thrive in unsanitary habitat and yet have prolonged lifespans. Such species are rarely reported to develop cancer, suggesting the presence of anticancer molecules/mechanisms. Here, we tested effects from sera of Asian water monitor lizard (Varanus salvator), python (Malayopython reticulatus) and tortoise (Cuora kamaroma amboinensis) against cancer cells. Sera were collected and cytotoxicity assays were performed using prostate cancer cells (PC3), Henrietta Lacks cervical adenocarcinoma cells (HeLa) and human breast adenocarcinoma cells (MCF7), as well as human keratinized skin cells (Hacat), by measuring lactate dehydrogenase release as an indicator for cell death. Growth inhibition assays were performed to determine the effects on cancer cell proliferation. Liquid chromatography mass spectrometry was performed for molecular identification. The findings revealed that reptilian sera, but not bovine serum, abolished viability of Hela, PC3 and MCF7 cells. Samples were subjected to liquid chromatography mass spectrometry, which detected 57 molecules from V. salvator, 81 molecules from Malayopython reticulatus and 33 molecules from C. kamaroma amboinensis and putatively identified 9 molecules from V. salvator, 20 molecules from Malayopython reticulatus and 9 molecules from C. kamaroma amboinensis when matched against METLIN database. Based on peptide amino acid composition, binary profile, dipeptide composition and pseudo-amino acid composition, 123 potential Anticancer Peptides (ACPs) were identified from 883 peptides from V. salvator, 306 potential ACPs from 1074 peptides from Malayopython reticulatus and 235 potential ACPs from 885 peptides from C. kamaroma amboinensis. To our knowledge, for the first time, we reported comprehensive analyses of selected reptiles' sera using liquid chromatography mass spectrometry, leading to the identification of potentially novel anticancer agents. We hope that the discovery of molecules from these animals will pave the way for the rational development of new anticancer agents.

Dehydration enhances innate immunity in a semiaquatic snake from the wet-dry tropics.

Dehydration is considered a physiological challenge, and many organisms live in environments that undergo periods of reduced water availability that can lead to dehydration. Recent studies have found a positive relationship between dehydration and innate immune function in animals adapted to xeric or semixeric environments. To explore the generality of this relationship, we examined the impact of dehydration on innate immune performance in water pythons (Liasis fuscus), a semiaquatic snake from the wet-dry tropics of Australia. We collected blood samples from male and female water pythons held in the laboratory without food and water for 4 weeks. We also collected blood from free-ranging snakes throughout the Austral dry-season. We evaluated plasma osmolality and innate immune function (agglutination, lysis, and bacterial-killing ability) and found that increased osmolality, whether manipulated in the laboratory or as a result of natural water limitation, resulted in enhanced aspects of innate immune performance. Counter-intuitively, snakes in the wild became more hydrated as the dry season progressed, suggesting the dehydrated snakes move to water sources periodically to rehydrate. Comparing our data with those from previous studies, we suspect species divergence in the level of dehydration (i.e., hyperosmolality) that triggers enhanced immune capabilities.

Floods and famine: climate‐induced collapse of a tropical predator‐prey community

Summary Will climate change threaten wildlife populations by gradual shifts in mean conditions, or by increased frequency of extreme weather events? Based on long‐term data (from 1991 to 2014), the aim of the present study was to analyse and compare the sensitivity of predator–prey demography to extreme climatic events versus normal, albeit highly variable, annual deviations in climatic conditions in the Australian wet–dry tropics. From 1991 to 2005, predators (water pythons, Liasis fuscus) and their main prey (dusky rats, Rattus colletti) showed significant climate‐driven fluctuations in numbers. These fluctuations were, however, trivial compared to the impact of two massive but brief deluges in 2007 and 2011, which virtually eliminated the dusky rats. The two floods resulted in the pythons experiencing an unprecedented famine in seven out of the last 8 years causing a massive shift in python demography, that is a significant reduction in feeding rates, reproductive output, growth rates, relative body mass, survival, mean body length and numbers (from 3173 in 1992 to 96 in 2013). Our results demonstrate that attempts to predict faunal responses to climate change, even if based on long‐term studies, may be doomed to failure. Consequently, biologists may need to confront the uncomfortable truth that increased frequency of brief unpredictable bouts of extreme weather can influence populations far more than gradual deviations in mean climatic conditions.

