Identification of extended-spectrum beta-lactamase (CTX-M)-producing Klebsiella pneumoniae belonging to ST37, ST290, and ST2640 in captive giant pandas.

BACKGROUND: Extended-spectrum ß-lactamases (ESBL)-producing strains of Klebsiella pneumoniae remain a worldwide, critical clinical concern. However, limited information was available concerning ESBL-producing Klebsiella pneumoniae in giant pandas. The objective of this study was to characterize ESBL-producing Klebsiella pneumoniae isolates from captive giant pandas. A total of 211 Klebsiella pneumoniae isolates were collected from 108 giant pandas housed at the Chengdu Research Base of Giant Panda Breeding (CRBGP), China. Samples were screened for the ESBL-producing phenotype via the double-disk synergy test. RESULT: A total of three (1.42%, n = 3/211) ESBL-producing Klebsiella pneumoniae strains were identified, and characterization of ESBL-producing Klebsiella pneumoniae isolates were studied by the detection of ESBL genes and mobile genetic elements (MGEs), evaluation of antimicrobial susceptibility and detection of associated resistance genes. Clonal analysis was performed by multi-locus sequencing type (MLST). Among the three ESBL-producing isolates, different ESBL-encoding genes, including blaCTX-M, and blaTEM, were detected. These three isolates were found to carry MGEs genes (i.e., IS903 and tnpU) and antimicrobial resistance genes (i.e., aac(6')-Ib, aac(6')-I, qnrA, and qnrB). Furthermore, it was found that the three isolates were not hypermucoviscosity, resistant to at least 13 antibiotics and belonged to different ST types (ST37, ST290, and ST2640). CONCLUSION: Effective surveillance and strict infection control strategies should be implemented to prevent outbreaks of ESBL-producing Klebsiella pneumoniae in giant pandas.

Mortality analysis of captive red panda cubs within Chengdu, China.

BACKGROUND: The red panda has been classified as an endangered species due to the decreased number in the world and disease is considered as a great threat to the health and survival of the cubs in captivity. RESULTS: This study analyzed 32 red panda cub mortalities (15 females and 17 males, age less than two months) through gross necropsy, microbiological examination, and histopathological observation at the Chengdu Research Base of Giant Panda Breeding, China, during 2014-2020. The results showed that screenings for canine distemper virus, canine parvovirus, rotavirus and parasite infection were all negative, however bacteria such as Klebsiella pneumoniae, Proteus mirabilis, Escherichia coli, Enterococcus faecalis, Pseudomonas were isolated from the tissue samples of some cubs. The major causes of death were respiratory (43.75%) and digestive system disease (28.13%), followed by cardiovascular disease (12.5%) and neonatal stillbirths (9.38%). Renal system diseases and trauma were also detected, at lower incidence (one case for each). The mortality rate within 15 days of birth was 68.75% and gradually decreased with age, there was no significant difference in gender. CONCLUSION: This study can provide a scientific basis for the analysis of the cause of death among red panda cubs in captivity, so as to improve the survival rate, help build the captive population and further the ex-situ conservation management of this endangered species. Additionally, our research may also provide insights into the in-situ conservation of wild red pandas by identifying emerging disease threats within the wild population and potential treatment for rescued individuals.

Isolation and characterization of Uropathogenic Escherichia coli (UPEC) from red panda (Ailurus fulgens).

