Glutathione-Mediated Metal Tolerance in an Amphibian Chytrid Fungus (Batrachochytrium dendrobatidis).

The spread of the amphibian chytrid fungus Batrachochytrium dendrobatidis, which causes the disease chytridiomycosis, has resulted in amphibian declines and extinctions worldwide. Some susceptible amphibian species can persist in contaminated habitats, prompting the hypothesis that B. dendrobatidis might be sensitive to heavy metals. We tested a panel of 12 metals to rank their toxicity to B. dendrobatidis zoospores and zoosporangia during a 6-h exposure. To better understand the mechanism for metal detoxification, we also evaluated whether glutathione is required for metal tolerance by depleting cellular glutathione before metal exposure. In addition, we investigated whether prior exposure to low metal concentrations impacted tolerance of subsequent exposure, as well as identifying metal combinations that may act synergistically. Silver (Ag), cadmium (Cd), and copper (Cu) were particularly toxic to B. dendrobatidis, with zoospore minimum lethal concentration values of 0.01 mM (Ag), 0.025 mM (Cd), and 0.5 mM (Cu). These three metals along with zinc (Zn) were also inhibitory to zoosporangia, with minimum inhibitory concentration values of 0.005 mM (Ag), 0.04 mM (Cd), 0.075 mM (Cu), and 0.04 mM (Zn). The fungicidal effects of several metals was reduced when assays were conducted in nutrient medium compared with synthetic pond water, highlighting the need for careful in vitro assay design and interpretation. Glutathione depletion strongly influenced tolerance of Cd and Ag (85% and 75% less growth, respectively) and moderately influenced tolerance of Cu, Zn, and lead (37%, 18%, and 14% less growth, respectively), indicating the importance of glutathione for metal detoxification. In general, the minimum metal concentrations that inhibited growth of B. dendrobatidis far exceeded values detected in contaminated amphibian habitats in Australia, suggesting that metal contamination alone may not have a strong protective effect against chytridiomycosis. We discuss future research directions to futher understand the potential for dissolved metals to create chytrid refuges. Environ Toxicol Chem 2024;00:1-9. © 2024 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

An examination of Homo naledi early juveniles recovered from the Rising Star cave system, South Africa.

BACKGROUND: Six Homo naledi early juveniles were recovered from U.W. 101 (Dinaledi Chamber), U.W. 102 (Lesedi Chamber), and U.W. 110 in the Rising Star cave system. AIM: This paper develops the information for the H. naledi early juvenile life stage, as defined by a combination of deciduous and permanent dentition, and the eruption of the first permanent molar. SUBJECTS AND METHODS: The growing number of young individuals recovered from the Rising Star cave system allows us to gain a better understanding of their variation, or lack thereof, and provides a basis to estimate broad ranges for age at death of the individuals. The individuals are identified and described through craniodental remains and spatial associations. RESULTS AND CONCLUSION: Our results show that the teeth are remarkably consistent across the localities in their metric and non-metric traits, and our analyses refine previous estimations on dental eruptions with the first permanent molar erupting first in the sequence among permanent teeth.

Advances in Managing Chytridiomycosis for Australian Frogs: Gradarius Firmus Victoria.

Extensive knowledge gains from research worldwide over the 25 years since the discovery of chytridiomycosis can be used for improved management. Strategies that have saved populations in the short term and/or enabled recovery include captive breeding, translocation into disease refugia, translocation from resistant populations, disease-free exclosures, and preservation of disease refuges with connectivity to previous habitat, while antifungal treatments have reduced mortality rates in the wild. Increasing host resistance is the goal of many strategies under development, including vaccination and targeted genetic interventions. Pathogen-directed strategies may be more challenging but would have broad applicability. While the search for the silver bullet solution continues, we should value targeted local interventions that stop extinction and buy time for evolution of resistance or development of novel solutions. As for most invasive species and infectious diseases, we need to accept that ongoing management is necessary. For species continuing to decline, proactive deployment and assessment of promising interventions are more valid than a hands-off, do-no-harm approach that will likely allow further extinctions.

Human-like enamel growth in Homo naledi.