The endoparasites of Liasis fuscus (Serpentes : Boidae) from the Adelaide River floodplain, Northern Territory, Australia

This study documents the parasite assemblage of Liasis fuscus Peters, 1873 from the Adelaide River flood plain, Northern Territory, Australia. In total, nine species of helminth (comprising three cestodes and six nematodes), one pentastome and one protozoan were found in 180 water pythons, with the Nematoda being the dominant and most diverse group, and a cestode, Bothridium ornatum Maplestone & Southwell, 1923, the most prevalent species. In spite of the hosts being present in high numbers the helminth assemblage was depauperate, characterised by low prevalence, intensity, frequency and abundance of species, with neither season nor sex of host affecting abundance. There were, however, significant differences between ages of hosts, juveniles having fewer parasites, and seasonal diversity, with the wet season of 2004 and the dry of 2005 the most similar. Of the Australian python species analysed Morelia spilota (Lacépède, 1804) had the highest species richness. The lower species richness found in L. fuscus was possibly due to its unusual biology. Biogeographic relationships of the parasite fauna of L. fuscus are diverse, connections with Africa, Asia, and South America being noted.

Mortality as a bivariate function of age and size in indeterminate growers

Mortality in organisms that grow indefinitely, known as indeterminate growers, is thought to be driven primarily by size. However, a number of ageing mechanisms also act as functions of age. Thus, to explain mortality in these species, both size and age need to be explicitly modeled. Here we developed a model that treats age‐ and size‐specific mortality as a bivariate process. This method facilitates the exploration of the underlying (unobserved) contributions of age and size to mortality. We show that, in theory, a population can show declining mortality with age and size while the underlying contribution of age, as a proxy for chronological deterioration, is of typical senescence; while a seemingly senescent population can have underlying age‐related negative senescence, which is, however, overcome by negative underlying size effects (i.e., increasing size‐specific mortality). We show how inference about these unobserved processes can be made using a simple Bayesian model, and how all of the mortality parameters are accurately retrieved. We then apply the methods to published datasets on water pythons and freshwater mussels and test different hypotheses regarding the effects of age and size on mortality. In both cases we found age‐dependent senescent mortality, with size having only negligible effects. The methods we present here can help to improve the demographic models commonly applied to a vast number of species of commercial and conservation importance such as fish, trees or bivalves, among many other, for which both age and size are relevant. In addition, the application of these methods can help to shed light on the existence of processes such as negative senescence, and contribute to our understanding of the evolutionary mechanisms of senescence in species that do not fit the established theories.

The consequences of alternative parental care tactics in free‐ranging pythons in tropical Australia

Summary1.Life‐history theory attributes the evolution of parental care to the benefits to offspring viability outweighing any costs to parental viability. However, the consequences of parental care tactics to parent(s) and the developmental environment have seldom been measured under field conditions.2.Laboratory research on pythons shows that maternal nest‐site selection and egg brooding benefit embryos, but prolonged nest attendance may impose fitness costs to free‐ranging females. A population of water pythons (Liasis fuscus) in tropical Australia provides an excellent opportunity to examine this parent–offspring trade‐off because females exhibit parental care polymorphism wherein some individuals brood their eggs only briefly (<10 days) post‐oviposition (‘short brooders’) while others remain with their eggs throughout the incubation period (>50 days; ‘long brooders’).3.We used radiotelemetry, temperature and humidity data loggers, ultrasonography, haematological techniques, and habitat analyses to examine the correlates and consequences of maternal nesting decisions in 14 free‐ranging female pythons over the 4‐month reproductive season.4.Nest‐site selection and maternal attendance enhanced thermal and hydric regimes within the nest. Egg production by reproducing female pythons resulted in high energetic costs (loss of 60% of maternal body mass) and increased parasite load. However, the estimated mass loss because of brooding was (i) low (<5%), (ii) inversely related to fecundity (females that produced relatively large clutches tended to select lower temperatures and thus lost less mass during brooding) and (iii) surprisingly unrelated to brooding duration. Phenotypic traits of short and long brooders were similar, but long brooders had higher haemoparasite burdens prior to oviposition. Clutches of long brooders were laid in more open sites (less canopy cover) and experienced warmer and more humid conditions than did those of short brooders.5.Together with previous research, we suggest several explanations for the maintenance of maternal care polymorphism within this population, such as a trade‐off between offspring number and quality (long brooders may produce fewer clutches during their lifespan but enhance offspring quality). Our study provides detailed measurements of the correlates and consequences of parental care in a free‐ranging reptile, and it clarifies the trade‐offs mediated by taxonomically widespread maternal decisions (e.g. nest‐ or oviposition‐site selection and nest attendance).

