Guinea pig as a commonly used laboratory animal is widely used in various fields of biomedical research.The stability of genetic quality directly affects its development and application .Genetic testing is designed to confirm the genetic characteristics of each strain , to verify whether there are genetic mutations and other genetic contamination, to ensure that the test object meets the requirements of this strain .Along with the emerge of biochemical and molecular marker technology , a more convenient and reliable means is provided for research of genetic homozygosity , genetic type detection and genetic quality monitoring of guinea pigs .In this paper, the application and research progress of biochemical, cytological and molecular markers in studies of guinea pig diversity will be summarized , and provide some help for genetic testing guinea pig.

To monitor and analyze Cricetulus migratorius fungal diversity ,60 adult Cricetulus migratorius brought from Xinjiang region of China were dissected after being euthanized and the specimens were collected .Fungal diversity research was carried out by TaqMan MGB probe real-time fluorescence quantitative PCR ,the ribosome cloning sequencing ,and fungal cul-ture identification techniques .The 60 fungal isolates were characterized from Cricetulus migratorius ,including Candida albi-cans ,Trichosporonasahii ,Aspergillus fumigatus ,Aspergillusniger ,Aspergillussydowii ,Aspergillus japonicus ,Asper-gillus ustus ,Aspergillus versicolor ,Penicillium chrysogenum ,Paecilomyces variotii ,Penicillium aurantiogriseum ,Neuros-pora sitophila ,Neurospora intermedia ,and Cladosporium cladosporioides .Many of them associated with grey hamster as zoonotic pathogens .The results showed that the most dominant fungal group was Aspergillus ,and Penicillium followed by it . These fungi were susceptible to nystatin ,clotrimazole and voriconazole .It’s indicated that application of TaqMan MGB probe real-time fluorescence quantitative PCR ,the ribosome cloning sequencing ,and fungal culture identification techniques could ef-fectively analyze Cricetulus migratorius .The results could provide a scientific basis for Cricetulus migratorius microbiological monitoring and quality standard establishment in China .Overall ,the findings of the present study constitute ,to the authors’ knowledge ,the first extensive report on the diversity of fungal flora associated with Cricetulus migratorius .

Objective Giardia lamblia is an important pathogen of zoonosis giardiasis , it poses a potential threat to the quality of SPF (specific pathogen-free) laboratory animals cannot be ignored.The aim of this study is to establish the method of rapid diagnosis of Giardia lamblia, and analyze the test results of 516 batches form 17 manufactures.Methods Direct microscopy, Giemsa-fast staining and multiplex polymerase chain reaction (multiplex PCR) were applied to detect Giardia lamblia.Results Numerous of Giardia lamblia trophozoites and cysts were detected in SPF laboratory animals by using direct microscopy and Giemsa-fast staining, and multiplex PCR were performed to identify 18S rDNA,β-giardin, TPI and GDH genes of DNA extracted from these trophozoites and cysts identified Giardia lamblia.Direct microscopy, Giemsa-fast staining, and multiplex PCR methods can be used to detect Giardia lamblia.Of the 2562 SPF laboratory animals studied, 22.9%(586/2562) were positive for Giardia lamblia.Conclusions Direct microscopy , Giemsa-fast staining , and multiplex PCR were effective techniques with high sensitivity and specificity for rapid diagnosis of Giardia lambliain.It is not satisfactory that the results of Giardia lamblia examination in 516 batches form 17 manufactures failed to meet the requirements 100%.

Objective To establish an effective PCR assay for leptospirosis detection , and applicate the assay in tree shrew, mongolian gerbil and gray hamster .Methods Sequence of leptospira was obtained from the NCBI Genbank , and primers were designed based on the sequences .The positive amplified fragments were sequenced to verify the reliability of the method.The samples from tree shrew, mongolian gerbils and hamsters were tested using this PCR method .Results The PCR method for detection of leptospirosis was successfully established .The positive rate of Leptospira was 8.33% in 60 samples of conventional tree shrews , 100% in 104 samples of the conventional Mongolian gerbils , and 0% in 60 samples of clean gray hamsters.Conclusions The establishment of this PCR assay is useful in the detection of leptospirosis in tree shrew, mongolian gerbil and gray hamster .The results of our investigation of leptospira infection levels of the three new experimental animals may promote their application in biomedical research .

