Comparative genomics provides insights into the aquatic adaptations of mammals.

The ancestors of marine mammals once roamed the land and independently committed to an aquatic lifestyle. These macroevolutionary transitions have intrigued scientists for centuries. Here, we generated high-quality genome assemblies of 17 marine mammals (11 cetaceans and six pinnipeds), including eight assemblies at the chromosome level. Incorporating previously published data, we reconstructed the marine mammal phylogeny and population histories and identified numerous idiosyncratic and convergent genomic variations that possibly contributed to the transition from land to water in marine mammal lineages. Genes associated with the formation of blubber (NFIA), vascular development (SEMA3E), and heat production by brown adipose tissue (UCP1) had unique changes that may contribute to marine mammal thermoregulation. We also observed many lineage-specific changes in the marine mammals, including genes associated with deep diving and navigation. Our study advances understanding of the timing, pattern, and molecular changes associated with the evolution of mammalian lineages adapting to aquatic life.

Ranging pattern development of a declining delphinid population: A potential cascade effect of vessel activities.

It is generally accepted that vessel activity causes various behavioral responses of cetaceans and undermines individual fitness. Whether or how it can lead to a demographic response of populations remains rarely examined. In the northern Beibu Gulf, China, vessel activities have sharply increased in the past two decades, while abnormal demographic dynamics was recently noted for the resident Indo-Pacific humpback dolphins. The present study first examined the humpback dolphins' utilization distribution (UD) from 2003 to 2019. Habitat suitability was then modeled with the sighting data collected before the most recent population reduction. Finally, we tried to disentangle the anthropogenic driver of dolphin demography by cross-referring the spatiotemporal development of dolphins' UD, vessel activities, and habitat suitability. Our results showed that the dolphins' UD shrank substantially during the port expansion in the early 2010s, and we suggest that the consequential increase in vessel activities might impose extra marine stressors on the resident humpback dolphins. To reduce the boat interaction, the dolphins steadily shifted their core area to a less suitable area in the east during 2015-2017, when unnaturally low survivals were recorded. Afterward, the dolphin core area partially shifted back to the more suitable area in the west, which corresponded to the improving dolphin survival in 2018. Our finding suggested that the vessel activity may be responsible for the dolphin displacement, while staying in the less suitable area may further lead to a more severe and acute demographic consequence on the population. The underlying and indirect impact of vessel activities as disclosed by the present study is particularly important for port management, marine planning, and conservation practice regarding coastal cetaceans, especially for those resident and endangered populations inhabiting the urbanized coastal areas.

Highly sensitive protein detection using recombinant spores and lateral flow immunoassay.

Lateral flow immunoassays (LFIs) can be used to detect intact bacteria or spores; when gold nanoparticles (AuNPs) are used as the signal reporters, the detection limits are very low. Spore-based surface display has been widely studied for enzyme immobilization and live-nontoxic oral vaccines. In this study, recombinant spores were used to improve the sensitivity of a LFI. We developed a test kit that combines streptavidin-displayed spores with a LFI assay for rapid protein detection. The recombinant spores served as a signal amplifier and AuNPs were used as the signal reporters. For detection of ß-galactosidase, which was used as the model protein, the detection limit was about 10-15 mol, while that of the conventional LFI is about 10-12 mol. In both methods, nanogold was used as the colorimetric signal and could be observed with the naked eye. This method improved LFI sensitivity without sacrificing its advantages. Furthermore, enhanced green fluorescent protein (eGFP) was also displayed on the surface of the streptavidin-displayed spores. Without AuNPs, the fluorescent recombinant spores acted as the signal, which could be detected by a fluorescence detector, such as a fluorescence microscope. The detection limit was 10-16 mol under fluorescence microscopy whose magnification was 25-fold. Therefore, in conclusion, in this proof of concept study, the detection limits of both proposed methods were far superior to those of traditional LFI assay.

Detection of biotin with zeptomole sensitivity using recombinant spores and a competition assay.