Temporal and Spatial Complexity of Maternal Thermoregulation in Tropical Pythons

Parental care is a widespread adaptation that evolved independently in a broad range of taxa. Although the dynamics by which two parents meet the developmental needs of offspring are well studied in birds, we lack understanding about the temporal and spatial complexity of parental care in taxa exhibiting female-only care, the predominant mode of parental care. Thus, we examined the behavioral and physiological mechanisms by which female water pythons Liasis fuscus meet a widespread developmental need (thermoregulation) in a natural setting. Although female L. fuscus were not facultatively thermogenic, they did use behaviors on multiple spatial scales (e.g., shifts in egg-brooding postures and surface activity patterns) to balance the thermal needs of their offspring throughout reproduction (gravidity and egg brooding). Maternal behaviors in L. fuscus varied by stage within reproduction and were mediated by interindividual variation in body size and fecundity. Female pythons with relatively larger clutch sizes were cooler during egg brooding, suggesting a trade-off between reproductive quantity (size of clutch) and quality (developmental temperature). In nature, caregiving parents of all taxa must navigate both extrinsic factors (temporal and spatial complexity) and intrinsic factors (body size and fecundity) to meet the needs of their offspring. Our study used a comprehensive approach that can be used as a general template for future research examining the dynamics by which parents meet other developmental needs (e.g., predation risk or energy balance).

Climate-induced reaction norms for life-history traits in pythons

Climate change modelers predict increasingly frequent "extreme events," so it is critical to quantify whether organismal responses (such as reproductive output) measured over the range of usual climatic conditions can predict responses under more extreme conditions. In a 20-year field study on water pythons (Liasis fuscus), we quantified the effects of climatically driven annual variation in food supply on demographic traits of female pythons (feeding rate, body size, body mass, and reproductive output). Reaction norms linking food supply to feeding rates and residual body mass were broadly linear, whereas norms linking food supply to female body size became curvilinear when a dramatic (flooding-induced) famine reduced the mean body size at sexual maturity. Thus, the reaction norms recorded over 16 years of mal" (albeit highly variable) climatic conditions gave little insight into the population's response to a more extreme nutritional crisis.

The use of ultrasonography to assess reproductive investment and output in pythons

Reproductive investment and output are integral fitness components, often incorporated into life-history trade-off models and important to population dynamics. The trade-offs associated with reproduction can be dramatic in species such as snakes that make especially large investments into reproduction. Unfortunately, traditional methods used to determine reproductive investment and output are effective in many (but not all) situations. Thus, we used portable ultrasonography to serially estimate reproductive investment and reproductive output in three python species that exhibit significant variation in phylogeny, geographic range, body size, egg size, and clutch size: ball pythons (Python regius), Children's pythons (Antaresia childreni), and water pythons (Liasis fuscus). At each time point of measurement (range: 1–49 days pre-oviposition), ultrasound estimates of viable clutch size were highly accurate in all three species. However, ultrasound estimates of mean viable egg mass, and thus viable clutch mass, significantly differed from the actual values (range: 23–73% error). Interestingly, this error was considerably smaller as females approached oviposition, suggesting that female pythons transfer a significant amount of water into their eggs during the week before oviposition. Thus, water balance during late-stage egg development may be an integral part of reproductive success. The results obtained in the present study form the foundation for future assessments of reproductive investment, and also provide insight into the use of ultrasound technology to assist such efforts.

Do natural antibodies compensate for humoral immunosenescence in tropical pythons?

1. Ageing is frequently claimed to result in an age-dependent deterioration in immune function. 2. Our results, however, based on 30 known-aged water pythons (Liasis fuscus; age ranging from <1 to 18 years) immunized with keyhole limpet haemocyanin (KLH), demonstrate that age-related changes in immune function may follow different and opposing pathways. 3. Python-specific humoral antibodies (SpAb) showed an age-dependent decrease in cross-reactivity to KLH, whereas natural antibodies (NAbs) ability to bind to this antigen increased with age. 4. Notably, when humoral SpAb and NAb titres were combined, no effect of age was detected in antibody cross-reactivity, strongly suggesting that NAbs may play a crucial role in maintaining immunocompetence during the ageing process.

