Temporal Dynamics of Plasma Catecholamines, Metabolic and Immune Markers, and the Corticosterone:DHEA Ratio in Farmed Crocodiles before and after an Acute Stressor.

Commercial crocodilian farms face significant economic and livestock losses attributed to stress, which may be linked to their adopted husbandry practices. The development of appropriate and modernized husbandry guidelines, particularly those focused on stress mitigation, is impeded by the limited understanding of the crocodilian stress response. Fifteen grower Nile crocodiles were subjected to simulated acute transport stress, with blood samples collected at various intervals post-stress. Plasma levels of corticosterone (CORT), dehydroepiandrosterone (DHEA), adrenaline, and noradrenaline were determined using high-performance liquid chromatography. Glucose and lactate were measured using portable meters and the heterophil-to-lymphocyte ratio (HLR) was determined via differential leucocyte counts. Significant differences were elicited after the stressor, with acute fluctuations observed in the fast-acting catecholamines (adrenaline and noradrenaline) when compared to the baseline. Downstream effects of these catecholamines and CORT appear to be associated with a persistent increase in plasma glucose and HLR. Lactate also showed acute fluctuations over time but returned to the baseline by the final measurement. DHEA, which is used in a ratio with CORT, showed fluctuations over time with an inverted release pattern to the catecholamines. The study highlights the temporal dynamics of physiological markers under acute stress, contributing to our understanding of crocodilian stress and potentially informing improved farming practices for conservation and sustainable management.

Interconnections between the dorsal thalamus and the basal nuclei in a reptile.

Reciprocal connections between the thalamus and the cortex are one of the most characteristic features of forebrain organization in mammals. To date, this circuit has been documented only in turtles. However, reptiles, including turtles, have an additional path from the dorsal thalamus to the telencephalon. This terminates in a pallial structure known as the dorsal ventricular ridge. Yet, no reciprocal connection from the dorsal ventricular ridge to thalamic nuclei has been uncovered. Since axons from the thalamus pass through the basal nuclei on route to the dorsal ventricular ridge, the basal nuclei might be a source of reciprocal connections. Accordingly, the location and distribution of neurons after retrograde tracer placement into the dorsal thalamus were examined. Retrogradely labeled neurons in the basal nuclei were indeed found. One possibility to explain this observation is that connections with the dorsal ventricular ridge are present during development but later pruned during embryogenesis.

Exploring the most common lesion of Australian farmed saltwater crocodile (Crocodylus porosus) belly skin in the Northern Territory.

This is the first descriptive study to characterise and identify the most common lesions on harvested Australian saltwater crocodiles (Crocodylus porosus). 88 skins were examined over a 17-month period as part of normal farming practices, 2901 lesions identified, with scale location, location of the lesion on the scale, and characteristics (contour, keratin normality, translucency and colour) recorded. The study determined that linear lesions accounted for 68.25 % of lesions followed by foci lesions 17.24 %. Lesions were distributed on the upper proportion of the belly skin (77.8 %) and along the midline (72 %). The most common lesion identified was a single translucent linear lesion across the scale that otherwise appeared normal (58.95 %). While there is extensive research into pathogenic agents, further research is recommended to explore further causation of linear lesions, and factors that may contribute to their prevention. Given the subjective nature of crocodile skin grading, it is recommended future research into lesions is required to ensure the sustainability and profitability of the industry.

Hidden social complexity behind vocal and acoustic communication in non-avian reptiles.

Social interactions are inevitable in the lives of most animals, since most essential behaviours require interaction with conspecifics, such as mating and competing for resources. Non-avian reptiles are typically viewed as solitary animals that predominantly use their vision and olfaction to communicate with conspecifics. Nevertheless, in recent years, evidence is mounting that some reptiles can produce sounds and have the potential for acoustic communication. Reptiles that can produce sound have an additional communicative channel (in addition to visual/olfactory channels), which could suggest they have a higher communicative complexity, the evolution of which is assumed to be driven by the need of social interactions. Thus, acoustic reptiles may provide an opportunity to unveil the true social complexity of reptiles that are usually thought of as solitary. This review aims to reveal the hidden social interactions behind the use of sounds in non-avian reptiles. Our review suggests that the potential of vocal and acoustic communication and the complexity of social interactions may be underestimated in non-avian reptiles, and that acoustic reptiles may provide a great opportunity to uncover the coevolution between sociality and communication in non-avian reptiles. This article is part of the theme issue 'The power of sound: unravelling how acoustic communication shapes group dynamics'.

