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Science and technological advancements drive human progress, with laboratory animals serving as essential resources for developments in life sciences and medicine. However, the waste generated by these animals presents new challenges for urban management. Issues such as classification, recycling, effective utilization, and biohazard elimination must be addressed, necessitating the development of regulations, standards, and norms to keep pace with advancements. The construction of quality management system is the foundation and framework for the management of inspection and testing organizations. It should have strong operability and inspectability, enabling continuous improvement of the management level and enhancing the stability of basic management. However, current quality management systems often lack clarity in managing laboratory animal waste, including undefined disposal processes for non-medical institutions, inaccurate waste classification, and inadequate disposal methods for different waste categories. This paper addresses these challenges by identifying necessary processes to be added or removed in the quality management system of National Institutes for Food and Drug Control, developing effective SOPs, proposing practical measures to strengthen supervision and management, and integrating 6S management principles into our quality management system. In conclusion, effective management of laboratory animal waste should be centered on improving the quality management system, emphasizing waste classification and management at the source, controlling biological hazards, minimizing environmental pollution and promoting conditions for sustainable development.

ObjectiveTo understand the testing level of the relevant laboratories and standardize the detection of porcine parvovirus (PPV) nucleic acid by implementing laboratory proficiency testing plan of the PPV nucleic acid testing.MethodsFrom January to August 2022, NIFDC organized the laboratory proficiency testing plan for PPV nucleic acid detection (plan number: NIFDC-PT-364). According to relevant CNAS standards, inactivated PPV was added into the serum of PPV negative pigs to prepare proficiency testing samples. After passing the homogeneity and stability tests, the samples should be distributed to the participating laboratories. Each laboratory was distributed 3 samples and was required to conduct qualitative detection of PPV nucleic acid, and submit the results and relevant original records within the specified time. Finally, the results submitted by each laboratory were summarized and analyzed.ResultsA total of 9 laboratories registered to participate in this proficiency testing, and all of them fed back the results within the specified time. Among them, the results of 8 laboratories met the expectations, accounting for 88.9%. Three samples from one laboratory were all tested positive for PPV, which was not consistent with the expected results, and it was judged as unsatisfactory. Most laboratories adopted real-time fluorescent PCR detection method, and used domestic reagents for nucleic acid extraction, while the PCR amplification reagents were mainly imported reagents. The use of different brands of reagents didn't have impact on the detection results. There were some problems in some laboratories, such as unclear diagram of experimental results and incomplete information of original records.ConclusionMost of the participating laboratories are capable of PPV nucleic acid detection, but the detection details need to be improved.

Objective To analyze the status of proficiency testing samples in the field of laboratory animal testing, and to evaluate the role of the testing samples in the laboratory animal laboratory proficiency testing program.MethodsA retrospective study was conducted to compare the 30 proficiency testing plans and reports in the field of laboratory animal organized by National Institute for Food and Drug Control (NIFDC) from 2013 to 2022. The type, quantity, preparation method, homogeneity and stability of the proficiency testing samples, as well as the satisfaction of the participating units to the samples were analyzed. Then, the supply of new samples, the addition of quantitative projects, the improvement of stability research and the attention to customer needs were discussed.ResultsThe 30 proficiency testing programs included 10 items in the direction of bacteria, 10 items in the direction of virus and 10 items in the direction of genetics in the field of experimental animals. A total of 80 non-nucleic acid samples (including serums, feces, tracheal flushing fluid and proteins) and 4 nucleic acid samples were provided, all of which were qualitative samples. Compared with nucleic acid samples, non-nucleic acid samples always required large numbers of animals for sample preparation. The pass rate of homogeneity verification of the sample was 100%, but there is no quantitative study at present. Stability verification can be conducted for the short-term storage, transportation, freeze-thaw, different concentrations and terminal stability of different samples, but it lacks long-term stability verification.ConclusionIn the past ten years, the proficiency testing samples have basically met the basic requirements of the proficiency testing plan of laboratory animal testing area. However, the provision of nucleic acid samples should be reasonably increased; quantitative research samples and projects should be added; the influence of proficiency testing in the field of laboratory animal testing should be continuously expanded and maintain high-quality services.