Detection of protein-binding analytes is important for many applications. Currently, various instrument-based techniques are used for detecting protein-binding analytes. However, such techniques have several limitations including high cost and time-consuming sample processing. In order to overcome these limitations, we developed a sensitive competition assay for the detection of protein-binding analytes using recombinant endospores as a sensing element. The method is based on the competition between the biotin, the model analyte, and a biotin-magnetic bead complex to bind the recombinant spores containing the biotin binding region of streptavidin. After magnetic attraction, the residual spores in the suspension are spread on plates to form colonies which are used to count the amount of the residual spores; the higher the residual ratio of spores, the more biotin in the samples. The linear range was from 150 zmol to 1.5 fmol and the limit of detection of the assay was 150 zmol. The assay proposed herein is sensitive and does not require any expensive equipment. It is suitable for qualitative or semi-quantitative analysis such as screening tests for the detection of toxic chemicals.

Whole Genome Sequencing of Chinese White Dolphin (Sousa chinensis) for High-Throughput Screening of Antihypertensive Peptides.

Chinese white dolphin (Sousa chinensis), also known as the Indo-Pacific humpback dolphin, has been classified as "Vulnerable" on the IUCN Red List of Threatened Species. It is a special cetacean species that lives in tropical and subtropical nearshore waters, with significant differences from other cetaceans. Here, we sequenced and assembled a draft genome of the Chinese white dolphin with a total length of 2.3 Gb and annotation of 18,387 protein-coding genes. Genes from certain expanded families are potentially involved in DNA replication and repairing, suggesting that they may be related to adaptation of this marine mammal to nearshore environments. We also discovered that its historical population had undergone a remarkable bottleneck incident before the Mindel glaciation. In addition, a comparative genomic survey on antihypertensive peptides (AHTPs) among five representative mammals with various residential habitats (such as remarkable differences in exogenous ion concentrations and sea depth) revealed that these small bioactive peptides were highly conserved among these examined mammals, and they had the most abundant hits in collagen subunit proteins, especially for two putative AHTP peptides Gly-Leu-Pro (GLP) and Leu-Gly-Pro (LGP). Our genome assembly will be a valuable resource for further genetic researches on adaptive ecology and conservation biology of cetaceans, and for in-depth investigations into bioactive peptides in aquatic and terrestrial mammals for development of peptide-based drugs to treat various human cardiovascular diseases.

In-plane Anisotropy of Quantum Transport in Artificial Two-dimensional Au Lattices.

We report an experimental observation and direct control of quantum transport in artificial two-dimensional Au lattices. Combining the advanced techniques of low-temperature deposition and newly developed double-probe scanning tunneling spectroscopy, we display a two-dimensional carrier transport and demonstrate a strong in-plane transport modulation in the two-dimensional Au lattices. In well-ordered Au lattices, we observe the carrier transport behavior manifesting as a band-like feature with an energy gap. Furthermore, controlled structural modification performed by constructing coupled "stadiums" enables a transition of system dynamics in the lattices, which in turn establishes tunable resonant transport throughout a wide energy range. Our findings open the possibility of the construction and transport engineering of artificial lattices by the geometrical arrangement of scatterers and quantum chaotic dynamics.

Dichroic Behavior of Gold Nanoparticles Synthesized in Aqueous Solution with Insufficient Reducing Agent.

The spherical gold nanoparticles (AuNPs) typically are red in solution. However, in this study, the dichroic and spherical AuNPs were synthesized using a modified seeding growth method under reducing agent insufficiency in an aqueous solution. This particular AuNP solution is orange in reflected light and red in transmitted light. The reflectance curves confirm that the dichroic AuNPs are different from the classic AuNPs. With particle assembling, the AuNP solution is fainter orange in reflected light, but purple in transmitted light when the color of classic spherical AuNP solution is purple in both lights. Furthermore, the aggregated-nanogold solutions were added to HAuCl4 solutions with the addition of an insufficient amount reducing agent. The solution changed from faint orange to bright orange in reflected light and from purple to blue in transmitted light. It indicates that the gold assembling under a reducing agent insufficiency, not the shape of AuNP, causes the dichroic phenomenon. To the best of our knowing, this is the first study to report how the AuNP is synthesized, not the shape, affects the color of the AuNP.