BACKGROUND: Disease prevention and control is a significant part in the ex-situ conservation of the endangered red panda (Ailurus fulgens), being bacterial infection is one of the most important health threats to the captive population. To date, studies about the infection caused by Escherichia coli in the red panda are scarce. This study was conducted to determine the cause of death of a captive red panda through clinical symptoms, complete blood count, biochemical analysis, pathological diagnosis and bacterial whole genome sequencing. CASE PRESENTATION: The following report describes a case of a 1.5 year old captive red panda (Ailurus fulgens) that was found lethargic and anorectic. She was moved to the quarantine area for daily treatment with 50 mg of Cefpodoxime Proxetil. During the three-day treatment, she did not eat or defecate, and then died. Clinical hematology revealed the values of neutrophils, alanine aminotransferase (ALT), aspartate aminotransferase (AST) and blood urea nitrogen (BUN) were significantly higher. Histological analysis demonstrated major pathological damage in the kidneys, liver and lungs, characterized by hyperemia, parenchymal cell degeneration and necrosis and inflammatory cell infiltration which were predominantly neutrophilic. A bacterial strain confirmed as Escherichia coli was isolated post mortem. Whole genome sequencing of the E. coli showed the complete genome size was 4.99 Mbp. PapA, PapC, OmpA, OmpU and other virulence factors which specific to Uropathogenic Escherichia coli (UPEC) were found in the isolate. Among the virulence factors, P pili, type I pili and related factors of the iron uptake system were associated with nephrotoxicity. CONCLUSION: The red panda died of bacterial infection caused by an uropathogenic strain of Escherichia coli. The pathogenic mechanisms of the strain are closely related to the expression of specific virulence genes.

High-throughput sequencing and characterization of potentially pathogenic fungi from the vaginal mycobiome of giant panda (Ailuropoda melanoleuca) in estrus and non-estrus.

Introduction: The giant panda (Ailuropoda melanoleuca) reproduction is of worldwide attention, and the vaginal microbiome is one of the most important factors affecting the reproductive rate of giant pandas. The aim of this study is to investigate the diversity of vaginal mycobiota structure, and potential pathogenic fungi in female giant pandas during estrus and non-estrus. Methods: This study combined with high-throughput sequencing and laboratory testing to compare the diversity of the vaginal mycobiota in giant pandas during estrus and non-estrus, and to investigate the presence of potentially pathogenic fungi. Potentially pathogenic fungi were studied in mice to explore their pathogenicity. Results and discussion: The results revealed that during estrus, the vaginal secretions of giant pandas play a crucial role in fungal colonization. Moreover, the diversity of the vaginal mycobiota is reduced and specificity is enhanced. The abundance of Trichosporon and Cutaneotrichosporon in the vaginal mycobiota of giant pandas during estrus was significantly higher than that during non-estrus periods. Apiotrichum and Cutaneotrichosporon were considered the most important genera, and they primarily originate from the environment owing to marking behavior exhibited during the estrous period of giant pandas. Trichosporon is considered a resident mycobiota of the vagina and is an important pathogen that causes infection when immune system is suppressed. Potentially pathogenic fungi were further isolated and identified from the vaginal secretions of giant pandas during estrus, and seven strains of Apiotrichum (A. brassicae), one strain of Cutaneotrichosporon (C. moniliiforme), and nine strains of Trichosporon (two strains of T. asteroides, one strain of T. inkin, one strain of T. insectorum, and five strains of T. japonicum) were identified. Pathogenicity results showed that T. asteroides was the most pathogenic strain, as it is associated with extensive connective tissue replacement and inflammatory cell infiltration in both liver and kidney tissues. The results of this study improve our understanding of the diversity of the vaginal fungi present in giant pandas and will significantly contribute to improving the reproductive health of giant pandas in the future.

Hematological and biochemical parameters of giant pandas (Ailuropoda melanoleuca) in captive and semi-natural environments.

Physiological indexes like blood parameters have been widely used to monitor the health of free-roaming animals. Attempts to reintroduce one of China's most endangered species, the giant panda (Ailuropoda melanoleuca), have been hampered by a lack of data on its ecology and physiology. We examined three giant pandas' hematological and blood chemistry parameters in a soft release program and 30 captive giant pandas as controls and determined the reference intervals (RIs) for those blood parameters in the captive animals. Elevation, captivity status and the interaction of those factors were statistically significant for hematologic measures. Release pandas had significantly higher hemoglobin and hematocrit values after they moved to high elevation locations. We also found significant difference in the enzyme parameters between high and low elevation pandas such as higher aspartate aminotransferase, alanine aminotransferase, creatinine kinase, amylase and lower lactate dehydrogenase and alkaline phosphatase. Release pandas also had higher nutrition parameter values such as higher albumin, globulin and creatinine. The RI for blood parameters in our study provides a baseline to monitor the health of captive animals and forms the basis for assessing the health of free-roaming giant pandas in future reintroduction efforts.