ObjectiveTo assess the ability of the inspection institutions of laboratory animals and to improve their detection levels through the implementation of the proficiency testing for the detection ability of Bordetella bronchiseptica, Pasteurella pneumotropica, Pasteurella multocida and Corynebacterium kutscheri in laboratory animals.MethodsAccording to the proficiency testing scheme approved by China National Accreditation Service for Conformity Assessment (CNAS), the two projects, NIFDC-PT-309 and NIFDC-PT-363 in 2021 and 2022 were set up, respectively. The samples of respiratory tract pathogens in laboratory animal were prepared by freeze-drying at low temperature. The samples were randomly grouped and numbered through the CNAS platform and issued to participating units after passing the homogeneity and stability tests. A work instruction was attached with each sample set. The inspection reports and original records were required to be submitted within the specified time. If the feedback results were consistent with the sample setting, the results were deemed as satisfactory, and if the feedback results were inconsistent or not submitted on time, the results were deemed as unsatisfactory.ResultsThere were 29 laboratories joined to NIFDC-PT-309, 27 laboratories (93.1%) achieved satisfactory test results. There were 30 laboratories joined to NIFDC-PT-363, the satisfaction rate is 100%.ConclusionLaboratory animal quality inspection institutions have the ability to detect respiratory pathogens, which gets steadily improved through the ability verification activities.

ObjectiveTo test the single nucleotide polymorphism (SNP) detection technology level of mice genetic quality control in laboratories, promote the application of nucleic acid samples and SNP detection technology by implementing a nationwide proficiency testing program.MethodsIn 2022, National Institutes for Food and Drug Control implemented domestic laboratory proficiency testing named“NIFDC-PT-365 Laboratory Proficiency Testing Program of SNP Marker Detection Ablity in Laboratory Mice DNA Samples”. According to the requirements of the program, each participating laboratory received 2 blind samples of DNA from BALB/c mice and C57BL/6 mice with random numbers, standard operation procedure and 2 pairs of primers for rs3023177 and rs3023382 SNP loci. The participants should submit the results and original records before the deadline. If the mutation site amplification and sequencing results were consistent with the preseted results, it was considered as satisfactory; otherwise, it was considered as unsatisfactory. Meanwhile, PCR amplification system, sequencing direction and alignment methods of participating laboratories were compared to analyze as the relevant factors which may affect the SNP detection results of nucleic acid standard samples.ResultsThe results of 10 participating laboratories were consistent with the preset. They all obtained satisfactory results. Differences in amplification volume (20 μL, 25 μL, 30 μL), Taq enzyme system (single component and PCR MIX), sequencing company, sequencing direction and alignment software had no influence on the accuracy of the results.ConclusionThe participating laboratories have high detection ability in SNP detection method of mice nucleic acid samples. SNP detection technology is simple and feasible in mouse genetic quality control and is worth popularizing.

ObjectiveTo analyze the genetic quality of an outbred stock of Wistar rats in different periods by T/CALAS 21—2017 method, and to evaluate the applicability of the association standard.MethodsWistar rats selected from the same outbred stock in 2015 and 2019 were named as group A and group B, respectively. Twenty-five pairs of microsatellite primers were used to get genetic parameters for quality analysis of the outbred stock Wistar rats according to the association standard T/CALAS 21—2017. Polymorphism information content (PIC) was used to analyze the polymorphism of the loci.ResultsOne hundred alleles in group A and 69 alleles in group B were obtained. The average heterozygosity of the two groups was 0.574 and 0.447, while the average PIC was 0.541 and 0.393, respectively.ConclusionThe association standard T/CALAS 21—2017 has good applicability in the genetic quality analysis of outbred stock rats, and the genetic diversity of group A is better than that of group B.

ObjectiveTo promote the development of domestic laboratory animal detection proficiency testing activities and testing levels by assessing self testing capabilities.MethodsThe National Institutes for Food and Drug Control participated in the International Council for Laboratory Animal Science (ICLAS) laboratory animal detection proficiency test (i.e. Performance Evaluation Program for Diagnostic Laboratories, PEP) in 2013—2020. PEP pathogenic bacteria samples were tested by culture, biochemical identification, PCR, and sequencing. Finally, the results were analyzed and summarized.ResultsThere were 40 samples in seven international PEP activities, involving 27 kinds of pathogenic bacteria. The coincidence rates of the seven results were 8/8, 2/2, 5/8, 9/9, 5/5, 6/7, and 1/1, respectively, including 33 pathogen detection items and 7 antibody detection items. Nine pathogens were missing from the national standards: Bordetella hinzii, Serratia marcescens, Corynebacterium mastitis, CAR (Cilia-associated respiratory) bacillus, Actinobacillus muris, Citrobacter freundii, Klebsiella oxytoca, Streptococcus agalactiae, and pathogenic Escherichia coli.ConclusionThrough participating in PEP, we can identify the differences between domestic and foreign standards in laboratory animal detection and the testing capabilities that we need to improve, expecting to be in line with international standards as soon as possible.