Low Major Histocompatibility Complex Class II Variation in the Endangered Indo-Pacific Humpback Dolphin (Sousa chinensis): Inferences About the Role of Balancing Selection.

It has been widely reported that the major histocompatibility complex (MHC) is under balancing selection due to its immune function across terrestrial and aquatic mammals. The comprehensive studies at MHC and other neutral loci could give us a synthetic evaluation about the major force determining genetic diversity of species. Previously, a low level of genetic diversity has been reported among the Indo-Pacific humpback dolphin (Sousa chinensis) in the Pearl River Estuary (PRE) using both mitochondrial marker and microsatellite loci. Here, the expression and sequence polymorphism of 2 MHC class II genes (DQB and DRB) in 32 S. chinensis from PRE collected between 2003 and 2011 were investigated. High ratios of non-synonymous to synonymous substitution rates, codon-based selection analysis, and trans-species polymorphism (TSP) support the hypothesis that balancing selection acted on S. chinensis MHC sequences. However, only 2 haplotypes were detected at either DQB or DRB loci. Moreover, the lack of deviation from the Hardy-Weinberg expectation at DRB locus combined with the relatively low heterozygosity at both DQB locus and microsatellite loci suggested that balancing selection might not be sufficient, which further suggested that genetic drift associated with historical bottlenecks was not mitigated by balancing selection in terms of the loss of MHC and neutral variation in S. chinensis. The combined results highlighted the importance of maintaining the genetic diversity of the endangered S. chinensis.

Humpback Dolphins in Hong Kong and the Pearl River Delta: Status, Threats and Conservation Challenges.

In coastal waters of the Pearl River Delta (PRD) region, the Indo-Pacific humpback dolphin (Sousa chinensis) is thought to number approximately 2500 individuals. Given these figures, the putative PRD population may appear strong enough to resist demographic stochasticity and environmental pressures. However, living in close proximity to the world's busiest seaport/airport and several densely populated urban centres with major coastal infrastructural developments comes with challenges to the long-term survival of these animals. There are few other small cetacean populations that face the range and intensity of human-induced pressures as those present in the PRD and current protection measures are severely inadequate. Recent mark-recapture analyses of the animals in Hong Kong waters indicate that in the past two decades the population parameters have not been well understood, and spatial analyses show that only a very small proportion of the dolphins' key habitats are given any form of protection. All current marine protected areas within the PRD fail to meet a minimum habitat requirement that could facilitate the population's long-term persistence. Demographic models indicate a continuous decline of 2.5% per annum, a rate at which the population is likely to drop below the demographic threshold within two generations and lose 74% of the current numbers within the lifespan of three generations. In Hong Kong, the case of humpback dolphins represents a particularly explicit example of inadequate management where a complete revision of the fundamental approach to conservation management is urgently needed.

Likely Age-Related Hearing Loss (Presbycusis) in a Stranded Indo-Pacific Humpback Dolphin (Sousa chinensis).

The hearing of a stranded Indo-Pacific humpback dolphin (Sousa chinensis) in Zhuhai, China, was measured. The age of this animal was estimated to be ~40 years. The animal's hearing was measured using a noninvasive auditory evoked potential (AEP) method. The results showed that the high-frequency hearing cutoff frequency of the studied dolphin was ~30-40 kHz lower than that of a conspecific younger individual ~13 year old. The lower high-frequency hearing range in the older dolphin was explained as a likely result of age-related hearing loss (presbycusis).

Electronic bandgap and edge reconstruction in phosphorene materials.