COST OF MULTIPLE MATINGS IN FEMALE ADDERS (VIPERA BERUS)

Female multiple mating with different partners (polyandry) is taxonomically widespread and appears to be the very prevalent among squamate reptiles (Wusterbarth et al. 2010, and references therein). In this group of animals, females who mate multiply do not gain any direct benefits from the males but may benefit from increased offspring viability as has been observed in some squamates such as adders (Vipera berus, Madsen et al. 1992), water pythons (Liasis fuscus, Madsen et al. 2005), and sand lizards (Lacerta agilis, Olsson et al. 1994). In a recent publication such fitness benefits were again demonstrated in the latter taxon (Olsson et al. 2011). However, in spite of this, the authors stated that “the widespread within‐clutch multiple paternity in reptiles are unlikely to be explained by these effects. A more parsimonious explanation to multiple paternity would be relatively low costs to females for mating multiply and high gains for males.” Although I am one of the authors of this publication, I do not agree with this explanation.

How well do predators adjust to climate-mediated shifts in prey distribution? A study on Australian water pythons

Climate change can move the spatial location of resources critical for population viability, and a species' resilience to such changes will depend upon its ability to flexibly shift its activities away from no-longer-suitable sites to exploit new opportunities. Intuition suggests that vagile predators should be able to track spatial shifts in prey availability, but our data on water pythons (Liasisfuscus) in tropical Australia suggest a less encouraging scenario. These pythons undergo regular long-range (to >10 kmin) seasonal migrations to follow flooding-induced migrations by their prey (native dusky rats, Rattus colletti). However, when an extreme flooding event virtually eliminated rats for a three-year period, the local pythons did not disperse despite the presence of abundant rats only 8 km away; instead, many pythons starved to death. This inflexibility suggests that some vagile species that track seasonally migrating prey may do so by responding to habitat attributes that have consistently predicted prey availability over evolutionary time, rather than reacting to proximate cues that signal the presence of prey per se. A species' vulnerability to climate change will be increased by an inability to shift its activities away from historical sites toward newly favorable areas.

Body condition and head size in snakes

AbstractOn several occasions during my many years of working with various snake species, I observed a significant positive relationship between body condition and head size (the former calculated from residual scores from a general linear regression of ln-transformed mass on ln-transformed snout-vent length). Based on results from my long-term study of water pythons (Liasis fuscus) this relationship is, however, most likely caused by a condition-dependent bias when recording snake snout-vent length (SVL). Water pythons in good condition were recorded as being "shorter" and hence having relatively larger head size, whereas snakes in poor condition were recorded as being "longer" with concomitant smaller head relative head size. Such a systematic bias may lead to spurious conclusions concerning the adaptive significance of the relationship between snake body condition and head size.

Non-homologous sex chromosomes of birds and snakes share repetitive sequences

Snake sex chromosomes provided Susumo Ohno with the material on which he based his theory of how sex chromosomes differentiate from autosomal pairs. Like birds, snakes have a ZZ male/ZW female sex chromosome system, in which the snake Z is a macrochromosome much the same size as the bird Z. However, the gene content shows clearly that the snake and bird Z chromosomes are completely non-homologous. The molecular aspect of W chromosome degeneration in snakes remains largely unexplored. We used comparative genomic hybridization to identify the female-specific region of the W chromosome in representative species of Australian snakes. Using this approach, we show that an increasingly complex suite of repeats accompanies the evolution of W chromosome heteromorphy. In particular, we found that while the python Liasis fuscus exhibits no sex-specific repeats and indeed, no cytologically recognizable sex-specific region, the colubrid Stegonotus cucullatus shows a large domain on the short arm of the W chromosome that consists of female-specific repeats, and the large W of Notechis scutatus is composed almost entirely of repetitive sequences, including Bkm and 18S rDNA-related elements. FISH mapping of both simple and complex probes shows patterns of repeat amplification concordant with the size of the female-specific region in each species examined. Mapping of intronic sequences of genes that are sex-linked in both birds (DMRT1) and snakes (CTNNB1) reveals massive amplification in discrete domains on the W chromosome of the elapid N. scutatus. Using chicken W chromosome paint, we demonstrate that repetitive sequences are shared between the sex chromosomes of birds and derived snakes. This could be explained by ancestral but as yet undetected shared synteny of bird and snake sex chromosomes or may indicate functional homology of the repeats and suggests that degeneration is a convergent property of sex chromosome evolution. We also establish that synteny of snake Z-linked genes has been conserved for at least 166 million years and that the snake Z consists of two conserved blocks derived from the same ancestral vertebrate chromosome.

