Pseudomonas-associated bacteria play a key role in obtaining nutrition from bamboo for the giant panda (Ailuropoda melanoleuca).

Gut microbiota plays a vital role in obtaining nutrition from bamboo for giant pandas. However, low cellulase activity has been observed in the panda's gut. Besides, no specific pathway has been implicated in lignin digestion by gut microbiota of pandas. Therefore, the mechanism by which they obtain nutrients is still controversial. It is necessary to elucidate the precise pathways employed by gut microbiota of pandas to degrade lignin. Here, the metabolic pathways for lignin degradation in pandas were explored by comparing 209 metagenomic sequencing data from wild species with different feeding habits. Lignin degradation central pathways, including beta-ketoadipate and homogentisate pathway, were enriched in the gut of wild bamboo-eating pandas. The gut microbiome of wild bamboo-eating specialists was enriched with genes from pathways implicated in degrading ferulate and p-coumarate into acetyl-CoA and succinyl-CoA, which can potentially provide the raw materials for metabolism in pandas. Specifically, Pseudomonas, as the most dominant gut bacteria genus, was found to be the main bacteria to provide genes involved in lignin or lignin derivative degradation. Herein, three Pseudomonas-associated strains isolated from the feces of wild pandas showed the laccase, lignin peroxidase, and manganese peroxidase activity and extracellular lignin degradation ability in vitro. A potential mechanism for pandas to obtain nutrition from bamboo was proposed based on the results. This study provides novel insights into the adaptive evolution of pandas from the perspective of lignin metabolism. IMPORTANCE: Although giant pandas only feed on bamboo, the mechanism of lignin digestion in pandas is unclear. Here, the metabolic pathways for lignin degradation in wild pandas were explored by comparing gut metagenomic from species with different feeding habits. Results showed that lignin degradation central pathways, including beta-ketoadipate and homogentisate pathway, were enriched in the gut of wild bamboo-eating pandas. Genes from pathways involved in degrading ferulate and p-coumarate via beta-ketoadipate pathway were also enriched in bamboo-eating pandas. The final products of the above process, such as acetyl-CoA, can potentially provide the raw materials for metabolism in pandas. Specifically, Pseudomonas, as the most dominant gut bacteria genus, mainly provides genes involved in lignin degradation. Herein, Pseudomonas-associated strains isolated from the feces of pandas could degrade extracellular lignin. These findings suggest that gut microbiome of pandas is crucial in obtaining nutrition from lignin via Pseudomonas, as the main lignin-degrading bacteria.

Effects of Age and Season on Blood Parameters of Captive Giant Pandas: A Pilot Study.

The giant panda, Ailuropoda melanoleuca, serves as a flagship species for biodiversity conservation, embodying the intersection of ecological, evolutionary, and anthropogenic forces shaping the natural world. Hematological parameters serve as crucial indicators for assessing the physiological status of animals. However, our understanding of blood parameters and hemorheology in captive giant pandas under non-anesthetic conditions is limited. In this study, from September 2018 to August 2020, we collected blood samples from captive giant pandas under non-anesthetic conditions. Twelve captive giant pandas, ranging in age from 2 to 28 years, were divided into three groups based on their age, and the variations in basic blood parameters and hemorheological parameters across four seasons were analyzed. This provided baseline data for future blood sample comparisons in non-anesthetized captive giant pandas. Additionally, we observed seasonal changes in hematological morphology, hemorheology, and serum enzymes. Moreover, seasonality had a regulatory effect on hemorheological parameters and negatively impacted blood viscosity. Age influenced changes in serum enzymes, serum protein content, and serum metabolites, indicating differences in overall metabolic processes among giant pandas of different age groups. Whether factors such as season and climate contribute to environmental stress in captive giant pandas requires further investigation. The findings of this study may help to protect the stability of the giant panda population better and provide a reference for the medical care of captive giant pandas.

Comparative proteomics in captive giant pandas to identify proteins involved in age-related cataract formation.

