Detection of new antibiotic resistance gene profile in Escherichia coliassociated with avian leukosis virus infection from broiler chickens

The Escherichia coli(E.coli)is prevailing worldwide,but the epidemiology of E.coli infections feature regional distribution characteristics to some extent.E.coli,as a zoonotic pathogen,can be transferred from animals to humans through food chain or via contact with wounds,causing a public health risk.We reported the swelling of proventriculus and tracheal bleeding following the death in two broiler chickens(Gallus gallus domesticus)from Beijing,China.To investigate whether a virus was involved in the infection,Madin Darby Bovine Kidney(MDCK)cells were co-cultured with supernatants of proventriculus,trachea and spleen homogenates.The avian leucosis virus was detected in the samples of proventriculus and trachea,but the avian influenza virus,the Newcastle disease virus and the avian infectious laryngotracheitis virus were not detected.E.coli isolates were resistant to almost all the antimicrobial as tested except for the combinations of amoxicillin/clavulanic acid and sulfamethoxazole/trimethoprim.PCR tests demonstrated the presence of antibiotic resistance genes in these E.coli isolates and further research revealed a novel gene profile with the presence of CTX-M-1,gyrA,gyrB,oqxA,oqxB,parC and Sul2 antibiotic resistance genes in a strain isolated from a proventriculus sample.These results demonstrated that the presence of antibiotic resistant E.coli would not necessarily cause outbreak of large-scale disease.However,when the bacteria carrying new antibiotic resistance genes enter the environment,it may result in the development of more virulent strains which will potentially impact human and animal health.

Characterization of multidrug-resistant Klebsiella pneumoniae isolated from the Chinese cobra Naja atra in a Beijing suburb

The emergence and spread of antibiotic resistance genes among Bacteria are a serious threat to global health.Their occurrence in animals which are in contact with humans is also important.The Chinese cobra(Naja atra,Elapidae),though a highly venomous species,is appreciated as food and as a source of materials used in traditional Chinese medicine.We are here reporting the isolation of multidrug-resistant Klebsiella pneumoniae(Enterobacteriaceae)from the lung of Naja atra,obtained from a snake farm in a Beijing suburb.Our study analyzed,using gene sequencing,the occurrence of antibiotic resistance genes(ARGs)in three K.pneumoniae isolates from two snakes.In addition,bacterial clones were identified by biochemical tests and phylogenetic analysis.Tests of antimicrobial susceptibility showed that all K.pneumoniae isolates were resistant to a host of antibiotics(piperacillin,cefazolin,gentamicin,tetracycline,doxycyclin,ciprofloxacin,levofloxacin,lomefloxacin,ofloxacin,norfloxacin,nalidixic acid,chloramphenicol,nitrofurantoin,sulfamethoxazole,and sulfamethoxazole/trimethoprim)but were susceptible to cefotaxime,cefixime,aztreonam,bramycin,amikacin,kanamycin,netilmicin,and streptomycin.Eighteen ARGs were detected in total DNA extracted from the isolates.Results showed three quinolone resistance genes(oqxA,oqxB,qnrB),the gyrA gene that confers resistance to beta-lactam antibiotics,and the emerging aac(3)-II gene that confers resistance to aminoglycosides.K.pneumoniae is an important opportunistic human pathogen and the emergence of multidrug-resistant K.pneumoniae in N.atra suggests the increasing risk of pathogen transmission between humans,livestock,and wildlife.Given the close association between foodborne pathogenic microorganisms and humans,it is key factor to identify these antibiotic resistance genes profile thereby minimize the risk of K.pneumoniae transmission.

Molecular evolution of Toll-like receptors in rodents

Toll-like receptors(TLRs),the key sensor molecules in vertebrates,trigger the innate immunity and prime the adaptive immune system.The TLR family of rodents,the largest order of mammals,typically contains 13 TLR genes.However,a clear picture of the evolution of the rodent TLR family has not yet emerged and the TLR evolutionary patterns are unclear in rodent clades.Here,we analyzed the natural variation and the evolutionary processes acting on the TLR family in rodents at both the interspecific and population levels.Our results showed that rodent TLRs were dominated by purifying selection,but a series of positively selected sites(PSSs)primarily located in the ligand-binding domain was also identified.The numbers of PSSs differed among TLRs,and nonviralsensing TLRs had more PSSs than those in viral-sensing TLRs.Gene-conversion events were found between TLR1 and TLR6 in most rodent species.Population genetic analyses showed that TLR2,TLR8,and TLR12 were under positive selection in Rattus norvegicus and R.tanezumi,whereas positive selection also acted on TLR5 and TLR9 in the former species,as well as TLR1 and TLR7 in the latter species.Moreover,we found that the proportion of polymorphisms with potentially functional change was much lower in viral-sensing TLRs than in nonviral-sensing TLRs in both of these rat species.Our findings revealed the first thorough insight into the evolution of the rodent TLR genetic variability and provided important novel insights into the evolutionary history of TLRs over long and short timescales.