Molecular characterization of Hepatozoon species in a Leopard Cat (Prionailurus bengalensis) from Thailand

Hepatozoon gamonts were observed by light microscopy in neutrophils of a male, wild-caught Leopard Cat. Complete blood counts at presentation and 6 months later were unremarkable. Serologic tests were negative for both FIV and FeLV. A partial sequence of the 18S rRNA gene from the Hepatozoon found in the cat indicated that, compared with all species examined, the protozoan had the closest relationship (99.2% sequence similarity) with the Hepatozoon of the water python (Stegonotus cucullatus). The cat was clinically healthy at last report. Although Hepatozoon has been found in another wild cat in Thailand, this is the first report in a Leopard Cat. The pathogenicity of Hepatozoon in these cats remains uncertain.

Short Telomeres in Hatchling Snakes: Erythrocyte Telomere Dynamics and Longevity in Tropical Pythons

BackgroundTelomere length (TL) has been found to be associated with life span in birds and humans. However, other studies have demonstrated that TL does not affect survival among old humans. Furthermore, replicative senescence has been shown to be induced by changes in the protected status of the telomeres rather than the loss of TL. In the present study we explore whether age- and sex-specific telomere dynamics affect life span in a long-lived snake, the water python (Liasis fuscus).Methodology/Principal FindingsErythrocyte TL was measured using the Telo TAGGG TL Assay Kit (Roche). In contrast to other vertebrates, TL of hatchling pythons was significantly shorter than that of older snakes. However, during their first year of life hatchling TL increased substantially. While TL of older snakes decreased with age, we did not observe any correlation between TL and age in cross-sectional sampling. In older snakes, female TL was longer than that of males. When using recapture as a proxy for survival, our results do not support that longer telomeres resulted in an increased water python survival/longevity.Conclusions/SignificanceIn fish high telomerase activity has been observed in somatic cells exhibiting high proliferation rates. Hatchling pythons show similar high somatic cell proliferation rates. Thus, the increase in TL of this group may have been caused by increased telomerase activity. In older humans female TL is longer than that of males. This has been suggested to be caused by high estrogen levels that stimulate increased telomerase activity. Thus, high estrogen levels may also have caused the longer telomeres in female pythons. The lack of correlation between TL and age among old snakes and the fact that longer telomeres did not appear to affect python survival do not support that erythrocyte telomere dynamics has a major impact on water python longevity.

Molecular and Morphological Description of a Hepatozoon Species in Reptiles and Their Ticks in the Northern Territory, Australia

Ticks, representing 3 species of Amblyomma, were collected from the water python (Liasis fuscus) and 3 additional reptile species in the Northern Territory, Australia, and tested for the presence of Hepatozoon sp., the most common blood parasites of snakes. In addition, blood smears were collected from 5 reptiles, including the water python, and examined for the presence of the parasite. Hepatozoon sp. DNA was detected in all tick and reptile species, with 57.7% of tick samples (n = 187) and 35.6% of blood smears (n=35) showing evidence of infection. Phylogenetic analysis of the 18S rRNA gene demonstrated that half of the sequences obtained from positive tick samples matched closest with a Hepatozoon species previously identified in the water python population. The remaining sequences were found to be more closely related to mammalian and amphibian Hepatozoon species. This study confirms that species of Amblyomma harbor DNA of the same Hepatozoon species detected in the water pythons. The detection of an additional genotype suggests the ticks may be exposed to 2 Hepatozoon species, providing further opportunity to study multiple host-vector-parasite relationships.

Spatial ecology of hatchling water pythons (Liasis fuscus) in tropical Australia

Abstract:Young snakes are rarely seen in the field and little is known about their habits, mostly because they are too small for radio-telemetry (the primary method for studying snake spatial ecology). However, the offspring of some larger species can be fitted with transmitters and we investigated the spatial ecology and habitat use of ten hatchling water pythons (Liasis fuscus: Pythonidae) in the floodplain of the Adelaide River, tropical Australia. Patterns of habitat use in the late wet season and during the dry season were similar to those of adults tracked in the same vicinity in an earlier study. Soon after release the young snakes moved to the floodplain, avoiding pasture areas. Diurnal refuge sites were typically in the base of grass clumps or below the soil surface, especially in sites with thick vegetation and deep, wide soil cracks. Adult snakes are more sedentary but move longer absolute distances (mean ± SE = 252 ± 50 m wk−1) than hatchlings (66.3 ± 41 m wk−1). However, hatchling snakes moved longer distances relative to body size (84.4 ± 1.1 body lengths wk−1) than did the previously studied adults (66.0 ± 1.1 body lengths wk−1). Mean and minimum body temperatures of the hatchlings were correlated with mean and minimum air temperatures, respectively.

On parsimonious paternity and scientific rigor: a reply to Madsen

On parsimonious paternity and scientific rigor: a reply to Madsen