Approximately 20% of aged captive giant pandas (Ailuropoda melanoleuca) have cataracts that impair their quality of life. To identify potential biomarkers of cataract formation, we carried out a quantitative proteomics analysis of 10 giant pandas to find proteins differing in abundance between healthy and cataract-bearing animals. We identified almost 150 proteins exceeding our threshold for differential abundance, most of which were associated with GO categories related to extracellular localization. The most significant differential abundance was associated with components of the proteasome and other proteins with a role in proteolysis or its regulation, most of which were depleted in pandas with cataracts. Other modulated proteins included components of the extracellular matrix or cytoskeleton, as well as associated signaling proteins and regulators, but we did not find any differentially expressed transcription factors. These results indicate that the formation of cataracts involves a complex post-transcriptional network of signaling inside and outside lens cells to drive stress responses as a means to address the accumulation of protein aggregates triggered by oxidative damage. The modulated proteins also indicate that it should be possible to predict the onset of cataracts in captive pandas by taking blood samples and testing them for the presence or absence of specific protein markers.

A rare case of giant panda cancer: Pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is a highly malignant cancer of the digestive system common among humans. However, it is rarely reported in wild animals. In 2018, a giant panda died in the Beijing Zoo. During subsequent histological observation of the pancreas, it was discovered that the glandular epithelial cells had lost the pancreatic acinar structure, tubular areas with obvious structure in the pancreas, and the ductal epithelium was substituted by high columnar mucus cells. Masson staining showed that there were several fibrous tissue proliferative reactions around the ductal adenocarcinoma and immunohistochemical staining revealed that CK7 and CK19 were positively expressed in the pancreatic tissue. Therefore, the pathological diagnosis indicated that the panda had PDAC. In this paper, the panda's living conditions and pathological diagnosis results are examined, with the aim of providing a reference point for the future diagnosis of wild animal tumors.

RNA-Seq analysis in giant pandas reveals the differential expression of multiple genes involved in cataract formation.

BACKGROUND: The giant panda (Ailuropoda melanoleuca) is an endangered mammalian species native to China. Fewer than 2500 giant pandas are known to exist, many of which are bred in captivity as a means to preserve and repopulate the species. Like other captive mammals, giant pandas acquire age-related cataracts, reducing their quality of life. Recent comparative genome-wide methylation analysis revealed 110 differentially methylated genes associated with cataract formation including six also associated with the formation of age-related cataracts in humans. RESULTS: To investigate the pathological pathway in greater detail, here we used RNA-Seq analysis to investigate the differential expression profiles of genes in three giant pandas with cataracts and three healthy controls. We identified more than 700 differentially expressed genes, 29 of which were selected for further analysis based on their low q-value. We found that many of the genes encoded regulatory and signaling proteins associated with the control of cell growth, migration, differentiation and apoptosis, supporting previous research indicating a key role for apoptosis in cataract formation. CONCLUSION: The identification of genes involved in the formation of age-related cataracts could facilitate the development of predictive markers, preventative measures and even new therapies to improve the life of captive animals.

A novel missense mutation in the HSF4 gene of giant pandas with senile congenital cataracts.

Cataracts are a common cause of visual impairment and blindness in mammals. They are usually associated with aging, but approximately one third of cases have a significant genetic component. Cataracts are increasingly prevalent among aging populations of captive giant pandas (Ailuropoda melanoleuca) and it is therefore important to identify genetic determinants that influence the likelihood of cataract development in order to distinguish between congenital and age-related disease. Here we screened for cataract-related genetic effects using a functional candidate gene approach combined with bioinformatics to identify the underlying genetic defect in a giant panda with congenital cataracts. We identified a missense mutation in exon 10 of the HSF4 gene encoding heat shock transcription factor 4. The mutation causes the amino acid substitution R377W in a highly conserved segment of the protein between the isoform-specific and downstream hydrophobic regions. Predictive modeling revealed that the substitution is likely to increase the hydrophobicity of the protein and disrupt interactions with spatially adjacent amino acid side chains. The mutation was not found in 13 unaffected unrelated animals but was found in an unrelated animal also diagnosed with senile congenital cataract. The novel missense mutation in the HSF4 gene therefore provides a potential new genetic determinant that could help to predict the risk of cataracts in giant pandas.