Objective To screen the markers of chromosome 12, which is vacant in the Beijing local standard DB11/T828.3-2011 for genetic quality control of miniature pigs in Beijing, and to study the applicability of the local standard by comparison of two different methods for evaluation of the genetic quality of the same population.Methods According to the literature, we selected four pairs of microsatellite markers of chromosome 12 and studied the polymorphism through monitoring the genetic quality of two populations of China Agricultural University miniature pigs.We screened the highly polymorphic markers of chromosome 12, combined them with the microsatellite primers of the standard 18 pairs of chromosomes to establish the whole chromosome method.We compared and analyzed the applicability of the local standard DB11/T828.3-2011 through monitoring the genetic quality of the same population of miniature pigs with different method.The data were processed and analyzed using software Popgen32.Results All the screened four pairs of microsatellite markers of chromosome 12 were highly polymorphic (PIC>0.5).The local standard showed a chromosome coverage of 94.7%, stability of amplification of 96.0%, and certified that the China Agricultural University miniature pig III was qualified, while the China Agricultural University miniature pig I was not qualified.When the markers of chromosome 12 were added, the whole chromosome method showed result of 100% chromosome coverage and 96.6% amplification stability, both of the two populations of pigs were certified as qualified.Conclusions The four screened markers of chromosome 12 are all highly polymorphic, and provide a support for supplement of the local standard DB11/T828.3-2011.

Objective To analyze the population genetics of two outbred colony guinea pigs from two institutions and provide basical information in developing genetic detection methods and standardization of the outbred guinea pigs.Methods 25 polymorphic microsatellite markers were screened by a fluorescent based semi-automated genotyping method for the two populations of guinea pigs, and the population genetic parameters were calculated.Results A total of 121 alleles were detected in the two populations, with 2-10 alleles and a mean 4.84 alleles at each locus.The mean expected heterozygosity was 0.6067, and the average polymorphism information content was 0.552.In the two populations, 103 and 116 alleles were detected, the mean expected heterozygosity was 0.5195 and 0.5838, and mean polymorphism information content was 0.459 and 0.518, respectively.Five loci and six loci, respectively, showed significant deviation from Hardy-Weinberg equilibrium (P<0.05) in the two populations, mostly resulted from heterozygote deficiency.The average Fst of all loci was 0.1056, which implied a moderate genetic differentiation between populations.The Nei’ (1972) genetic distance and Nei ‘(1978) unbiased genetic distance between the two populations were 0.3302 and 0.3204, respectively.Conclusions Both the two populations are consistent with a closed group of animal population genetic characteristics.Several loci deviate from HWE, which probably indicates that a certain degree of inbreeding phenomenon exists during the breeding process.

Objective To test the potency of hepatitis B vaccine in China. Methods Two inbred strains(DBA/1 and BALB/c) and two NIH closes-colonies of mice were typed in the H-2 region by microcy-totoxicity method and PCR. Groups of mice of the tested strains were immunized with the same hepatitis B vaccine, the titre of anti-HBsAg antibody was analyzed by microplate, and the ED50 was then estimated by Karder method for each strain. Results Significant differences were found between potency estimates de-rived from assays using different strains of mice. Conclusion It is likely that the variation of immune re-sponse to hepatitis B vaccine in mice is correlative with the H-2 haplotype. In some special case, the bet-erozygosity in H-2 region found in NIH stock could influence the accuracy in such testing even a reference preparation of hepatitis B vaccine was used. Base on our experiment, to select an appropriate NIH stocks with the H-2q haplotype for potency testing of hepatitis B vaccine in China.