OBJECTIVES: A modern pattern (rate and duration) of dental development occurs relatively recently during human evolution. Given the temporal overlap of Homo naledi with the first appearance of fossil Homo sapiens in Africa, this small-bodied and small-brained hominin presents an opportunity to elucidate the evolution of enamel growth in the hominin clade. Here we conduct the first histological study of two permanent mandibular canines and one permanent maxillary first molar, representing three individuals attributed to H. naledi. We reconstruct the rate and duration of enamel growth and compare these findings to those reported for other fossil hominins and recent humans. MATERIALS AND METHODS: Thin sections of each tooth were produced using standard histological methods. Daily and longer period incremental markings were measured to reconstruct enamel secretion and extension rates, Retzius periodicity, canine crown and molar cusp formation time. RESULTS: Daily enamel secretion rates overlapped with those from recent hominins. Canine crown formation time is similar to that observed in recent Europeans but is longer than canine formation times reported for most other hominins including Australopithecus and H. neanderthalensis. The extended period of canine formation appears to be due to a relatively tall enamel crown and a sustained slow rate of enamel extension in the cervical portion of the crown. A Retzius periodicity of 11 days for the canines, and nine days for the molar, in H. naledi parallel results found in recent humans. An 11-day periodicity has not been reported for Late Pleistocene Homo (H. erectus, H. neanderthalensis) and is rarely found in Australopithecus and Paranthropus species. DISCUSSION: Enamel growth of H. naledi is most similar to recent humans though comparative data are limited for most fossil hominin species. The high Retzius periodicity values do not follow expectations for a small-brained hominin.

Sex-biased sampling may influence Homo naledi tooth size variation.

A frequent source of debate in paleoanthropology concerns the taxonomic unity of fossil assemblages, with many hominin samples exhibiting elevated levels of variation that can be interpreted as indicating the presence of multiple species. By contrast, the large assemblage of hominin fossils from the Rising Star cave system, assigned to Homo naledi, exhibits a remarkably low degree of variation for most skeletal elements. Many factors can contribute to low sample variation, including genetic drift, strong natural selection, biased sex ratios, and sampling of closely related individuals. In this study, we tested for potential sex-biased sampling in the Rising Star dental sample. We compared coefficients of variation for the H. naledi teeth to those for eight extant hominoid samples. We used a resampling procedure that generated samples from the extant taxa that matched the sample size of the fossil sample for each possible Rising Star dental sex ratio. We found that variation at four H. naledi tooth positions-I2, M1, P4, M1-is so low that the possibility that one sex is represented by few or no individuals in the sample cannot be excluded. Additional evidence is needed to corroborate this inference, such as ancient DNA or enamel proteome data, and our study design does not address other potential factors that would account for low sample variation. Nevertheless, our results highlight the importance of considering the taphonomic history of a hominin assemblage and suggest that sex-biased sampling is a plausible explanation for the low level of phenotypic variation found in some aspects of the current H. naledi assemblage.

Glutathione is required for growth and cadmium tolerance in the amphibian chytrid fungus, Batrachochytrium dendrobatidis.

Batrachochytrium dendrobatidis (Bd) is a lethal amphibian pathogen, partly due to its ability to evade the immune system of susceptible frog species. In many pathogenic fungi, the antioxidant glutathione is a virulence factor that helps neutralise oxidative stressors generated from host immune cells, as well as other environmental stressors such as heavy metals. The role of glutathione in stress tolerance in Bd has not been investigated. Here, we examine the changes in the glutathione pool after stress exposure and quantify the effect of glutathione depletion on cell growth and stress tolerance. Depletion of glutathione repressed growth and release of zoospores, suggesting that glutathione is essential for life cycle completion in Bd. Supplementation with <2 mM exogenous glutathione accelerated zoospore development, but concentrations >2 mM were strongly inhibitory to Bd cells. While hydrogen peroxide exposure lowered the total cellular glutathione levels by 42 %, glutathione depletion did not increase the sensitivity to hydrogen peroxide. Exposure to cadmium increased total cellular glutathione levels by 93 %. Glutathione-depleted cells were more sensitive to cadmium, and this effect was attenuated by glutathione supplementation, suggesting that glutathione plays an important role in cadmium tolerance. The effects of heat and salt were exacerbated by the addition of exogenous glutathione. The impact of glutathione levels on Bd stress sensitivity may help explain differences in host susceptibility to chytridiomycosis and may provide opportunities for synergistic therapeutics.