Single-layer black phosphorus (BP), or phosphorene, is a highly anisotropic two-dimensional elemental material possessing promising semiconductor properties for flexible electronics. However, the direct bandgap of single-layer black phosphorus predicted theoretically has not been directly measured, and the properties of its edges have not been considered in detail. Here we report atomic scale electronic variation related to strain-induced anisotropic deformation of the puckered honeycomb structure of freshly cleaved black phosphorus using a high-resolution scanning tunneling spectroscopy (STS) survey along the light (x) and heavy (y) effective mass directions. Through a combination of STS measurements and first-principles calculations, a model for edge reconstruction is also determined. The reconstruction is shown to self-passivate most dangling bonds by switching the coordination number of phosphorus from 3 to 5 or 3 to 4.

Skinny dolphins: Can poor body condition explain population decline in Indo-Pacific humpback dolphins (Sousa chinensis)?

Indo-Pacific humpback dolphins (IPHDs) who form resident populations along the Chinese coastline are facing a wide range of anthropogenic disturbances including intense fishing and some populations have been shown to experience a severe decline. Body condition is thought to be a good indicator of health since it is linked to survival and reproductive success. In order to better understand population trends, we investigated whether the body condition of IPHDs is poorer in populations whose status is alarming than in other populations. UAV flights were conducted from 2022 to 2023 in four locations (i.e., Sanniang Bay, Leizhou Bay, Jiangmen, and Lingding Bay) in the northern South China Sea. Body ratios were calculated using the body length and widths of IPHDs and were used to analyze differences among seasons, locations, and demographic parameters. A PCA was then used to obtain a detailed picture of the body condition composition of dolphins at each location. Results showed that dolphins from Leizhou Bay and Jiangmen were in better body condition than those from Sanniang Bay and Lingding Bay. Since populations inhabiting Sanniang Bay and Lingding Bay have been shown to experience a sharp decline, it can be hypothesized that poor body condition may have played a role in such a trend. Further investigations of the factors impacting IPHDs' body condition are needed, including monitoring of prey density, contaminant concentration, stress levels, and impacts of human activities on dolphins' behavior. In addition, the creation of a robust scoring method would allow for regular monitoring of IPHDs' body condition to inform conservation measures.

Enhancing antibody detection sensitivity in lateral flow immunoassays using endospores of Bacillus subtilis as signal amplifiers.

Antibody detection is the critical first step for tracking the spread of many diseases including COVID-19. Lateral flow immunoassay (LFIA) is the most commonly used method for rapid antibody detection because it is easy-to-use and inexpensive. However, LFIA has limited sensitivity when gold nanoparticles (AuNPs) are used as the signals. In this study, the endospores of Bacillus subtilis were used in combination with AuNP in a LFIA to detect antibodies. The endospores serve as a signal amplifier. The detection limit was about 10-8 M for anti-beta galactosidase antibody detection whereas the detection limit of conventional LFIA is about 10-6 M. Furthermore, the proposed methods have no additional user steps compared with the traditional LFIA. This method, therefore, improved the sensitivity 100-fold without compromising any advantages of LFIA. We believe that the proposed method will be useful for detection of antibodies against HIV, Zika virus, SARS-CoV-2, and so on.

Possible age-related hearing loss (presbycusis) and corresponding change in echolocation parameters in a stranded Indo-Pacific humpback dolphin.

The hearing and echolocation clicks of a stranded Indo-Pacific humpback dolphin (Sousa chinensis) in Zhuhai, China, were studied. This animal had been repeatedly observed in the wild before it was stranded and its age was estimated to be ~40 years. The animal's hearing was measured using a non-invasive auditory evoked potential (AEP) method. Echolocation clicks produced by the dolphin were recorded when the animal was freely swimming in a 7.5 m (width)×22 m (length)×4.8 m (structural depth) pool with a water depth of ~2.5 m. The hearing and echolocation clicks of the studied dolphin were compared with those of a conspecific younger individual, ~13 years of age. The results suggested that the cut-off frequency of the high-frequency hearing of the studied dolphin was ~30-40 kHz lower than that of the younger individual. The peak and centre frequencies of the clicks produced by the older dolphin were ~16 kHz lower than those of the clicks produced by the younger animal. Considering that the older dolphin was ~40 years old, its lower high-frequency hearing range with lower click peak and centre frequencies could probably be explained by age-related hearing loss (presbycusis).

