Lifeng Zhu

Evidence of cellulose metabolism by the giant panda gut microbiome

The giant panda genome codes for all necessary enzymes associated with a carnivorous digestive system but lacks genes for enzymes needed to digest cellulose, the principal component of their bamboo diet. It has been posited that this iconic species must therefore possess microbial symbionts capable of metabolizing cellulose, but these symbionts have remained undetected. Here we examined 5,522 prokaryotic ribosomal RNA gene sequences in wild and captive giant panda fecal samples. We found lower species richness of the panda microbiome than of mammalian microbiomes for herbivores and nonherbivorous carnivores. We detected 13 operational taxonomic units closely related to Clostridium groups I and XIVa, both of which contain taxa known to digest cellulose. Seven of these 13 operational taxonomic units were unique to pandas compared with other mammals. Metagenomic analysis using ∼37-Mbp contig sequences from gut microbes recovered putative genes coding two cellulose-digesting enzymes and one hemicellulose-digesting enzyme, cellulase, β-glucosidase, and xylan 1,4-β-xylosidase, in Clostridium group I. Comparing glycoside hydrolase profiles of pandas with those of herbivores and omnivores, we found a moderate abundance of oligosaccharide-degrading enzymes for pandas (36%), close to that for humans (37%), and the lowest abundance of cellulases and endohemicellulases (2%), which may reflect low digestibility of cellulose and hemicellulose in the pandas unique bamboo diet. The presence of putative cellulose-digesting microbes, in combination with adaptations related to feeding, physiology, and morphology, show that giant pandas have evolved a number of traits to overcome the anatomical and physiological challenge of digesting a diet high in fibrous matter.

Whole-genome sequencing of giant pandas provides insights into demographic history and local adaptation

The panda lineage dates back to the late Miocene and ultimately leads to only one extant species, the giant panda (Ailuropoda melanoleuca). Although global climate change and anthropogenic disturbances are recognized to shape animal population demography their contribution to panda population dynamics remains largely unknown. We sequenced the whole genomes of 34 pandas at an average 4.7-fold coverage and used this data set together with the previously deep-sequenced panda genome to reconstruct a continuous demographic history of pandas from their origin to the present. We identify two population expansions, two bottlenecks and two divergences. Evidence indicated that, whereas global changes in climate were the primary drivers of population fluctuation for millions of years, human activities likely underlie recent population divergence and serious decline. We identified three distinct panda populations that show genetic adaptation to their environments. However, in all three populations, anthropogenic activities have negatively affected pandas for 3,000 years.

Black and white and read all over: the past, present and future of giant panda genetics

Few species attract much more attention from the public and scientists than the giant panda (Ailuropoda melanoleuca), a popular, enigmatic but highly endangered species. The application of molecular genetics to its biology and conservation has facilitated surprising insights into the biology of giant pandas as well as the effectiveness of conservation efforts during the past decades. Here, we review the history of genetic advances in this species, from phylogeny, demographical history, genetic variation, population structure, noninvasive population census and adaptive evolution to reveal to what extent the current status of the giant panda is a reflection of its evolutionary legacy, as opposed to the influence of anthropogenic factors that have negatively impacted this species. In addition, we summarize the conservation implications of these genetic findings applied for the management of this high‐profile species. Finally, on the basis of these advances and predictable future changes in genetic technology, we discuss future research directions that seem promising for giant panda biology and conservation.

Significant genetic boundaries and spatial dynamics of giant pandas occupying fragmented habitat across southwest China

Understanding population history and genetic structure are key drivers of ecological research. Here, we studied two highly fragmented and isolated populations (Xiaoxiangling and Daxiangling) of giant pandas (Ailuropoda melanoleuca) at the extreme southwestern edge of their distribution. This area also contains the Dadu River, national road 108 and various human infrastructure and development, providing an ideal region in which we can identify the effects of different barriers on animal movements. We used partial mitochondrial control region (mtDNA) and nine microsatellite loci (nuclear DNA) data derived from 192 faecal and one blood sample collected from the wild. We found 136 genotypes corresponding to 53 unique multilocus genotypes and eight unique control region haplotypes (653 bp). Significant genetic boundaries correlated spatially with the Dadu River (K = 2). We estimate that a major divergence took place between these populations 26 000 years bp, at around the similar time the rock surface of valley bottom formed in Dadu River. The national road has resulted in further recent population differentiation (Pairwise FS on mtDNA and nuclear DNA) so that in effect, four smaller sub‐populations now exist. Promisingly, we identified two possible first‐generation migrants and their migration paths, and recommended the immediate construction of a number of corridors. Fortunately, the Chinese government has accepted our advice and is now planning corridor construction.