The prevalence of rodent-borne zoonotic pathogens in the South Gobi desert region of Mongolia.

The alpine ecosystems and communities of central Asia are currently undergoing large-scale ecological and socio-ecological changes likely to affect wildlife-livestock-human disease interactions and zoonosis transmission risk. However, relatively little is known about the prevalence of pathogens in this region. Between 2012 and 2015 we screened 142 rodents in Mongolia's Gobi desert for exposure to important zoonotic and livestock pathogens. Rodent seroprevalence to Leptospira spp. was >1/3 of tested animals, Toxoplasma gondii and Coxiella burnetii approximately 1/8 animals, and the hantaviruses being between 1/20 (Puumala-like hantavirus) and <1/100 (Seoul-like hantavirus). Gerbils trapped inside local dwellings were one of the species seropositive to Puumala-like hantavirus, suggesting a potential zoonotic transmission pathway. Seventeen genera of zoonotic bacteria were also detected in the faeces and ticks collected from these rodents, with one tick testing positive to Yersinia. Our study helps provide baseline patterns of disease prevalence needed to infer potential transmission between source and target populations in this region, and to help shift the focus of epidemiological research towards understanding disease transmission among species and proactive disease mitigation strategies within a broader One Health framework.

One Health Approach to Globalizing, Accelerating, and Focusing Amphibian and Reptile Disease Research-Reflections and Opinions from the First Global Amphibian and Reptile Disease Conference.

The world's reptiles and amphibians are experiencing dramatic and ongoing losses in biodiversity, changes that can have substantial effects on ecosystems and human health. In 2022, the first Global Amphibian and Reptile Disease Conference was held, using One Health as a guiding principle. The conference showcased knowledge on numerous reptile and amphibian pathogens from several standpoints, including epidemiology, host immune defenses, wild population effects, and mitigation. The conference also provided field experts the opportunity to discuss and identify the most urgent herpetofaunal disease research directions necessary to address current and future threats to reptile and amphibian biodiversity.

Conserved Evolution of MHC Supertypes among Japanese Frogs Suggests Selection for Bd Resistance.

The chytrid fungus Batrachochytrium dendrobatidis (Bd) is a major threat to amphibians, yet there are no reports of major disease impacts in East Asian frogs. Genetic variation of the major histocompatibility complex (MHC) has been associated with resistance to Bd in frogs from East Asia and worldwide. Using transcriptomic data collated from 11 Japanese frog species (one individual per species), we isolated MHC class I and IIb sequences and validated using molecular cloning. We then compared MHC from Japanese frogs and other species worldwide, with varying Bd susceptibility. Supertyping analysis, which groups MHC alleles based on physicochemical properties of peptide binding sites, identified that all examined East Asian frogs contained at least one MHC-IIb allele belonging to supertype ST-1. This indicates that, despite the large divergence times between some Japanese frogs (up to 145 million years), particular functional properties in the peptide binding sites of MHC-II are conserved among East Asian frogs. Furthermore, preliminary analysis using NetMHCIIpan-4.0, which predicts potential Bd-peptide binding ability, suggests that MHC-IIb ST-1 and ST-2 have higher overall peptide binding ability than other supertypes, irrespective of whether the peptides are derived from Bd, other fungi, or bacteria. Our findings suggest that MHC-IIb among East Asian frogs may have co-evolved under the same selective pressure. Given that Bd originated in this region, it may be a major driver of MHC evolution in East Asian frogs.

Lead exposure of mainland Australia's top avian predator.

Lead (Pb) toxicity, through ingestion of lead ammunition in carcasses, is a threat to scavenging birds worldwide, but has received little attention in Australia. We analyzed lead exposure in the wedge-tailed eagle (Aquila audax), the largest raptor species found in mainland Australia and a facultative scavenger. Eagle carcasses were collected opportunistically throughout south-eastern mainland Australia between 1996 and 2022. Lead concentrations were measured in bone samples from 62 animals via portable X-ray fluorescence (XRF). Lead was detected (concentration >1 ppm) in 84% (n = 52) of the bone samples. The mean lead concentration of birds in which lead was detected was 9.10 ppm (±SE 1.66). Bone lead concentrations were elevated (10-20 ppm) in 12.9% of samples, and severe (>20 ppm) in 4.8% of samples. These proportions are moderately higher than equivalent data for the same species from the island of Tasmania, and are comparable to data from threatened eagle species from other continents. Lead exposure at these levels is likely to have negative impacts on wedge-tailed eagles at the level of the individual and perhaps at a population level. Our results suggest that studies of lead exposure in other Australian avian scavenger species are warranted.