Pharmacokinetics of tildipirosin in estuarine (Crocodylus porosus) and freshwater (Crocodylus siamensis) crocodiles.

Tildipirosin is a macrolide antimicrobial. It is authorised for the treatment and prevention of respiratory disease in cattle and pigs. There are no data on its administration in crocodiles. Therefore, this study evaluated the disposition kinetics of tildipirosin after intravenous (dose: 2 mg/kg) and intramuscular (doses: 2 and 4 mg/kg) administration in two crocodilian species (estuarine and freshwater; n = 5). Tildipirosin plasma concentrations were quantified by a validated HPLC method. Plasma concentrations obtained at each extraction time were analysed by non-compartmental methods. In the estuarine and freshwater crocodiles, the apparent volumes of distribution of tildipirosin after intravenous administration were 0.36 ± 0.10 and 1.48 ± 0.26 L/kg, respectively. These values, suggesting poorer tissue distribution, were much lower than those obtained in mammals. There was complete bioavailability of tildipirosin after intramuscular route at a dose of 2 mg/kg; however, at a dose of 4 mg/kg the bioavailability decreased by about 20-25 %. Furthermore, the pharmacokinetics of tildipirosin were markedly different in the two crocodilian species. Considering a MIC of 0.5 µg/mL, the surrogate marker AUC0-24/MIC indicates that tildipirosin would greatly exceed the value of 65 h for both crocodile species and dose levels tested. This suggests that both doses (2 and 4 mg/kg) may provide a bactericidal effect. Therefore, based on the absence of adverse reactions following the administration of tildipirosin in both crocodilian species, and considering its favourable pharmacokinetic properties, tildipirosin may be useful in treating infections in these reptiles.

Detection of organochlorine pesticides in infertile eggs of crocodylus acutus from sinaloa.

Environmental contaminants endanger human health and non-target organisms such as crocodiles (Crocodylus acutus) that live in aquatic bodies surrounding agricultural areas. Due to their intrinsic characteristics, these organisms could be bioaccumulating and transmitting organochlorine pesticides (OCs) to their eggs. The objectives of this study were to determine the OCs in infertile eggs of C. acutus from Sinaloa and their correlation with the morphometric characteristics (MC), and to perform a preliminary estimate of the ecological risk due to the presence of pesticides using the PERPEST model. In June 2022, 76 infertile eggs (Ie) were collected: 57 from wild areas (Wa) and 19 from a crocodile farm (CSMf). Determination of OC in Ie was performed according to the USEPA method 8081b, modified. The observed percentages of Ie in Wa were 31.48% and 21.33% in CSMf. Twenty OCs were detected in the Ie, where dieldrin recorded the highest average concentration in Wa (6542.6 ng/g), and endosulfan-II in the CSMf (2172.8 ng/g). Bad negative and positive correlations were observed between OCs and MC, standing out the correlations between endosulfan-II and %Ie (-0.688) in the Wa, Cedritos drain, and between endrin and the weight of Ie (0.786) of the CSMf. The evaluation of the ecological risks of the aquatic environment due to the presence of OCs follow the sequence cyclodienes > aromatic > alicyclic hydrocarbons. A potential risk to the endocrine health of the species C. acutus was observed. Crocodiles are excellent biological models for monitoring the effects of OCs.

Pharmacokinetic disposition of marbofloxacin after intramuscular administration in estuarine crocodiles (Crocodylus porosus).