Local crystallography analysis for atomically resolved scanning tunneling microscopy images.

Scanning probe microscopy has emerged as a powerful and flexible tool for atomically resolved imaging of surface structures. However, due to the amount of information extracted, in many cases the interpretation of such data is limited to being qualitative and semi-quantitative in nature. At the same time, much can be learned from local atom parameters, such as distances and angles, that can be analyzed and interpreted as variations of local chemical bonding, or order parameter fields. Here, we demonstrate an iterative algorithm for indexing and determining atomic positions that allows the analysis of inhomogeneous surfaces. This approach is further illustrated by local crystallographic analysis of several real surfaces, including highly ordered pyrolytic graphite and an Fe-based superconductor FeTe0.55Se0.45. This study provides a new pathway to extract and quantify local properties for scanning probe microscopy images.

A genome and single-nucleus cerebral cortex transcriptome atlas of the short-finned pilot whale Globicephala macrorhynchus.

Cetaceans (dolphins, whales, and porpoises) have large and anatomically sophisticated brains. To expand our understanding of the cellular makeup of cetacean brains and the similarities and divergence between the brains of cetaceans and terrestrial mammals, we report a short-finned pilot whale (Globicephala macrorhynchus) single-nucleus transcriptome atlas. To achieve this goal, we assembled a chromosome-scale reference genome spanning 2.25 Gb on 22 chromosomes and profiled the gene expression of five major anatomical cortical regions of the short-finned pilot whale by single-nucleus RNA-sequencing (snRNA-seq). We identified six major cell lineages in the cerebral cortex (excitatory neurons, inhibitory neurons, oligodendrocytes, oligodendrocyte precursor cells, astrocytes, and endothelial cells), eight molecularly distinct subclusters of excitatory neurons, and four subclusters of inhibitory neurons. Finally, a comparison of snRNA-seq data from the short-finned pilot whale, human, and rhesus macaque revealed a broadly conserved cellular makeup of brain cell types. Our study provides genomic resources and molecular insights into cetacean brain evolution.

Microbial diversity and structure in the gastrointestinal tracts of two stranded short-finned pilot whales (Globicephala macrorhynchus) and a pygmy sperm whale (Kogia breviceps).

Information on the gut microbiome composition of different mammals could provide novel insights into the evolution of mammals and succession of microbial communities in different hosts. However, there is limited information on the gut microbiome composition of marine mammals, especially cetaceans because of sampling constraints. In this study, we investigated the diversity and composition of microbial communities in the stomach, midgut, and hindgut of 2 stranded short-finned pilot whales (Globicephala macrorhynchus) and hindgut of a stranded pygmy sperm whale (Kogia breviceps) by using 16S rRNA gene amplicon sequencing technology. On the basis of the 50 most abundant operational taxonomic units, principal coordinate analysis, and non-metric multidimensional scaling analysis, we confirmed that the gut microbial communities of the 3 whales were different. Our results revealed that the gut microbiome of 1 stranded short-finned pilot whale GM16 was dominated by Firmicutes (mainly Clostridium) and Fusobacteria; whereas that of the other pilot whale GM19 was composed of Gammaproteobacteria and Bacteroidetes (mainly Vibrio and Bacteroides, respectively), probably caused by intestinal disease and antibiotic treatment. The gut microbiome of the pygmy sperm whale was dominated by Firmicutes and Bacteroidetes. Moreover, different gastrointestinal tract regions harbored different microbial community structures. To our knowledge, this is the first report of the gut microbiome of short-finned pilot whales, and our findings will expand our current knowledge on microbial diversity and composition in the gastrointestinal tract of cetaceans.

Prey decline leads to diet shift in the largest population of Indo-Pacific humpback dolphins?