Exogenous ARC down-regulates caspase-3 expression and inhibits apoptosis of broiler chicken cardiomyocytes exposed to hydrogen peroxide.

Apoptosis repressor with caspase recruitment domain (ARC) is highly involved in apoptosis induced by oxidative stress or ischaemia/reperfusion injury. Furthermore, even though the exact mechanism is still unknown, some studies suggest that exogenous ARC also possesses anti-apoptotic ability. The study investigated whether mouse-derived ARC acquires anti-apoptotic ability and the pathway of regulation in chick embryo cardiomyocytes. To evaluate whether mouse-derived ARC can inhibit chick embryo cardiomyocyte apoptosis induced by hydrogen peroxide, recombinant pcDNA3.1/ARC plasmid was acquired and transfected into chick embryo cardiomyocytes. ARC-related gene (caspase-2, caspase-8, caspase-3, and caspase-9, cytochrome C, bcl-2, and XIAP) mRNA and protein expression levels were detected by real-time polymerase chain reaction and western blotting, respectively. Here we demonstrate that hydrogen peroxide induced apoptosis in chick embryo cardiomyocytes in a time-dependent manner and that this effect could be suppressed by mouse-derived ARC expression. Moreover, unlike endogenous ARC, exogenous ARC was exclusively expressed in the cytoplasm and down-regulated caspase-2, caspase-8, and caspase-3, bcl-2, and XIAP gene expression levels. However, only caspase-3 protein levels were decreased. In addition, threonine 149 phosphorylation by CK2 was required for exogenous ARC to exert an anti-apoptotic effect in chicken embryo cardiomyocytes and suggested exogenous ARC may in part share the same pathway of regulation with endogenous ARC. These results indicate that mouse-derived ARC plays an important role in protection of chick embryo cardiomyocytes against oxidative stress apoptosis by inhibiting caspase-3 mRNA and protein expression levels.

First Isolation and Identification of Aeromonas veronii in a Captive Giant Panda (Ailuropoda melanoleuca).

The objective of this study was to understand biological characteristics of one bacteria strain named as VPG which was isolated from multiple organs of a dead captive giant panda cub. Here, we use biochemical tests, 16S rRNA and gyrB genes for bacterial identification, the disk diffusion method for antibiotic resistance phenotype, smart chip real-time PCR for the antibiotic resistance genotype, multiplex PCR for determination of virulence genes, and the acute toxicity test in mice for testing the pathogenicity of isolates. The isolate was identified as A. veronii strain based on the biochemical properties and genetic analysis. We found that the strain carried 31 antibiotic resistance genes, revealed antimicrobial resistance phenotypically to several antibiotics including penicillin, ampicillin, oxacillin, amoxicillin, imipenem, and vancomycin, and carried virulence genes including aer, act, lip, exu, ser, luxs, and tapA. The main pathological changes in giant panda were congestion, necrotic lesions and a large number of bacteria in multiple organs. In addition, the LD50 in Kunming mice infected with strain VGP was 5.14 × 107 CFU/mL by intraperitoneal injection. Infection with strain VGP led to considerable histological lesions such as hemorrhage of internal organs, necrosis of lymphocytes and neurons in Kunming mice. Taken together, these results suggest that infection with strain VGP would be an important causes of death in this giant panda cub.

Occurrence and Molecular Characteristics of Microsporidia in Captive Red Pandas (Ailurus fulgens) in China.