To date, the pharmacokinetics of fluoroquinolones in estuarine crocodiles (Crocodylus porosus) have been reported for enrofloxacin but not for marbofloxacin (MBF), which is a broad-spectrum antibiotic used only in veterinary medicine. This study investigated the pharmacokinetics of MBF after intramuscular administration at two difference dosages (2 and 4 mg/kg body weight) in estuarine crocodiles and estimated pharmacokinetic/pharmacodynamic (PK/PD) surrogate parameters for the optimization of dosage regimens. Ten treated estuarine crocodiles were divided into two groups (n = 5) using a randomization procedure according to a parallel study design. Blood samples were collected at assigned times up to 168 h. MBF plasma samples were cleaned up using liquid-liquid extraction and analyzed using a validated high-performance liquid chromatography method with fluorescence detection. A non-compartment approach was used to fit the plasma concentration of MBF vs time curve for each crocodile. The plasma concentrations of MBF were quantifiable for up to 168 h in both groups. The elimination half-life values of MBF were long (33.99 and 39.28 h for 2 and 4 mg/kg, respectively) with no significant differences between the groups. The average plasma protein binding of MBF was 30.85%. According to the surrogated PK/PD parameter (AUC0-24 -to-MIC ratio >100-125), the 2 and 4 mg/kg dosing rates should be effective for bacteria with MIC values lower than 0.125 µg/mL and 0.35 µg/mL, respectively.

The Various Ways in Which Birds Blink.

There has, to date, been no systematic study of the various ways in which birds blink. Digital video recordings were made, and studied using still frames, of 524 bird species, mainly in zoos but also in the wild. Videos on 106 species from various sites on the internet were studied, some of which we had also videoed, giving a total of 591 (out of a possible 10,000) species from all 43 orders and 125 (out of a possible 249) families. Digital video recordings were also made of 15 (out of a possible 24) species of crocodile. Three types of blink were observed in birds: (1) Nictitating membrane blinks were rapid and brief (phasic) and occurred mainly on head movement. (2) Upper lid blinks were seen in parrots, owls, pigeons and some others. These were also rapid and brief and accompanied nictitating membrane blinks. (3) Lower lid blinks were slow and sustained (tonic) and occurred with drowsiness and preening. Nictitating membrane blinks and lower lid blinks were seen in crocodiles but not upper lid blinks. Globe retraction, where the eyeball is pulled into the orbit of the skull during a blink, was seen in crocodiles but not birds. Phasic blinks remove debris and moisten the cornea, essential for allowing oxygen to diffuse into the cornea, which has no blood supply. Tonic blinks are probably mainly protective. The orders of birds which have upper lid blinking are not closely related and this feature is probably the result of convergent evolution.

Virulence genes identification in Salmonella enterica isolates from humans, crocodiles, and poultry farms from two regions in Colombia.

Background and Aim: Salmonella spp. is frequently found in the digestive tract of birds and reptiles and transmitted to humans through food. Salmonellosis is a public health problem because of pathogenicity variability in strains for virulence factors. This study aimed to identify the virulence genes in Salmonella isolates from humans, crocodiles, broiler cloacas, and broiler carcasses from two departments of Colombia. Materials and Methods: This study was conducted on 31 Salmonella enterica strains from humans with gastroenteritis (seven), crocodiles (seven), broiler cloacas (six), and broiler carcasses (12) from Tolima and Santander departments of Colombia, belonging to 21 serotypes. All samples were tested for Salmonella spp. using culture method on selective and non-selective mediums. Extraction of genomic DNA was performed from fresh colonies, DNA quality was verified by spectrophotometry and confirmed by amplification of InvA gene using conventional polymerase chain reaction (PCR). bapA, fimA, icmF, IroB, marT, mgtC, nlpI, oafA, pagN, siiD, spvC, spvR, spvB, Stn, and vexA genes were amplified by PCR. Results: The most prevalent gene was bapA (100%), followed by marT (96.77%), mgtC (93.55%), and fimA (83.87%). Likewise, IroB (70.97%), Stn (67.74%), spvR (61.29%), pagN (54.84%), icmF (54.8%), and SiiD (45.16%) were positive for more than 50% of the strains. Furthermore, none of the isolates tested positive for the vexA gene. Salmonella isolates presented 26 virulence profiles. Conclusion: This study reported 14 virulence genes in Salmonella spp. isolates from humans with gastroenteritis, crocodiles, and broiler cloacas and carcasses. The distribution of virulence genes differed among sources. This study could help in decision-making by health and sanitary authorities.