Yersinia pestis: examining wildlife plague surveillance in China and the USA

Plague is a zoonotic disease caused by the bacterium Yersinia pestis Lehmann and Neumann,1896.Although it is essentially a disease of rodents,plague can also be transmitted to people.Historically,plague has caused mas-sive morbidity and mortality events in human populations,and has recently been classified as a reemerging dis-ease in many parts of the world.This public health threat has led many countries to set up wild and domestic an-imal surveillance programs in an attempt to monitor plague activity that could potentially spill over into human populations.Both China and the USA have plague surveillance programs in place,but the disease dynamics dif-fer in each country.We present data on plague seroprevalence in wildlife and review different approaches for plague surveillance in the 2 countries.The need to better comprehend plague dynamics,combined with the fact that there are still several thousand human plague cases per year,make well-designed wildlife surveillance pro-grams a critical part of both understanding plague risks to humans and preventing disease outbreaks in the future.

Genetic characterizations of Toll-like receptors in the brown rat and their associations with pathogen infections

Toll-like receptors(TLRs)are important initiators of innate immune responses that target host-pathogen interactions.However,further research into the molecular characteristics of TLRs in wild populations is required,as well as how TLRs genetically influenced pathogen infections in the brown rat(Rattus norvegicus).Here,we explored the genetic characterization and evolution of 2 sensing nucleic acid TLRs(TLR7 and TLR8)and 2 sensing nonnucleic acid TLRs(TLR2 and TLR4)in the wild brown rat,and assessed their associations with 2 RNA viruses(Seoul hantavirus and rat hepatitis E virus(HEV))and 2 bacteria(Leptospira and Bartonella).In these 4 TLRs,we discovered a total of 16 variants.Furthermore,TLR8 had high genetic diversity among 7 variants,while TLR2 had low genetic diversity with only 1 variant.According to selective pressure analyses,TLR4,TLR7,and TLR8 genes evolved under purifying selection.Interestingly,significant associations were found between 3 TLR8 variants and HEV infection,as well as 1 TLR2 variant and Bartonella infection.Overall,our findings provided a glimpse into the genetic characterization of TLRs in the brown rat,and further demonstrated that TLR2 and TLR8 genetic variations were related to Bartonella and HEV infection,respectively.Especially,TLR8 may be a good candidate immune gene for future research on molecular ecology and functional adaptation in wild populations.

Isolation and characterization of a Neisseria sp. from the captive wild goose (Anser anser)

The present study investigated 15 dead cases of captive wild goslings(Anser anser),which were bred in a small poultry farm in Shandong Province,China.The examined cases presented diverse clinical signs accompanied with neurological manifestations and fatal outcomes.Bacterial culture identified the gram-negative Neisseria sp.from the brain homogenate of most examined cases(10/15,66.7%).The isolated bacteria were identified based on morphologic characteristics,biochemical tests and 16S rDNA typing.Results proved that 1 identical bacteri­al strain(BNO09-3)was isolated from the positive cases.The phylogeny based on the 16S rDNA gene sequenc­es indicated that this isolate has a close relationship with various strains of genus Neisseria sp.isolated from liv­er and feces of duck.This is the first report of Neisseria sp.causing fatality in captive wild geese in China.