To investigate the prevalence and the antibiotic resistance of bacteria isolated from 25 miniature pigs. 45 bacterial strains were isolated, which were identified by biochemical assays, amplification of 16S rRNA genes by PCR and sequence analysis, and were evaluated for resistance to 30 antibiotics. The identification results showed that these bacteria belonged to Campylobacter (Campylobacter jejuni), Helicobacterium (Helicobacter pylori), Klebsiella (Klebsiella pneumoniae), Escherichia (Escherichia coli, Escherichia fergusonii), Pseudomonas (Pseudomonas aeruginosa), Stenotrophomonas (Stenotrophomonas maltophilia), Staphylococcus (Staphylococcus aureus, Staphylococcus haemolyticus, Staphylococcus simulans), Streptococcus (Streptococcus pneumoniae, Streptococcus suis, Streptococcus vestibularis, Streptococcus mitis, Gemella measles, Aerococcus viridans) and Bacillus (Bacillus subtilis, Bacillus licheniformis, Bacillus alvei, Bacterium megaterium). These bacteria were all susceptible to aztreonam and cephalothin. However, the resistence to furazolidone was found. Microbial population carried by miniature pigs in China had characters of diversity. Results of this study provided scientifical accordance for the microorganism monitoring of miniature pigs in China.

Objective To develop a TaqMan MGB probe-based,sensitive and specific real-time fluorescence quantitative PCR assay for rapid detection of Helicobacter hepaticus.Methods Primers and probes specific toflaB gene of Helicobacter hepaticus were designed.A TaqMan MGB probe-based,real-time fluorescence quantitative PCR was established.The specificity,sensitivity and stability of the assay were assassed.Then,the established TaqMan MGB probe real-time fluorescence quantitative PCR assay was applied to detect Helicobacter hepaticus in 1081 clinical specimens during 2008-2011,compared with bacterial isolation and culture method and conventional PCR assay.Results The specificity of this established TaqMan MGB probe-based real-time fluorescence quantitative PCR was high and there were no cross-reactivity with Helicobacter pylori,Campylobacter jejuni,Clostridium piliforme,Pasteurella pneumotropica,Escherichia coli,Pseudomonas aeruginosa.The detection limits was 8.3 copies.The correlation coefficient and slope value of standard curve were 0.999 and -3.227,respectively and the efficiency of TaqMan MGB-based probe realtime fluorescence quantitative PCR assay was 100％.The TaqMan MGB-based probe real-time fluorescence quantitative PCR and conventional PCR were preformed to detect Helicobacter hepaticus in 1081 clinical specimens,a total of 86 specimens were positive for Helicobacter hepaticus.However,there was only 4 specimens were positive by bacteria isolation and culture method.The results showed that TaqMan MGB -based probe real-time fluorescence quantitative PCR for Helicobacter hepaticas was more sensitive than bacteria isolation and culture method,and it could detect Helicobacter hepaticus DNA from clinical specimens directly,and detection time is only 2 hours.Conclusion The TaqMan MGB-based probe real-time fluorescence quantitative PCR assay was a reliable,specific,sensitive and useful tool for rapid detection of Helicobacter hepaticus.

Objective To improve the accuracy of detection through analyzing the phenotypes of P.pneumotropica isolates in laboratory animals in Beijing area .Methods 306 suspicious P.pneumotropica strains were identified by biochemical identification and 16S rDNA sequencing.Then, to obtain the phylogenetic relationships combined with colony characteristics on blood agar plates and biochemical characteristics of 53 biotypes .Results BD Phoenix 100 automated bacterial identification system and 16S rDNA sequencing identified P.pneumotropica positive rate of 306 isolates were 164/306 and 227/306, respectively.There were 140 phenotypes in 227 true-positive strains, of which 106 were biotype Heyl and 23 were biotype Jawetz .Conclusions In the samples of laboratory animals in Beijing area , P.pneumotropica infection mainly are of biotype Heyl , and less is of biotype Jawetz .The phenotypes are diverse and widely distributed .