A cluster of leptospirosis cases associated with crocodile workers in the Northern Territory, Australia, 2022.

Leptospirosis is a worldwide zoonotic waterborne disease endemic in tropical and subtropical climates. Outbreaks have been observed in the Northern Territory (NT) of Australia. We briefly described the epidemiology of leptospirosis in the NT between 2012 and 2022, and undertook an investigation of a cluster of three leptospirosis cases observed in crocodile workers between January and December 2022 in the Top End of the NT. A descriptive case series was conducted to investigate the cluster; all three cases were male and non-Aboriginal with a median age of 46.5 years; none took chemoprophylaxis; only one of the three cases reported wearing appropriate protective attire; all reported receiving limited to no education about personal protective measures from their associated workplaces. Higher than average rainfall in both February and December 2022 likely contributed to the increased risk of infection in those months. Changing climate patterns are likely to result in more frequent periods of heavy rain, and risk of contracting leptospirosis in the NT may increase, particularly for those who work in wet and muddy conditions. Promoting the use of protective workplace clothing and equipment, the use of waterproof dressings for skin abrasions, regular hand hygiene, and the consideration of chemoprophylaxis in certain circumstances may prevent future cases.

Crocodile perception of distress in hominid baby cries.

It is generally argued that distress vocalizations, a common modality for alerting conspecifics across a wide range of terrestrial vertebrates, share acoustic features that allow heterospecific communication. Yet studies suggest that the acoustic traits used to decode distress may vary between species, leading to decoding errors. Here we found through playback experiments that Nile crocodiles are attracted to infant hominid cries (bonobo, chimpanzee and human), and that the intensity of crocodile response depends critically on a set of specific acoustic features (mainly deterministic chaos, harmonicity and spectral prominences). Our results suggest that crocodiles are sensitive to the degree of distress encoded in the vocalizations of phylogenetically very distant vertebrates. A comparison of these results with those obtained with human subjects confronted with the same stimuli further indicates that crocodiles and humans use different acoustic criteria to assess the distress encoded in infant cries. Interestingly, the acoustic features driving crocodile reaction are likely to be more reliable markers of distress than those used by humans. These results highlight that the acoustic features encoding information in vertebrate sound signals are not necessarily identical across species.

How do maternal androgens and estrogens affect sex determination in reptiles with temperature-dependent sex?

Temperature sex determination (TSD) in reptiles has been studied to elucidate the mechanisms by which temperature is transformed into a biological signal that determines the sex of the embryo. Temperature is thought to trigger signals that alter gene expression and hormone metabolism, which will determine the development of female or male gonads. In this review, we focus on collecting and discussing important and recent information on the role of maternal steroid hormones in sex determination in oviparous reptiles such as crocodiles, turtles, and lizards that possess TSD. In particular, we focus on maternal androgens and estrogens deposited in the egg yolk and their metabolites that could also influence the sex of offspring. Finally, we suggest guidelines for future research to help clarify the link between maternal steroid hormones and offspring sex.

A different framework to classify preoptic and hypothalamic nuclei in reptiles.

The preoptic area and the hypothalamus are inextricably linked. Together, they represent an area of the forebrain that is essential for survival of the species. Observations in mammals have suggested a classification of these structures into four rostrocaudal areas and three mediolateral zones. Two species of crocodiles were investigated to determine if this scheme or a modification of it could be applied to these reptiles. The resulting classification identified three rostrocaudal areas based on their respective relationship to the ventricular system: preoptic, anterior, and tuberal and four mediolateral zones: ependyma, periventricular, medial, and lateral. This scheme avoided the cumbersome and complicated nomenclature that has traditionally been used for morphologic studies of these areas in other reptiles, including crocodiles. The present classification is simple, straightforward, and readily applicable to other reptiles.