Enterocytozoon bieneusi and Encephalitozoon spp. are microsporidian pathogens with zoonotic potential that pose significant public health concerns. To ascertain the occurrence and genotypes of E. bieneusi and Encephalitozoon spp., we used nested PCR to amplify the internal transcribed spacer (ITS) gene and DNA sequencing to analyze 198 fecal samples from red pandas from 6 zoos in China. The total rate of microsporidial infection was 15.7% (31/198), with 12.1% (24/198), 1.0% (2/198), 2.0% (4/198) and 1.0% (2/198) for infection rate of E. bieneusi, Encephalitozoon cuniculi, Encephalitozoon intestinalis and Encephalitozoon hellem, respectively. One red panda was detected positive for a mixed infection (E. bieneusi and E. intestinalis). Red pandas living in semi-free conditions are more likely to be infected with microsporidia (χ2 = 6.212, df = 1, p < 0.05). Three known (SC02, D, and PL2) and one novel (SCR1) genotypes of E. bieneusi were found. Three genotypes of E. bieneusi (SC02, D, SCR1) were grouped into group 1 with public health importance, while genotype PL2 formed a separate clade associated with group 2. These findings suggest that red pandas may serve as a host reservoir for zoonotic microsporidia, potentially allowing transmission from red pandas to humans and other animals.

Gut phageome of the giant panda (Ailuropoda melanoleuca) reveals greater diversity than relative species.

The gut flora is a treasure house of diverse bacteriophages maintaining a harmonious and coexistent relationship with their hosts. The giant panda (Ailuropoda melanoleuca), as a vulnerable endemic species in China, has existed for millions of years and is regarded as a flagship species for biodiversity conservation. And yet, limited studies have analyzed the phage communities in the gut of giant pandas. Using viral metagenomic analysis, the phageomes of giant pandas and other relative species were investigated. Our study explored and compared the composition of phage communities from different animal sources. Giant pandas possessed more diverse and abundant phage communities in the gut compared with other relevant animals. Phylogenetic analyses based on the phage terminase large subunit (TerL) showed that the Caudovirales phages in giant pandas also presented highly genetic diversity. Our study revealed the diversity of phage communities in giant pandas and other relative species, contributing to the health maintenance of giant pandas and laying the groundwork for molecular evolution research of bacteriophages in mammals. IMPORTANCE Gut phageome plays an important role in shaping gut microbiomes by direct interactions with bacteria or indirect influences on the host immune system, potentially regulating host health and disease status. The giant panda (Ailuropoda melanoleuca) is a vulnerable and umbrella species for biodiversity conservation. Our work explored and compared the gut phageome of giant pandas and relative species, contributing to the health maintenance of giant pandas.

A snapshot of climate drivers and temporal variation of Ixodes ovatus abundance from a giant panda living in the wild.

Ticks and tick-borne diseases have negative impacts on the health of wild animals including endangered and vulnerable species. The giant panda (Ailuropoda melanoleuca), a vulnerable and iconic flagship species, is threatened by tick infestation as well. Not only can ticks cause anemia and immunosuppression in the giant panda, but also bacterial and viral diseases. However, previous studies regarding tick infestation on giant pandas were limited in scope as case reports from sick or dead animals. In this study, an investigation focusing on the tick infestation of a reintroduced giant panda at the Daxiangling Reintroduction Base in Sichuan, China was conducted. Ticks were routinely collected and identified from the ears of the giant panda from March to September in 2021. A linear model was used to test the correlation between tick abundance and climate factors. All ticks were identified as Ixodes ovatus. Tick abundance was significantly different among months. Results from the linear model showed temperature positively correlated to tick abundance, while air pressure had a negative correlation with tick abundance. To the best of our knowledge, this study is the first reported investigation of tick species and abundance on a healthy giant panda living in the natural environment, and provides important information for the conservation of giant pandas and other species sharing the same habitat.

