1. Application of OrthoANI combined with MALDI-TOF MS in identification and typing of Campylobacter fetus
Song LI ; Pinghua QU ; Yanjiao YAO ; Cha CHEN ; Qiwei LI ; Guangyuan DENG ; Zhenjie XU ; Fu CHEN
Chinese Journal of Laboratory Medicine 2018;41(8):583-588
Objective:
Establishing the mass spectrum library of a new Campylobacter- " C.fetus subsp.testudinum" for rapid species identification in clinical microbiology laboratory.
Methods:
Illumina second generation sequencing platform 2000/miSeq was used to carry out high flux genome sequencing for the strains which were collected to establish mass spectrum library.The analysis oforthologous average nucleotide identity (OrthoANI) between collected strains and reference strains was performed at JAVA 8 operation environment. Then, the mass spectrums ofcollected strains andreference strains were acquired using MALDI-TOF MS. And the mass spectrum library of C. fetus subsp.testudinum. were established and verified.
Results:
The OrthoANI analysis showed that the OrthoANI value of the collected strains and the reference strain C. fetus subsp.testudinum03-427 was 99.30%-99.96%, while the OrthoANI values of collected strains and C. fetus subsp.venerealisNCTC10354 orC.fetus subsp.fetus82-40 were 91.05%-92.26%. With reference to OrthoANI ≥ 95% as the basis for the determination of the same strain, the strains which collected to establish mass spectrum library was finally identified as " C. fetus subsp.testudinum" . The identification accuracy rate of the mass spectrum library was 100% (consistent with gene sequencing), and the confidence interval was 82.3%-99.9%, identification of the same strain is 100% reproducible.
Conclusions
The new" gold standard" based on high throughput sequencing and total genome analysis has provided the ideal reference value for the establishment of mass spectrum library.And the accurate and objective reference spectrum of the" C.fetus subsp.testudinum" provides a new platform for the rapid diagnosis of fetal Campylobacter infection. (
2.Identification and classification of non-tuberculosis Mycobacterium by matrix-assisted laser desorption ionization-time of flight mass spectrometry
Song LI ; Yanjiao YAO ; Pinghua QU ; Cha CHEN ; Weizheng ZHANG ; Qiwei LI ; Zhenjie XU ; Fu CHEN
Chinese Journal of Laboratory Medicine 2018;41(2):109-115
Objective Reference standard of the RPOB(rifampin resistance)gene recommended by CLSI-MM18A(Interpretive Criteria for Identification of Bacteria and Fungi by DNA Target Sequencing) was used to evaluate the ability of MALDI-TOFMS techniques for the identification and classification of non-tuberculous Mycobacterium.Methods Fifty five clinicalstrains were collected from 2012 to 2016 with different sources.The RPOB gene was sequenced, and results were applied to phylogenetics analysis. MALDI-TOF MS technology was implemented to identify the strains, and cluster analysis was conducted based on protein fingerprint.The consistency of two methods for NTM identification and typing was evaluated.Results The RPOB gene method showed a good ability of identification(similarity>99.0%) and subtyping(to subspeciesof the complex level).The French BioMérieux MALDI-TOF MS identified 89.1% of 55 strains to genus level and 78.2% to species level.The phylogeneticsanalysis of protein fingerprint by SARAMS Premium software also showed good typing ability.Conclusions MALDI-TOF MS technology can identify and classify non-tuberculous Mycobacterium effectively,which is rapid and easy.It is complementary to RPOB gene method in laboratory application.
3.Characteristics of skin microbiota in an atopic dermatitis-like mouse model induced by 2,4-dinitrochlorobenzene
Yuehua MEN ; Huipu HAN ; ·Alifu AKADE ; Yifan YAO ; Tingting HUANG ; Jiarui KANG ; Tieshan WANG ; Mingbo ZHANG ; Yanjiao JU ; Ming CHU ; Yuedan WANG
Chinese Journal of Dermatology 2021;54(7):605-611
Objective:To investigate characteristics and changes of skin microbiota in atopic dermatitis-like mouse models induced by different concentrations of 2,4-dinitrochlorobenzene (DNCB) .Methods:Totally, 30 male specific-pathogen-free BALB/c mice were randomly divided into 3 groups by using a random number table: negative control group topically treated with 200 μl of mixture of acetone and olive oil at a volume ratio of 3∶1 on the back twice a week for 6 consecutive weeks; high-and low-concentration DNCB groups both topically treated with 200 μl of 1% DNCB on the first and third day at the first week, followed by topical application of 200 μl of 0.5% and 0.1% DNCB, respectively, twice a week for 5 weeks from the second week. Twenty-four hours after the last treatment, the severity of skin lesions was evaluated, and the transepidermal water loss and stratum corneum hydration were measured. After the experiment, the mice were sacrificed, and skin tissues were resected from the back of the mice for histopathological examination. Full-thickness skin tissue samples were obtained from the back of 3 mice in each group. Illumina Miseq PE300 high-throughput sequencing was performed to sequence the V3-V4 variable region of 16S rRNA gene of skin microbiota on the back of the mice, and the composition and structure of the skin microbiota and changes in the relative abundance of different genera were analyzed. One-way analysis of variance was used to analyze differences in indices among the 3 groups, and the Games-Howell method was used for multiple comparisons.Results:The severity scores of skin lesions were significantly higher in the high-and low-concentration DNCB groups (9.83 ± 2.45 points, 2.71 ± 0.56 points, respectively) than in the negative control group (0.51 ± 0.12 points, t=-7.19,-2.85, respectively, both P < 0.05) . Compared with the negative control group, the high-and low-concentration DNCB groups showed significantly increased transepidermal water loss ( t=-7.72,-2.68, respectively, both P < 0.05) , but significantly decreased stratum corneum hydration ( t=6.77, 5.99, respectively, both P < 0.05) ; the transepidermal water loss was significantly higher in the high-concentration DNCB group than in the low-concentration DNCB group ( t=2.76, P < 0.05) , while no significant difference in the stratum corneum hydration was observed between the high-and low-concentration DNCB groups ( P > 0.05) . There was a significant difference in the relative abundance of Corynebacterium among the 3 groups ( F=249.85, P < 0.001) , which was highest in the high-concentration DNCB group. No significant differences in the observed species and Chao1 index of the skin samples were observed among the 3 groups (both P > 0.05) , and the Shannon index was significantly lower in the high-concentration DNCB group than in the low-concentration DNCB group and negative control group ( t=6.96,-6.37, respectively, both P < 0.05) . Conclusion:DNCB could induce atopic dermatitis-like dermatitis in mice, and the severity of skin lesions and degree of barrier function impairment were related to the concentration of DNCB; the species diversity of skin microbiota markedly decreased in the high-concentration DNCB group, indicating that high-concentration DNCB modeling has more advantages in studying microbiological changes associated with atopic dermatitis.