The ultrastructure of a stria vascularis in the auditory organ of the cuban crocodile (Crocodylus rhombifer).

Background: An endocochlear potential (EP) exists in the mammalian cochlea generated by the stria vascularis and an associated fibrocyte network. It plays an essential role for sensory cell function and hearing sensitivity. In non-mammalian ectothermic animals the endocochlear potential is low and its origin somewhat unclear. In this study, we explored the crocodilian auditory organ and describe the fine structure of a stria vascularis epithelium that has not been verified in birds. Material and Methods: Three Cuban crocodiles (Crocodylus rhombifer) were analyzed with light and transmission electron microscopy. The ears were fixed in glutaraldehyde The temporal bones were drilled out and decalcified. The ears were dehydrated, and embedded and was followed by semi-thin and thin sectioning. Results: The fine structure of the crocodile auditory organ including the papilla basilaris and endolymph system was outlined. The upper roof of the endolymph compartment was specialized into a Reissner membrane and tegmentum vasculosum. At the lateral limbus an organized, multilayered, vascularized epithelium or stria vascularis was identified. Discussion: Electron microscopy demonstrates that the auditory organ in Crocodylus rhombifer, unlike in birds, contains a stria vascularis epithelium separate from the tegmentum vasculosum. It is believed to secrete endolymph and to generate a low grade endocochlear potential. It may regulate endolymph composition and optimize hearing sensitivity alongside the tegmentum vasculosum. It could represent a parallel evolution essential for the adaptation of crocodiles to their diverse habitats.

Never smile at a crocodile: Gaping behaviour in the Nile crocodile at Ndumo Game Reserve, South Africa.

Gaping is a regularly observed behaviour in crocodilians globally but is still poorly understood in relation to external variables which could trigger this behaviour. The occurrence of gaping behaviour was investigated in a large wild population of Nile crocodiles (Crocodylus niloticus) during the dry and wet seasons at Nyamithi Pan, Ndumo Game Reserve, South Africa. This is one of the longest crocodile gaping behaviour studies conducted in the wild, with 300 h of observations conducted over two seasons resulting in 1120 gaping behaviours recorded. The most common size class observed was between 1.5 and 2.5 m (n = 697), which accounted for 62.2% of the gaping observations. A significant decrease in gaping duration was observed as ambient temperatures increased, and the gape duration was longer at a higher degree angle. In addition, an increase in gape duration was observed as the number of neighbouring crocodiles increased. These results suggest gaping behaviour occurs when there are other crocodiles nearby and when temperatures do not necessitate thermoregulation. The study suggests that gaping could be used as a form of thermoregulation and intra-species communication.

The challenge of compassion in predator conservation.

This paper argues that compassion for wild animals and the humans living alongside them should be integral to wildlife conservation. Nowhere is this more apparent than in predator conservation, and case studies are used to explore the consequences of wild animal attacks for human victims. Some arguments for extending compassionate consideration to animals seen as individuals are considered, along with the challenges these pose for predator conservation. A way forward from this apparent impasse is suggested, drawing on the capacity approach to embrace human with animal actors. The paper concludes with implications for predator conservation and recommendations, including incident responses sensitive to the traumatic impacts of attacks, and more collaborative approaches to handling human-wildlife interactions taking account of the capacities of local humans and wildlife.

Pharmacokinetic characteristics of azithromycin in freshwater crocodiles (Crocodylus siamensis) after intramuscular administration at three different dosages.