Effect of epidemic diseases on wild animal conservation

Under the background of global species extinction,the impact of epidemic diseases on wild animal protection is increasingly prominent.Here,we review and synthesize the literature on this topic,and discuss the relationship between diseases and biodiversity.Diseases usually reduce species diversity by decreasing or extinction of species populations,but also accelerate species evolution and promote species diversity.At the same time,species diversity can regulate disease outbreaks through dilution or amplification effects.The synergistic effect of human activities and global change is emphasized,which further aggravates the complex relationship between biodiversity and diseases.Finally,we emphasize the importance of active surveillance of wild animal diseases,which can protect wild animals from potential diseases,maintain population size and genetic variation,and reduce the damage of diseases to the balance of the whole ecosystem and human health.Therefore,we suggest that a background survey of wild animal populations and their pathogens should be carried out to assess the impact of potential outbreaks on the population or species level.The mechanism of dilution and amplification effect between species diversity and diseases of wild animals should be further studied to provide a theoretical basis and technical support for human intervention measures to change biodiversity.Most importantly,we should closely combine the protection of wild animals with the establishment of an active surveillance,prevention,and control system for wild animal epidemics,in an effort to achieve a win-win situation between wild animal protection and disease control.

Novel human H7N9 influenza virus in China

Outbreaks of H7N9 avian influenza in humans in 5 provinces and 2 municipalities of China have reawakened concern that avian influenza viruses may again cross species barriers to infect the human population and thereby initiate a new influenza pandemic.Evolutionary analysis shows that human H7N9 influenza viruses originated from the H9N2,H7N3 and H11N9 avian viruses,and that it is as a novel reassortment influenza virus.This ar­ticle reviews current knowledge on 11 subtypes of influenza A virus from human which can cause human infec­tions.

Identification and genetic characterization of Contracaecum sp.(Nematoda:Anisakidae)from China

Contracaecum species are economically important fish-borne larval nematodes with zoonotic significance.In June 2019,more than 100 piscivorous birds died in their habitats close to the Wild Duck Lake,located in Yanqing,northwest of Beijing,China.Post-mortem examination of Black Night Herons(Nycticorax)revealed the presence of numerous anisakid nematodes in the proventriculus.Recovered nematodes were identified as Contracaecum sp.based on morphological description.Phylogenetic analysis of the mitochondrial(mt)genome and the ITS gene showed that sequences of Contracaecum sp.Beijing isolates were grouped into a new individual cluster.Furthermore,the parasite was successfully isolated from fresh dead birds,feces of piscivorous birds,and fish and prevalence ranged from 8.0%to 81.8%.Consequently,our study demonstrated Contracaecum sp.infections in different sources from China,which might constitute a threat to wildlife,aquaculture,and public health.

Different infection routes of avian influenza A (H5N1) virus in mice

The continuing outbreaks of avian influenza A H5N1 virus infection in Asia and Africa have caused worldwide concern because of the high mortality rates in poultry,suggesting its potential to become a pandemic influenza virus in humans.The transmission route of the virus among either the same species or different species is not yet clear.Broilers and BABL/c mice were inoculated with the H5N1 strain of influenza A virus isolated from birds.The animals were inoculated with 0.1 mL 106.83 TCID50 of H5N1 virus oronasally,intraperitoneally and using eye drops.The viruses were examined by virological and pathological assays.In addition,to detect horizontal transmission,in each group,healthy chicks and mice were mixed with those infected.Viruses were detected in homogenates of the heart,liver,spleen,kidney and blood of the infected mice and chickens.Virus antigen was not detected in the spleen,kidney or gastrointestinal tract,but detected by Plaque Forming Unit(PFU)assay in the brain,liver and lung without degenerative change in these organs(in the group inoculated using eye drops.The detection results for mice inoculated using eye drops suggest that this virus might have a different tissue tropism from other influenza viruses mainly restricted to the respiratory tract in mice.All chicken samples tested positive for the virus,regardless of the method of inoculation.Avian influenza A H5N1 viruses are highly pathogenic to chickens,but its virulence in other animals is not yet known.To sum up,the results suggest that the virus replicates not only in different animal species but also through different routes of infection.In addition,the virus was detection not only in the respiratory tract but also in multiple extra-respiratory tissues.This study demonstrates that H5N1 virus infection in mice can cause systemic disease and spread through potentially novel routes within and between mammalian hosts.

Importance of collaboration on wildlife disease issues

Wildlife species are important components in disease transmission and persistence.They serve a critical role when viewing disease from a One Health perspective(One Health Initiative 2009).Although often misunderstood or underestimated,disease issues within wildlife species could greatly impact concerns in domestic animal or human health.Recent outbreaks of some zoonotic diseases can be traced,at least in part,to wildlife species.Beyond the potential threat to humans and agricultural resources,wildlife diseases also threaten biodiversity and animal welfare within natural ecosystems.