Is skin pressure in load carriage over-evaluated?

Skin pressure is a biomechanical measure widely used in the assessment of load carriage systems. However, because of the complicated contour of human body, the stiffness of the pressure sensor array, and the large range of measurable pressures, there is much variability in previously reported results. In this paper, a simple mechanical model for load carriage was proposed, and the skin pressures beneath the shoulder and hip straps were predicted from the strap forces based on Laplace's law. The proposed model was used to analyze data from literature with an aim to check the reliability of existing pressure measurements. The static and dynamic pressures at five locations on eight subjects wearing a backpack with a 10 kg load, while standing and walking on a treadmill, were measured respectively using pressure sensors of the air pack type. The combination of literature data analysis and experimental testing proved that the existing measurement method of interface pressure in load carriage systems often leads to over-estimation and this might misguide the pressure criteria set for load carriage system usage and design. The proposed model will be useful for quick prediction of the interface pressure in load carriage systems.

Surveillance Study of Klebsiella pneumoniae in the Giant Panda Revealed High Genetic Diversity and Antibiotic Therapy Challenge.

Klebsiella pneumoniae is not only a worldwide human pathogen, it also effects wildlife, such as the giant panda (Ailuropoda melanoleuca), in which it has recently been evidenced to result in diarrhea, organ failure, and even death. A K. pneumoniae investigation was carried out at the Chengdu Research Base of Giant Panda Breeding in 2018. As part of the investigation, the pulsed-field gel electrophoresis (PFGE) typing, multilocus-sequence typing (MLST), antibiotic resistance profiles (ARPs), and antibiotic resistance genes (ARGs) were studied based on all isolates. Fecal samples were collected from 72 A. melanoleuca from May to December 2018, and a total of 90 K. pneumoniae were isolated from 153 fecal samples. The genotyping results showed that the isolates had high diversity, of which 84 clusters were obtained by PFGE and 57 STs by MLST. The overall trend of the similarity of isolates was the first sample period > second sample period > third sample period, which showed the increasement of genome variability of K. pneumoniae. In addition, 90 isolates showed high resistance to ampicillin, rifampicin, and compound sulfamethoxazole. Of the obtained isolates, 50% carried 6~8 ARPs, and the carrying volume increased during three sample periods, in which we found two isolates carrying 12 and 13 ARPs during the third sample period, respectively. Moreover, a total of 65 ARGs were detected (90.28%, 65/72) in 90 K. pneumoniae samples. Almost all bacteria sampled contained 17 ARGs that belonged to the ß-lactamase, Multidrug, MGEs, Aminoglycoside, and Tetracycline, which may be the basis of ARPs of K. pneumoniae. Moreover, the types of Multidrug and MGEs had a greater impact on antibiotic susceptivity of K. pneumoniae. Our results showed that K. pneumoniae has a serious risk of transmission in A. melanoleuca and K. pneumoniae had a high possibility of genome diversity and the risk of drugs tolerance under the large antibiotic usage.

Comparison of a commercial ELISA and indirect hemagglutination assay with the modified agglutination test for detection of Toxoplasma gondii antibodies in giant panda (Ailuropoda melanoleuca).