4.Establishment of Drug Registration Applicant Compliance Management Organizations in the United States and Its Enlightenment to China
Yi YAO ; Yanjiao ZHAO ; Xin FAN ; Yongfa CHEN
China Pharmacy 2020;31(12):1414-1418
OBJECTIVE:To provide reference for further improving the establishment of drug registration applicant compliance management organizations in China. METHODS :Through introducing and analyzing the establishment ,responsibility configuration and operation mechanism of drug registration applicant compliance management organizations in the United States ,the suggestions were put forward for improving the establishment of drug registration applicant compliance management institutions in China. RESULTS & CONCLUSIONS :There were two levels of compliance management organizations for drug registration applicant in the United States ;at Office of the Commissioner of FDA level ,the Compliance Policy Council was responsible for ensuring consistency in the implementation of all compliance policies in the FDA regulated area ;at the level of Center for Drug Evalution and Research (CDER)and Office of Regulatory Affairs (ORA),the compliance management function was mainly undertaken by Office of New Drugs (OND),Office of Generic Drugs (OGD),Office of Compliance (OC) and Office of Communication (OCOMM)under CDER ,as well as Office of Policy and Risk Management (OPRM),OC,Office of Bioresearch Monitoring Operations(OBIMO)and Office of Pharmaceuticl Quality Operations (OPQO)under ORA ,which were responsible for developing compliance documents and procedures ,carrying out compliance education ,compliance guidance ,compliance inspection and punishment for violations. Before submission stage of drug registration application ,the compliance education function of OC and the compliance guidance function of OND/OGD played an important role ;in the application submission stage ,OND/OGD was responsible for the compliance review of application materials ;in the stage of reviewing the application materials ,OND/OGD and OC of CDER ensured drug compliance through collaboration with OC ,OBIMO and OPQO of ORA. The establishment of drug registration compliance management institutions in China was still in the preliminary exploration stage. It is suggested that China can learn from the relevant experience of the United States ,establish a multi-level or ganization structure ,set up an independent compliance education department , and strengthen thecooperation between evaluation department and compliance inspection department ,and to improve the level of compliance management of drug registration applicants in China.
5.Integrated Traditional Chinese and Western Medicine Treatment for Acute Lymphoblastic Leukemia Complicated with Severe COVID-19 in Recovery Stage: A Case Report
Xinglin GUO ; Jianzhu SHAO ; Jing JING ; Mingzhong XIAO ; Chongxiang XUE ; Qingwei LI ; Yanjiao ZHANG ; Chensi YAO ; Xuefei ZHAO ; Keyu CHEN ; Yingying YANG ; Xiuyang LI ; Yusheng BAI
Journal of Traditional Chinese Medicine 2023;64(23):2466-2470
This paper reported a case of severe COVID-19 in the recovery stage with acute lymphoblastic leukemia treated by integrated traditional Chinese and western medicine, with the intention of shedding light on the clinical diagnosis and treatment of similar conditions. The patient, who had acute lymphoblastic leukemia, developed COVID-19 infection during the bone marrow suppression period after chemotherapy. Treatment with western medicine was mainly anti-infection, symptomatic management, and supportive care. During the recovery stage, considering the patient's chemotherapy history and disease progression, the overall syndrome was identified as deficiency of both qi and yin and binding of phlegm and blood. Based on the “state-target” combined treatment strategy, herbal prescriptions were selected and modified to address the “deficiency state”, “disease target”, and “symptom target”. In addition to western medicine, the patient was administered with Shengmai Powder (生脉散) and Compound Zhebei Granules (复方浙贝颗粒) in its modifications to boost qi, nourish yin, and reinforce healthy qi, nourish and cool the blood, ultimately achieving satisfactory therapeutic effects.