Descriptive catalog of Homo naledi dental remains from the 2013 to 2015 excavations of the Dinaledi Chamber, site U.W. 101, within the Rising Star cave system, South Africa.

More than 150 hominin teeth, dated to ∼330-241 thousand years ago, were recovered during the 2013-2015 excavations of the Dinaledi Chamber of the Rising Star cave system, South Africa. These fossils comprise the first large single-site sample of hominin teeth from the Middle Pleistocene of Africa. Though scattered remains attributable to Homo sapiens, or their possible lineal ancestors, are known from older and younger sites across the continent, the distinctive morphological feature set of the Dinaledi teeth supports the recognition of a novel hominin species, Homo naledi. This material provides evidence of African Homo lineage diversity that lasts until at least the Middle Pleistocene. Here, a catalog, anatomical descriptions, and details of preservation and taphonomic alteration are provided for the Dinaledi teeth. Where possible, provisional associations among teeth are also proposed. To facilitate future research, we also provide access to a catalog of surface files of the Rising Star jaws and teeth.

Revisiting the age of the Florisbad hominin material.

In 1996, Grün and coworkers provided an ESR age determination for a hominin molar from Florisbad, South Africa, at 259 ± 35 ka. The most anatomically informative hominin specimen from Florisbad is a fragmentary craniofacial specimen, attributed by many to early Homo sapiens, which has been assumed to derive from the same individual as this tooth. Since 1996, evidence about the evolutionary context for the later Middle Pleistocene and its associated hominins in Africa had markedly changed. If the Florisbad date is accurate and if the molar is associated with the craniofacial specimen, this evidence may suggest that Homo naledi coexisted in time and geography with H. sapiens in southern Africa. Thus, the accuracy for all dates needs critical investigation. This study examines the published record of excavation and subsequent analyses for the Florisbad material. We find that the contemporary records raise doubt about the contemporaneity of the tooth and calvaria. The site's complex stratigraphy and the details about the tooth's discovery also pose challenges for the previous ESR age estimate. Because the fragmentary cranial specimen has value for morphological comparisons, developing additional means of understanding its geological age is necessary. The current data do not demonstrate the presence of H. sapiens in southern Africa at ~260 ka.

Perioperative escape from dormancy of spontaneous micro-metastases: A role for malignant secretion of IL-6, IL-8, and VEGF, through adrenergic and prostaglandin signaling.

We recently showed that a minimally-invasive removal of MDA-MB-231HM primary tumors (PTs) and elimination of their secreted factors (including IL-6, IL-8, VEGF, EGF, PDGF-aa, MIF, SerpinE1, and M-CSF), caused regression of spontaneous micro-metastases into a non-growing dormant state. To explore the underlying mechanisms and potential clinical ramifications of this phenomenon, we herein used the MDA-MB-231HM human breast cancer cell-line, in-vitro, and in vivo following orthotopic implantation in immune-deficient BALB/C nu/nu mice. Employing bioluminescence imaging, we found that adding laparotomy to minimally-invasive removal of the PT caused an outbreak of micro-metastases. However, perioperative ß-adrenergic and COX-2 inhibition, using propranolol + etodolac, maintained metastatic dormancy following laparotomy. In-vitro, ß-adrenergic agonists (epinephrine or metaproterenol) and prostaglandin-E2 markedly increased MDA-MB-231HM secretion of the pro-metastatic factors IL-6, IL-8, and VEGF, whereas cortisol reduced their secretion, effects that were maintained even 12 h after the washout of these agonists. In-vivo, laparotomy elevated IL-6 and IL-8 levels in both plasma and ex-vivo PT spontaneous secretion, whereas perioperative propranolol + etodolac administration blocked these effects. Similar trends were evident for EGF and MIF. Promoter-based bioinformatics analyses of excised PT transcriptomes implicated elevated NF-kB activity and reduced IRF1 activity in the gene regulatory effects of laparotomy, and these effects were inhibited by pre-surgical propranolol + etodolac. Taken together, our findings suggest a novel mechanism of post-operative metastatic outbreak, where surgery-induced adrenergic and prostanoid signaling increase the secretion of pro-metastatic factors, including IL-6, IL-8, and VEGF, from PT and possibly residual malignant tissue, and thereby prevent residual disease from entering dormancy.