Toxoplasma gondii is a worldwide-distributed zoonotic protozoan parasite which causes toxoplasmosis and has a significant effect on public health. In the giant panda (Ailuropoda melanoleuca), toxoplasmosis can cause asymptomatic infections, reproductive disorder and even death, which poses a serious threat to the conservation of this rare protected species. Therefore, serological investigation of T. gondii is essential to understanding its risk to giant pandas, however, there are no specific testing kits for giant pandas. Previous research has used MAT as the reference method for screening T. gondii, to investigate this further, this study focused on the agreement comparing of MAT with ELISA and IHA tests for detecting T. gondii antibodies in 100 blood samples from 55 captive giant pandas in Chengdu, China. The results showed 87.0%, 87.0%, 84.0%, samples were sero-positive for T. gondii using ELISA (kits a, b, c), respectively, while MAT and IHA tests were 84.0% and 9.0% sero-positive, respectively. There was no significant difference between MAT and the three ELISA kits and these two methods had substantial agreement (0.61 < Ò› ≤ 0.80). Meanwhile, there was a significant difference (P < 0.001) between MAT and IHA, and these two methods had only a slight agreement (қ ≤ 0.20). The relative sensitivity of the ELISA (kits a, b, c) were 89.0%, 91.5% and 95.1%, and the specificity were 86.7%, 80.0% and 80.0%, respectively, which showed these three ELISA kits all had great accuracy. It is suggested that MAT is the recommended test method for primary screening T. gondii in giant pandas and then verified by ELISA.

Viral metagenomics unveiled extensive communications of viruses within giant pandas and their associated organisms in the same ecosystem.

Cross-species transmission events were commonplace, with numerous cases of host-switching during the viral evolutionary history, but relatively little evidence for onward transmission in different species living in the same ecosystem. For understanding the communications of viruses in giant pandas (Ailuropoda melanoleuca) and their associated organisms, based on a large size of samples (N = 2305) collected between 2015 and 2020 from giant panda (N = 776) and other four giant panda-associated organisms in the same ecosystem, red pandas (N = 700), stray cats (N = 32), wild rats (N = 42), and mosquitoes (N = 755), viromics was used for the virus identification and subsequent virus traceability. The results showed that a feline panleukopenia virus (FPV) was found in giant pandas with clinical signs of vomiting and mild diarrhea. Meanwhile, the same FPV strain was also prevalent in the healthy red panda (Ailurus fulgens) population. From the viromes of the five different organisms, 250 virus genomes were determined. Our data revealed that besides FPV, other putative pathogenic viruses, such as red panda amdoparvoviruses (RPAVs) and Getah viruses (GETVs) were responsible for previous disease or death of some red pandas. We also demonstrated that a number of viruses were involved in potential interspecies jumping events between giant pandas and their associated species. Collectively, our results shed light on the genetic diversity and relationship of diverse viral pathogens in 'Giant pandas-Associated animals-Arthropods' and report some cases of possible viral host-switching among these host species living in the same ecosystem.

Seroprevalence and risk factors of Toxoplasma gondii infection in captive giant panda (Ailuropoda melanoleuca).

Introduction: Toxoplasma gondii, a globally zoonotic protozoan parasite, infects most warm-blooded animals including the giant panda, and poses a serious threat to the giant panda conservation. However, the seroprevalence and the risk factors for toxoplasmosis in giant pandas are unknown. Here we aimed to determine the seroprevalence of T. gondii in the captive population of giant pandas and analyze the factors associated with the increased risk of infection. Methods: A total of 203 serum samples were collected from 157 (95 females and 62 males) captive giant pandas from 2007 to 2022, antibodies against T. gondii were screened using commercial ELISA and MAT kits. Results: The results showed 56 (35.67%) giant pandas were seropositive, age and transfer history between institutions were identifified as risk factors for T. gondii infection. It is suggested that age-related seroprevalence was the main factor, and housing multiple species in the same environment may increase the chance of cross-infection of T. gondii. Discussion: This study can provide research data for developing policies for the prevention and control of T. gondii and protecting the health of captive giant pandas and other wildlife.

Virome of Giant Panda-Infesting Ticks Reveals Novel Bunyaviruses and Other Viruses That Are Genetically Close to Those from Giant Pandas.