ObjectiveTo study the changes of total IgG antibodies and neutralizing antibodies in KM, BALB/c, NIH, C57BL/6J and BALB/c-nu mice infected with murine norovirus (MNV).MethodsFive strains of KM, BALB/c, NIH, C57BL/6J and BALB/c-nu mice were collected. Each strain was divided into control group and infection group. The infection group was inoculated with MNV by gavage, and the control group was inoculated with the same volume of normal saline. Blood and serum were collected from 1 to 9 weeks after infection, total IgG antibody level (mean absorbance) was determined by ELISA, and neutralizing antibody titer (half protective dose, PD50) was determined by RAW264.7 cytopathic method. T-test was used to compare the mean absorbance difference between the infection group and the control group at each time point; One way ANOVA was used to compare the total antibody and neutralizing antibody titer of each strain of the infection group, and to analyze the antibody changes of each strain of mice.ResultsT-test showed that the mean absorbance of total IgG antibody in KM, BALB/c, NIH, C57BL/6J and BALB/c-nu mice infected on the day 21, 35, 14, 14 and 28 was significantly higher than that of the control group (P < 0.01). To compared the total IgG antibody level of each strain of mice, the results showed NIH＞C57BL/6J＞KM＞BALB/c＞BALB/c-nu. The total IgG antibody levels of BALB/c and BALB/c-nu mice were significantly lower than that of NIH and C57BL/6J mice (P < 0.05); The antibody level of NIH, C57BL/6J, KM and BALB/c mice increased rapidly at the 6th week of infection, and NIH mice entered the plateau stage at the 49th day. The IgG antibody of the other three strains was still on the rise during the experimental period, and the overall antibody level of BALB-/c-nu mice was the lowest. The overall neutralizing antibody titers of each strain of mice were compared, showing that C57BL/6J>NIH>KM>BALB/c>BALB/c-nu, and the mean values of lgPD50 were 3.881, 3.819, 3.746, 3.146, and 2.338, respectively. Analysis of variance showed that neutralizing antibody titer of BALB/c and BALB/c-nu mice was significantly lower than that of C57BL/6J, NIH and KM mice (P < 0.01). Except for BALB/c-nu mice, the neutralizing antibody of the other four strains increased rapidly at the second week of infection, and the neutralizing antibody of BALB/c-nu mice was always at a low level.ConclusionThe humoral immune response of NIH, C57BL/6J and KM mice infected with MNV is stronger, and that of BALB/c and BALB/c-nu mice is weaker.

Background: The genetic quality of laboratory mice may have a direct impact on the results of research. Therefore, it is essential to improve genetic monitoring methods to guarantee research quality. However, few current methods boast high efficiency, high throughput, low cost, and general applicability at the same time. Methods: First, we got 34 SNP loci from previous studies for inbred strains and screened out 15 loci with good polymorphism for outbred groups from these 34 loci. Then, by using the Luminex xTAG assay, we tested inbred strains and outbred groups. Results: We tested commonly used inbred strains and five DNA samples from the International Council for Laboratory Animal Science, obtaining correct genotyping results. Additionally, some loci were potentially confirmed to be useful for distinguishing C57BL/6 and BALB/c mouse substrains. Furthermore, we tested three outbred groups and analyzed the genetic structure, and we compared the results of the SNP markers by xTAG assay to the STR markers by PCR, the trends of the three groups are the same. Conclusions: In our studies, the panels could meet the requirements for method promotion and provide a good choice for the genetic monitoring of inbred and outbred mice. [ABSTRACT FROM AUTHOR]

Background: The genetic background of Toll-like receptor 4 (TLR4) proved to be important in the induction of immune protection against Bordetella pertussis infection in humans. Currently, the evaluation of the acellular pertussis (aP) vaccine depends largely on using different mouse strains, while the TLR4 genotype of different mouse strains in response to pertussis toxin (PT) is not carefully determined. The current study was designed to determine the differences in TLR4 genotype and TLR4 pathway-related cytokines in response to PT stimulation among mouse strains of ICR, NIH, and BALB/c. Method: We first determined the single-nucleotide polymorphisms (SNPs) in the TLR4 gene by using first-generation sequencing. Then, the cellular response, including the TLR4 mRNA expression and TLR4 signaling-related cytokines, of immune cells from different mouse strains after PT stimulation was determined. Result: Three missense mutation sites (rs13489092, rs13489093, rs13489097) of the TLR4 gene were found. ICR mice were homozygous without mutation, NIH mice were partially heterozygous, and BALB/c mice were homozygous with a missense mutation. The expression of TLR4 was repressed while the downstream cytokines were upregulated after PT stimulation differently among mouse strains. The IFN-β cytokine of the TRIF pathway was significantly increased in ICR mice (p < 0.05). The IL-6 cytokine of the MyD88-dependent pathway was significantly increased in BALB/c mice (p < 0.05). The identified SNPs of the TLR4 gene in different mouse strains might account for the differences in cytokines levels determined after PT stimulation. Conclusions: Our studies might provide useful referees to reduce the mouse-derived difference in the determination of vaccine titer and increase the comparability of the vaccine from different origins, as different mouse strains were used for vaccine development in different countries. [ABSTRACT FROM AUTHOR]