A novel missense mutation in the gene encoding major intrinsic protein (MIP) in a Giant panda with unilateral cataract formation.

BACKGROUND: Cataracts are defects of the lens that cause progressive visual impairment and ultimately blindness in many vertebrate species. Most cataracts are age-related, but up to one third have an underlying genetic cause. Cataracts are common in captive zoo animals, but it is often unclear whether these are congenital or acquired (age-related) lesions. RESULTS: Here we used a functional candidate gene screening approach to identify mutations associated with cataracts in a captive giant panda (Ailuropoda melanoleuca). We screened 11 genes often associated with human cataracts and identified a novel missense mutation (c.686G > A) in the MIP gene encoding major intrinsic protein. This is expressed in the lens and normally accumulates in the plasma membrane of lens fiber cells, where it plays an important role in fluid transport and cell adhesion. The mutation causes the replacement of serine with asparagine (p.S229N) in the C-terminal tail of the protein, and modeling predicts that the mutation induces conformational changes that may interfere with lens permeability and cell-cell interactions. CONCLUSION: The c.686G > A mutation was found in a captive giant panda with a unilateral cataract but not in 18 controls from diverse regions in China, suggesting it is most likely a genuine disease-associated mutation rather than a single-nucleotide polymorphism. The mutation could therefore serve as a new genetic marker to predict the risk of congenital cataracts in captive giant pandas.

Genome-wide analysis of methylation in giant pandas with cataract by methylation-dependent restriction-site associated DNA sequencing (MethylRAD).

The giant panda (Ailuropoda melanoleuca) is a native species to China. They are rare and endangered and are regarded as the 'national treasure' and 'living fossil' in China. For the time being, there are only about 2500 giant pandas in the world. Therefore, we still have to do much more efforts to protect the giant pandas. In captive wildlife, the cataract incidence of mammalian always increases with age. Currently, in China, the proportion of elderly giant pandas who suffering from cataract has reached 20%. The eye disorder thus has a strong influence on the physical health and life quality of the elderly giant pandas. To discover the genes associated with the pathogenesis of cataract in the elderly giant panda and achieve the goal of early assessment and diagnosis of cataract in giant pandas during aging, we performed whole genome methylation sequencing in 3 giant pandas with cataract and 3 healthy giant pandas using methylation-dependent restriction-site associated DNA sequencing (MethylRAD). In the present study, we obtained 3.62M reads, on average, for each sample, and identified 116 and 242 differentially methylated genes (DMGs) between the two groups under the context of CCGG and CCWGG on genome, respectively. Further KEGG and GO enrichment analyses determined a total of 110 DMGs that are involved in the biological functions associated with pathogenesis of cataract. Among them, 6 DMGs including EEA1, GARS, SLITRK4, GSTM3, CASP3, and EGLN3 have been linked with cataract in old age.

ANIMAL PHYSIOLOGY. Exceptionally low daily energy expenditure in the bamboo-eating giant panda.

The carnivoran giant panda has a specialized bamboo diet, to which its alimentary tract is poorly adapted. Measurements of daily energy expenditure across five captive and three wild pandas averaged 5.2 megajoules (MJ)/day, only 37.7% of the predicted value (13.8 MJ/day). For the wild pandas, the mean was 6.2 MJ/day, or 45% of the mammalian expectation. Pandas achieve this exceptionally low expenditure in part by reduced sizes of several vital organs and low physical activity. In addition, circulating levels of thyroid hormones thyroxine (T4) and triiodothyronine (T3) averaged 46.9 and 64%, respectively, of the levels expected for a eutherian mammal of comparable size. A giant panda-unique mutation in the DUOX2 gene, critical for thyroid hormone synthesis, might explain these low thyroid hormone levels. A combination of morphological, behavioral, physiological, and genetic adaptations, leading to low energy expenditure, likely enables giant pandas to survive on a bamboo diet.