Tick infestations have been reported as one of the factors threatening the health of giant pandas, but studies of viral pathogens carried by ticks feeding on the blood of giant pandas are limited. To assess whether blood-sucking ticks of giant pandas can carry viral pathogens and if so, whether the viruses in ticks are associated with those previously detected in giant panda hosts, we determined the viromes of ticks detached from giant pandas in a field stocking area in Sichuan Province, southwest China. Using viral metagenomics we identified 32 viral species in ticks, half of which (including anellovirus [n = 9], circovirus [n = 3], and gemycircularvirus [n = 4]) showed homology to viruses carried by giant pandas and their associated host species (such as red pandas and mosquitoes) in the same living domain. Remarkably, several viruses in this study phylogenetically assigned as bunyavirus, hepe-like virus, and circovirus were detected with relatively high abundance, but whether these newly identified tick-associated viruses can replicate in ticks and then transmit to host animals during a blood meal will require further investigation. These findings further expand our understanding of the role of giant panda-infesting ticks in the local ecosystem, especially related to viral acquisition and transmission, and lay a foundation to assess the risk for giant panda exposure to tick-borne viruses. IMPORTANCE Ticks rank only second to mosquitoes as blood-feeding arthropods, capable of spreading pathogens (including viruses, bacteria, and parasites) to hosts during a blood meal. To better understand the relationship between viruses carried by ticks and viruses that have been reported in giant pandas, it is necessary to analyze the viromes of giant panda-parasitic blood-sucking ticks. This study collected 421 ticks on the body surface of giant pandas in Sichuan Province, China. We characterized the extensive genetic diversity of viruses harbored by these ticks and reported frequent communication of viruses between giant pandas and their ticks. While most of the virome discovered here are nonpathogenic viruses from giant pandas and potentially tick-specific viruses, we revealed some possible tick-borne viruses, represented by novel bunyaviruses. This research contributes to the literature because currently there are few studies on the virome of giant panda-infesting ticks.

First report of fatal baylisascariasis-induced acute pancreatitis in a giant panda.

A wild adult male giant panda that was rescued from a nature reserve in Sichuan Province, China, has died. The panda had been in poor physical condition: it was wheezing and had increased serum amylase. A pathological examination was performed in order to determine the cause of death. Gross examination revealed 1380 mL of yellowish fluid in the abdominal cavity, 356 nematodes in the digestive tract and one filling the pancreatic duct, contractions and variably-sized dark purple areas in the spleen, a collapsed right lung and consolidation of the left lung. Acute pancreatitis was confirmed histopathologically via edema, focal necrosis and hemorrhage with inflammatory cell infiltration. Other major histopathological changes included serous-hemorrhagic pneumonia, lymphocytic necrosis and depletion in the spleen, and degeneration and necrosis of renal tubular epithelial cells. The nematodes were identified as Baylisascaris schroederi via molecular assays. In conclusion, the cause of death of the giant panda was determined to be multiple organ dysfunction syndrome caused by baylisascariasis-induced acute pancreatitis. To our knowledge, this is the first report of fatal baylisascariasis-induced acute pancreatitis in the giant panda.

High Prevalence of Antimicrobial Resistance and Integron Gene Cassettes in Multi-Drug-Resistant Klebsiella pneumoniae Isolates From Captive Giant Pandas (Ailuropoda melanoleuca).