Constant-temperature predictions underestimate growth of a fungal amphibian pathogen under individual host thermal profiles.

Ectotherm body temperatures fluctuate with environmental variability and host behavior, which may influence host-pathogen interactions. Fungal pathogens are a major threat to ectotherms and may be highly responsive to the fluctuating thermal profiles of individual hosts, especially cool-loving fungi exposed to high host temperatures. However, most studies estimate pathogen thermal performance based on averages of host or surrogate environmental temperatures, potentially missing effects of short-term host temperature shifts such as daily or hourly heat spikes. We recorded individual thermal profiles of Australian rainforest frogs using temperature-sensitive radio-transmitters. We then reproduced a subset of individual thermal profiles in growth chambers containing cultures of the near-global amphibian pathogen Batrachochytrium dendrobatidis (Bd) to investigate how realistic host temperature profiles affect Bd growth. We focused on thermal profiles that exceed the thermal optimum of Bd because the effects of realistic heat spikes on Bd growth are unresolved. Our laboratory incubation experiment revealed that Bd growth varied in response to relatively small differences in heat spike characteristics of individual frog thermal profiles, such as a single degree or a few hours, highlighting the importance of individual host behaviors in predicting population-level disease dynamics. The fungus also grew better than predicted under the most extreme and unpredictable frog temperature profile, recovering from two days of extreme (nearly 32 °C) heat spikes without negative effects on overall growth, suggesting we are underestimating the growth potential of the pathogen in nature. Combined with the previous finding that Bd reduces host heat tolerance, our study suggests that this pathogen may carry a competitive edge over hosts in the face of anthropogenic climate change.

Frontal sinuses and human evolution.

The frontal sinuses are cavities inside the frontal bone located at the junction between the face and the cranial vault and close to the brain. Despite a long history of study, understanding of their origin and variation through evolution is limited. This work compares most hominin species' holotypes and other key individuals with extant hominids. It provides a unique and valuable perspective of the variation in sinuses position, shape, and dimensions based on a simple and reproducible methodology. We also observed a covariation between the size and shape of the sinuses and the underlying frontal lobes in hominin species from at least the appearance of Homo erectus. Our results additionally undermine hypotheses stating that hominin frontal sinuses were directly affected by biomechanical constraints resulting from either chewing or adaptation to climate. Last, we demonstrate their substantial potential for discussions of the evolutionary relationships between hominin species.

Genetic approaches for increasing fitness in endangered species.

The global rate of wildlife extinctions is accelerating, and the persistence of many species requires conservation breeding programs. A central paradigm of these programs is to preserve the genetic diversity of the founder populations. However, this may preserve original characteristics that make them vulnerable to extinction. We introduce targeted genetic intervention (TGI) as an alternative approach that promotes traits that enable species to persist in the face of threats by changing the incidence of alleles that impact on fitness. The TGI toolkit includes methods with established efficacy in model organisms and agriculture but are largely untried for conservation, such as synthetic biology and artificial selection. We explore TGI approaches as a species-restoration tool for intractable threats including infectious disease and climate change.

Non-detection of mycoviruses in amphibian chytrid fungus (Batrachochytrium dendrobatidis) from Australia.

Mycoviruses may influence the pathogenicity of disease-causing fungi. Although mycoviruses have been found in some chytrid fungi, limited testing has not detected them in Batrachochytrium dendrobatidis (Bd), the cause of the devastating amphibian disease, chytridiomycosis. Here we conducted a survey for mycovirus presence in 38 Bd isolates from Australia (n = 31), Brazil (n = 5) and South Korea (n = 2) with a combination of modern high-throughput sequencing and conventional dsRNA cellulose chromatography. Mycoviruses were not detected in any isolates. This result was unexpected, given the long evolutionary history of Bd, as well as the high prevalence of mycoviruses in related fungal species. Given our widespread sampling in Australia and the limited number of Bd introductions, we suggest that mycoviruses are uncommon or absent from Australian Bd. Testing more isolates from regions where Bd originated, as well as regions with high diversity or low fungal virulence may identify mycoviruses that could aid in disease control.