The study evaluated the pharmacokinetic features of azithromycin (AZM) in 15 freshwater crocodiles (Crocodylus siamensis) in Thailand. The crocodiles were administered a single intramuscular (i.m.) injection of AZM at three different dosages of 2.5, 5, and 10 mg/kg body weight (b.w.). Blood samples were collected at pre-assigned times up to 168 h. The plasma concentrations of AZM were measured using a validated liquid chromatography-tandem mass spectrometry method. The plasma concentration of AZM were quantifiable for up to 168 h after i.m. administration at the three different dosages. A non-compartmental model was used to fit the plasma concentration of AZM versus the time curve for each crocodile. The elimination half-life values of AZM were 33.70, 38.11, and 34.80 h following i.m. injection after dosages of 2.5, 5, and 10 mg/kg b.w., respectively. There were no significant differences among groups. The results indicated that the overall rate of elimination of AZM in freshwater crocodiles was relatively slow. The maximum concentration and area under the curve from zero to the last values of AZM increased in a dose-dependent fashion. The average binding percentage of AZM to plasma protein was 48.66%. Based on the pharmacokinetic data, the susceptibility break-point and the surrogate PK-PD index (T > MIC), the intramuscular administration of AZM at a dose of 10 mg/kg b.w. might be appropriate for the treatment of susceptible bacterial infections (MIC < 4 µg/ml) in freshwater crocodiles.

Pharmacokinetics of danofloxacin in freshwater crocodiles (Crocodylus siamensis) after intramuscular injection.

The present study evaluated the pharmacokinetic profiles of danofloxacin (DNX) in freshwater crocodiles (Crocodylus siamensis), following single intramuscular (i.m.) administrations at two different dosages of 6 and 12 mg/kg body weight (b.w.). Blood samples were collected at assigned times up to 168 h. DNX in the harvested crocodile plasma was extracted using liquid-liquid extraction and analyzed using a validated high-performance liquid chromatography method equipped with fluorescence detection. The pharmacokinetic analysis was performed using a non-compartmental approach. DNX in plasma was quantifiable from 5 min to 168 h after i.m. administration at the two dosages in freshwater crocodiles. The area under the curve (AUC) and maximum concentration (Cmax ) values increased in a dose-dependent fashion. Long elimination half-life (48.18 and 67.29 h) and low clearance (0.024 and 0.020 ml/g h) were obtained in the high- and low-dose groups, respectively. According to the pharmacokinetic-pharmacodynamic surrogate (AUC0-24h /MIC > 125), i.m. single administration of DNX at a dosage of 6 mg/kg b.w. is predicted to have antibacterial success for disease caused by bacteria with MIC < 0.04 µg/ml in the freshwater crocodile, C. siamensis.

Pharmacokinetic profiles of clarithromycin in freshwater crocodiles (Crocodylus siamensis).

Clarithromycin (CLA) is a new ß-lactamase-resistant macrolide antibiotic with potent activity against gram-positive and some gram-negative bacteria. To the authors' best knowledge, limited pharmacokinetic information to establish suitable therapeutic plans is available for freshwater crocodiles. To assess the prudent use of antibiotic in reptiles, this study was conducted to explore the pharmacokinetic characteristics of CLA in the freshwater crocodile, Crocodylus siamensis, following either single intravenous (i.v.) or intramuscular (i.m.) administration at a dosage of 2.5 mg/kg body weight (b.w.). Blood samples were collected at assigned times up to 168 h. CLA plasma samples were cleaned up using liquid-liquid extraction, and analysed by a validated liquid chromatography tandem-mass spectrometry (LC-MS/MS). CLA was quantifiable from 5 min to 72 h after i.v. administration, whereas it was detectable for 168 after i.m. administration at an identical dose rate. A non-compartmental model was used to fit the plasma concentration of CLA versus time curve for each crocodile. The t1/2λz value, similar for both routes (20 h), indicated that the overall rate of elimination of CLA in crocodiles is relatively slow. The average i.m. F% was complete. The protein plasma bound was found to be about 30%. CLA is a time-dependent antibiotic, and the T > MIC is the best PK/PD predictor for its efficacy. The CLA dosage of 2.5 mg/kg appeared to produce an appropriate value of the PK-PD surrogate that predicts antibacterial success for disease caused by susceptible bacteria.