Multi-drug-resistant Klebsiella pneumoniae (MDR K. pneumonia) is increasingly being reported with corresponding increase in morbidity and mortality all over the world. However, limited information is available concerning MDR K. pneumonia in giant pandas. The objective of this study was to grasp the drug resistance profile of MDR K. pneumonia isolated from giant pandas. A total of 182 K. pneumoniae isolates were collected from fresh feces of 94 captive giant pandas of different ages and sex and separated by season. We performed a standard disk diffusion antimicrobial susceptibility test with the isolates and further evaluated the antibiotic resistance genes (ARGs) of multi-drug-resistant strains by high-throughput quantitative PCR. In addition, we then analyzed mobile genetic elements (MGEs), integron gene cassettes, and the multi-locus sequence typing of multi-drug-resistant strains by PCR. Antimicrobial susceptibility testing results demonstrated that a total of 30 (16.5%) K. pneumoniae isolates showed multiple drug resistances. The thirty MDR K. pneumonia isolates were mainly resistant to amoxicillin (100.0%), doxycycline (86.7%), chloramphenicol (60.0%), compound trimethoprim (60.0%) and trimethoprim (56.7%). Fifty different types of antibiotic resistance genes were found, which included a total of 671 antibiotic resistance genes, in the 30 multi-drug-resistant isolates. The top ten resistance genes were: vanTC-02, aacC, blaCTX-M-04, blaSHV-01, blaSHV-02, ampC-04, blaOXY, tetD, blaTEM and tetA-02. Thirteen mobile genetic elements were detected, of which IS26 (96.67%) and intI1 (96.67%) had the highest frequency. The thirty MDR K. pneumonia isolates were negative for the traA, traF, tnsA, IS1133, ISpa7, ISkpn6, intI2 and intI3 genes. Moreover, a further investigation of integrons revealed that two types of specific gene cassettes (dfrA12 + orfF + aadA2 and dfrA12 + orfF) were identified in class 1 integrons. Multi-locus sequence typing results showed that 22 STs in the thirty MDR K. pneumonia isolates were identified, the main type was ST37 (5/30). Our results illustrate that effective surveillance and strict biosecurity strategies should be taken to prevent the spread of multi-drug-resistant bacteria, and monitor the emergence of mobile genetic elements and integrons.

Feline Panleukopenia Virus With G299E Substitution in the VP2 Protein First Identified From a Captive Giant Panda in China.

A feline panleukopenia virus (FPV), Giant panda/CD/2018, was isolated from a captive giant panda with mild diarrhea in 2018 in Chengdu, China, and further identified via indirect immunofluorescence assay (IFA), transmission electron microscopy (TEM) observation, and genetic analysis. Phylogenetic analysis based on the complete VP2 nucleotide sequences showed that it shared high homology with Chinese FPV isolates and grouped within FPV cluster 1. One unique substitution Gly(G)299Glu(E) in the capsid protein VP2 was first identified with Giant panda/CD/2018. The presence of the G299E substitution is notable as it is located on the top region of the interconnecting surface loop 3, which may be involved in controlling the host range and antigenicity of FPV. These findings first demonstrate that FPV with natural point mutation G299E in the VP2 gene is prevalent in giant panda and suggest that etiological surveillance and vaccination among all giant pandas are urgently needed to protect this endangered species against FPV infection.

Contrasting Vaginal Bacterial Communities Between Estrus and Non-estrus of Giant Pandas (Ailuropoda melanoleuca).

Bacterial infection and imbalance of bacterial community in the genitourinary system of giant panda could affect the reproductive health. In severe cases, it can also lead to abortion. In this study, 13 of vaginal secretions in the estrue (E) group and seven of vaginal secretions in the non-estrue (NE) group were used to study the composition and diversity of vaginal bacterial communities between estrus and non-estrus by 16S rRNA gene sequencing analysis. The results showed that the vaginal microbiome in giant pandas shared the same top five abundant species between estrus and non-estrus at the phylum level. However, the vaginal microbiome changed significantly during estrus at the genus level. In top 10 genera, the abundance of Escherichia, Streptococcus, and Bacteroides in the E group was significantly higher than that in the NE group (p<0.05); Azomonas, Porphyromonas, Prevotella, Campylobacter, and Peptoniphilus in the NE group was significantly higher than that in the E group (p<0.05). The richness and diversity of vaginal microbiome in giant panda on estrus were significantly lower than those on non-estrus (p<0.05). It is noteworthy that the abundance of Streptococcus, Escherichia, and Bacteroides of vagina in giant pandas maintained low abundance in the daily. Whereas, they increased significantly during estrus period, which may play an important role in female giant pandas during estrus period. It was hypothesized that hormones may be responsible for the changes in the vaginal microbiome of giant pandas between estrus and no-estrus stages.