New fossils of Australopithecus sediba reveal a nearly complete lower back.

Adaptations of the lower back to bipedalism are frequently discussed but infrequently demonstrated in early fossil hominins. Newly discovered lumbar vertebrae contribute to a near-complete lower back of Malapa Hominin 2 (MH2), offering additional insights into posture and locomotion in Australopithecus sediba. We show that MH2 possessed a lower back consistent with lumbar lordosis and other adaptations to bipedalism, including an increase in the width of intervertebral articular facets from the upper to lower lumbar column ('pyramidal configuration'). These results contrast with some recent work on lordosis in fossil hominins, where MH2 was argued to demonstrate no appreciable lordosis ('hypolordosis') similar to Neandertals. Our three-dimensional geometric morphometric (3D GM) analyses show that MH2's nearly complete middle lumbar vertebra is human-like in overall shape but its vertebral body is somewhat intermediate in shape between modern humans and great apes. Additionally, it bears long, cranially and ventrally oriented costal (transverse) processes, implying powerful trunk musculature. We interpret this combination of features to indicate that A. sediba used its lower back in both bipedal and arboreal positional behaviors, as previously suggested based on multiple lines of evidence from other parts of the skeleton and reconstructed paleobiology of A. sediba.

One of the defining features of humans is our ability to walk comfortably on two legs. To achieve this, our skeletons have evolved certain physical characteristics. For example, the lower part of the human spine has a forward curve that supports an upright posture; whereas the lower backs of chimpanzees and other apes ­ which walk around on four limbs and spend much of their time in trees ­ lack this curvature. Studying the fossilized back bones of ancient human remains can help us to understand how we evolved these features, and whether our ancestors moved in a similar way. Australopithecus sediba was a close-relative of modern humans that lived about two million years ago. In 2008, fossils from an adult female were discovered at a cave site in South Africa called Malapa. However, the fossils of the lower back region were incomplete, so it was unclear whether the female ­ referred to as Malapa Hominin 2 (MH2) ­ had a forward-curving spine and other adaptations needed to walk on two legs. Here, Williams et al. report the discovery of new A. sediba fossils from Malapa. The new fossils are mainly bones from the lower back, and they fit together with the previously discovered MH2 fossils, providing a nearly complete lower spine. Analysis of the fossils suggested that MH2 would have had an upright posture and comfortably walked on two legs, and the curvature of their lower back was similar to modern females. However, other aspects of the bones' shape suggest that as well as walking, A. sediba probably spent a significant amount of time climbing in trees. The findings of Williams et al. provide new insights in to our evolutionary history, and ultimately, our place in the natural world around us. Our lower back is prone to injury and pain associated with posture, pregnancy and exercise (or lack thereof). Therefore, understanding how the lower back evolved may help us to learn how to prevent injuries and maintain a healthy back.

Declining amphibians might be evolving increased reproductive effort in the face of devastating disease.

The devastating infectious disease chytridiomycosis has caused declines of amphibians across the globe, yet some populations are persisting and even recovering. One understudied effect of wildlife disease is changes in reproductive effort. Here, we aimed to understand if the disease has plastic effects on reproduction and if reproductive effort could evolve with disease endemism. We compared the effects of experimental pathogen exposure (trait plasticity) and population-level disease history (evolution in trait baseline) on reproductive effort using gametogenesis as a proxy in the declining and endangered frog Litoria verreauxii alpina. We found that unexposed males from disease-endemic populations had higher reproductive effort, which is consistent with an evolutionary response to chytridiomycosis. We also found evidence of trait plasticity, where males and females were affected differently by infection: pathogen exposed males had higher reproductive effort (larger testes), whereas females had reduced reproductive effort (smaller and fewer developed eggs) regardless of the population of origin. Infectious diseases can cause plastic changes in the reproductive effort at an individual level, and population-level disease exposure can result in changes to baseline reproductive effort; therefore, individual- and population-level effects of disease should be considered when designing management and conservation programs for threatened and declining species.