Conservation Implications of Drastic Reductions in the Smallest and Most Isolated Populations of Giant Pandas

In conservation biology, understanding the causes of endangerment is a key step to devising effective conservation strategies. We used molecular evidence (coalescent simulations of population changes from microsatellite data) and historical information (habitat and human population changes) to investigate how the most-isolated populations of giant pandas (Ailuropoda melanoleuca) in the Xiaoxiangling Mountains became highly endangered. These populations experienced a strong, recent demographic reduction (60-fold), starting approximately 250 years BP. Explosion of the human population and use of non-native crop species at the peak of the Qing Empire resulted in land-use changes, deforestation, and habitat fragmentation, which are likely to have led to the drastic reduction of the most-isolated populations of giant pandas. We predict that demographic, genetic, and environmental factors will lead to extinction of giant pandas in the Xiaoxiangling Mountains in the future if the population remains isolated. Therefore, a targeted conservation action--translocation--has been proposed and is being implemented by the Chinese government.

Thirty-three microsatellite loci for noninvasive genetic studies of the giant panda (Ailuropoda melanoleuca)

Limited microsatellite markers useable in noninvasive genetic methods have hampered the studies of dispersal patterns and mating systems of giant pandas. Therefore, we describe in this paper the characterization of 15 novel microsatellite loci from genomic DNA-enriched libraries and 18 redesigned microsatellite loci from published papers on the giant panda. The number of alleles per locus in 60 individuals ranged from 2 to 13, the average observed heterozygosity per locus from 0.168 to 0.800, and the average expected heterozygosity per locus from 0.152 to 0.882. All loci followed Hardy-Weinberg expectations. Four pairs of significant linkage association were found among all these loci. Moreover, the 33 microsatellite loci showed high amplification successes rate in noninvasive samples, which indicated that these loci will be of use in studying dispersal patterns and mating systems of giant pandas using noninvasive genetic methods.

Large-Scale Genetic Survey Provides Insights into the Captive Management and Reintroduction of Giant Pandas

The captive genetic management of threatened species strives to preserve genetic diversity and avoid inbreeding to ensure populations remain available, healthy, and viable for future reintroduction. Determining and responding to the genetic status of captive populations is therefore paramount to these programs. Here, we genotyped 19 microsatellite loci for 240 captive giant pandas (Ailuropoda melanoleuca) (∼64% of the captive population) from four breeding centers, Wolong (WL), Chengdu (CD), Louguantai (LGT), and Beijing (BJ), and analyzed 655 bp of mitochondrial DNA control region sequence for 220 of these animals. High levels of genetic diversity and low levels of inbreeding were estimated in the breeding centers, indicating that the captive population is genetically healthy and deliberate further genetic input from wild animals is unnecessary. However, the LGT population faces a higher risk of inbreeding, and significant genetic structure was detected among breeding centers, with LGT-CD and WL-BJ clustering separately. Based on these findings, we highlight that: 1) the LGT population should be managed as an independent captive population to resemble the genetic distinctness of their Qinling Mountain origins; 2) exchange between CD and WL should be encouraged because of similar wild founder sources; 3) the selection of captive individuals for reintroduction should consider their geographic origin, genetic background, and genetic contribution to wild populations; and 4) combining our molecular genetic data with existing pedigree data will better guide giant panda breeding and further reduce inbreeding into the future.

Isolation and characterization of 12 novel microsatellite loci for the red panda (Ailurus fulgens)

The information on dispersal patterns and mating systems of red pandas is quite important for the understanding of the genetic diversity and divergence of this species. And microsatellite marker is an ideal tool to analyze dispersal patterns and mating systems. Thus, we describe in this paper the isolation and characterization of 12 microsatellite loci in the red panda from genomic DNA-enriched libraries. These loci were highly polymorphic with numbers of alleles per locus in 24 individuals ranging from 2 to 14, observed heterozygosity from 0.143 to 0.864 and expected heterozygosity from 0.297 to 0.872. All loci except for RP6 locus followed Hardy–Weinberg expectations. No significant linkage association was found among all these loci. The 12 novel polymorphic microsatellite loci will be of use in studying dispersal patterns and mating systems of red pandas.

Seasonal variation in nutrient utilization shapes gut microbiome structure and function in wild giant pandas

Wild giant pandas use different parts of bamboo (shoots, leaves and stems) and different bamboo species at different times of the year. Their usage of bamboo can be classified temporally into a distinct leaf stage, shoot stage and transition stage. An association between this usage pattern and variation in the giant panda gut microbiome remains unknown. Here, we found associations using a gut metagenomic approach and nutritional analyses whereby diversity of the gut microbial community in the leaf and shoot stages was significantly different. Functional metagenomic analysis showed that in the leaf stage, bacteria species over-represented genes involved in raw fibre utilization and cell cycle control. Thus, raw fibre utilization by the gut microbiome was guaranteed during the nutrient-deficient leaf stage by reinforcing gut microbiome robustness. During the protein-abundant shoot stage, the functional capacity of the gut microbiome expanded to include prokaryotic secretion and signal transduction activity, suggesting active interactions between the gut microbiome and host. These results illustrate that seasonal nutrient variation in wild giant pandas substantially influences gut microbiome composition and function. Nutritional interactions between gut microbiomes and hosts appear to be complex and further work is needed.