The Pearl River Delta (PRD) region on the southeast coast of China has long been known as a highly productive fishing ground. Since the late 1980s, fishing pressure in the PRD has been intense, which warrants concerns of potential fishery-related impacts on the food resources and foraging ecology of apex marine predators in this region, such as the Indo-Pacific humpback dolphin (Sousa chinensis). In this study, we examined 54 stomachs with food remains, collected from beached carcasses of humpback dolphins recovered during fifteen years between 2003 and 2017. The 6043 identified prey items represent 62 teleost taxa, primarily small estuarine fish, but also larger reef fish. The dolphins appear to be opportunistic foragers, hunting across the water-column, with preference for shoaling and meaty fishes (e.g. Collichthys lucidus IRI% = 38.6%, Johnius belangerii IRI% = 23.1%, Mugil cephalus IRI% = 14.0%). Our findings suggest a dietary shift in recent years, from primarily demersal (as previously reported) to greater intake of neritic and pelagic fish. Dolphin foraging group size has decreased in recent years, which corresponds with declining size and numbers of prey items retrieved from dolphin stomachs. We suggest that these are indicators of declining food resources. Faced with a shortage of preferred prey, humpback dolphins may have broadened their dietary spectrum to maintain their daily energy intake, while their foraging group size decreased in response to the altered tradeoff between the costs and benefits of group foraging.

Early divergence and differential population histories of the Indo-Pacific humpback dolphin in the Pacific and Indian Oceans.

The currently recognized Indo-Pacific humpback dolphin occurs in estuaries and surrounding shallow waters from the South China Sea to the Asian coast of the Indian Ocean. However, a recent study suggested that the humpback dolphin from the Bay of Bengal may represent a distinct phylogenetic species. In this study, we sequenced 915-bp mtDNA segments from five geographic populations in both Chinese and Thai waters; together with previously published sequences, these data revealed that the ancestral Indo-Pacific humpback dolphin might have split during the transition from the Oligocene to Miocene (23.45 Mya, 95% HPD: 16.65-26.55 Mya), and then dispersed along the Pacific and Indian Ocean coasts of Asia. Genetic differentiation was detected between most of the examined populations, except for only a few pairwise populations in the northern South China Sea. Genetic differentiation/distance between the humpback dolphins from the northern and southern South China Sea met the sub-species threshold value proposed for marine mammals, whereas that between the humpback dolphins in the Pacific and the Indian Ocean was above the species threshold. Bayesian inference of historic gene flow indicated low but constant northward gene flow along the Indian Ocean coast; however, there was a recent abrupt increase in gene flow in the Pacific region, likely due to the shortening coastline at the low stand of sea level. Our results revealed that the current taxonomic classification of Indo-Pacific humpback dolphins may not reflect their phylogeography.

Cetacean occurrence and diversity in whale-watching waters off Mirissa, Southern Sri Lanka.

Scientific information is vital to the conservation of cetaceans and the management of whale-watching activities. The southern coastal waters of Sri Lanka are near a narrow continental shelf and biologically abundant in cetacean species. Although the occurrence of cetaceans has been investigated in certain waters of Sri Lanka, few surveys have been conducted along the southern coast. To fill this gap, we conducted boat-based surveys from January to May 2017 to investigate the occurrence, diversity, and behavior of cetaceans in the waters off Mirissa, covering a survey area of 788.9 km2 . During 55 survey days, we recorded a total of 242 cetacean sightings and identified at least 9 species (3 mysticetes and 6 odontocetes). The blue whale was the most common mysticete species (167 of 174 mysticete encounters), followed by the Omura's whale (4 of 174) and Bryde's whale (3 of 174). The spinner dolphin was the most common odontocete species (28 of 68 odontocete encounters), followed by the sperm whale (18 of 68), common bottlenose dolphin (13 of 68), short-finned pilot whale (5 of 68), melon-headed whale (2 of 68), and killer whale. Blue whales and sperm whales exhibited a clear preference for outer shelf and high slope areas, and blue whales were observed feeding along these waters. The present study provides near-baseline information on cetacean occurrence and diversity in whale-watching waters off southern Sri Lanka, and highlights the urgent need for proper management strategies for whale-watching activities.