AIM: To evaluate the level of sperm chromosome aberrations in male patients with hepatitis B, and to directly detect whether there are HBV DNA integrations in sperm chromosomes of hepatitis B patients.METHODS: Sperm c...AIM: To evaluate the level of sperm chromosome aberrations in male patients with hepatitis B, and to directly detect whether there are HBV DNA integrations in sperm chromosomes of hepatitis B patients.METHODS: Sperm chromosomes of 14 tested subjects (5healthy controls, 9 patients with HBV infection, including 1with acute hepatitis B, 2 with chronic active hepatitis B, 4with chronic persistent hepatitis B, 2 chronic HBsAg carriers with no clinical symptoms) were prepared using interspecific in vitro fertilization between zona-free golden hamster ova and human spermatozoa, and the frequencies of aberration spermatozoa were compared between subjects of HBV infection and controls. Fluorescence in situ hybridization (FISH) to sperm chromosome spreads was carried out with biotin-labeled full length HBV DNA probe to detect the specific HBV DNA sequences in the sperm chromosomes.RESULTS: The total frequency of sperm chromosome aberrations in HBV infection group (14.8%, 33/223) was significantly higher than that in the control group (4.3%,5/116). Moreover, the sperm chromosomes in HBV infection patients commonly presented stickiness, clumping, failure to staining, etc, which would affect the analysis of sperm chromosomes. Specific fluorescent signal spots for HBV DNA were seen in sperm chromosomes of one patient with chronic persistent hepatitis. In 9 (9/42) sperm chromosome complements containing fluorescent signal spots, one presented 5 obvious FISH spots, others presented 2 to 4signals. There was significant difference of fluorescence intensity among the signal spots. The distribution of signal sites among chromosomes was random.CONCLUSION: HBV infection can bring about mutagenic effects on sperm chromosomes. Integrations of viral DNA into sperm chromosomes which are multisites and nonspecific, can further increase the instability of sperm chromosomes. This study suggested that HBV infection can create extensively hereditary effects by alteration genetic constituent and/or induction chromosome aberrations, as well as t展开更多
A pair of primers created from information of PmNOBⅢ genome DNA Sal I fragment produced a 355bp band by using Penaeus chinensis non occluded baculovirus (PcNOBV),the WSBV isolate from P.hinensis in China's Ma...A pair of primers created from information of PmNOBⅢ genome DNA Sal I fragment produced a 355bp band by using Penaeus chinensis non occluded baculovirus (PcNOBV),the WSBV isolate from P.hinensis in China's Mainland,as the DNA template.The specific PCR product was cloned,sequenced and labeled with digoxigenin (DIG)DNA labeling kit(Boehringer Mannheim).The DIG labeled fragment was tested by dot blot hybridization for sensitivity and specificity with purified PcNOBV nucleocapsid,PcNOBV infected shrimp tissues and healthy shrimp tissues.The detection limit of the DNA probe is 6.8pg of purified PcNOBV DNA.No hybridization signals were observed using DNA from healthy shrimp as template.Healthy P.chinensis,artificially infected P.chinensis and pond reared adult P.chinensis were screened for PcNOBV infection by both PCR and the hybridization assay.The results showed a good relationship between PCR and the hybridization assay.These findings demonstrate that the DIG labeled probe can be used as a sensitive,specific and cost effective reagent for detection of PcNOBV.展开更多
基金the Natural Science Foundation of Guangdong Province,No.940567the National Natural Science Foundation of China,No.39970374
文摘AIM: To evaluate the level of sperm chromosome aberrations in male patients with hepatitis B, and to directly detect whether there are HBV DNA integrations in sperm chromosomes of hepatitis B patients.METHODS: Sperm chromosomes of 14 tested subjects (5healthy controls, 9 patients with HBV infection, including 1with acute hepatitis B, 2 with chronic active hepatitis B, 4with chronic persistent hepatitis B, 2 chronic HBsAg carriers with no clinical symptoms) were prepared using interspecific in vitro fertilization between zona-free golden hamster ova and human spermatozoa, and the frequencies of aberration spermatozoa were compared between subjects of HBV infection and controls. Fluorescence in situ hybridization (FISH) to sperm chromosome spreads was carried out with biotin-labeled full length HBV DNA probe to detect the specific HBV DNA sequences in the sperm chromosomes.RESULTS: The total frequency of sperm chromosome aberrations in HBV infection group (14.8%, 33/223) was significantly higher than that in the control group (4.3%,5/116). Moreover, the sperm chromosomes in HBV infection patients commonly presented stickiness, clumping, failure to staining, etc, which would affect the analysis of sperm chromosomes. Specific fluorescent signal spots for HBV DNA were seen in sperm chromosomes of one patient with chronic persistent hepatitis. In 9 (9/42) sperm chromosome complements containing fluorescent signal spots, one presented 5 obvious FISH spots, others presented 2 to 4signals. There was significant difference of fluorescence intensity among the signal spots. The distribution of signal sites among chromosomes was random.CONCLUSION: HBV infection can bring about mutagenic effects on sperm chromosomes. Integrations of viral DNA into sperm chromosomes which are multisites and nonspecific, can further increase the instability of sperm chromosomes. This study suggested that HBV infection can create extensively hereditary effects by alteration genetic constituent and/or induction chromosome aberrations, as well as t
文摘A pair of primers created from information of PmNOBⅢ genome DNA Sal I fragment produced a 355bp band by using Penaeus chinensis non occluded baculovirus (PcNOBV),the WSBV isolate from P.hinensis in China's Mainland,as the DNA template.The specific PCR product was cloned,sequenced and labeled with digoxigenin (DIG)DNA labeling kit(Boehringer Mannheim).The DIG labeled fragment was tested by dot blot hybridization for sensitivity and specificity with purified PcNOBV nucleocapsid,PcNOBV infected shrimp tissues and healthy shrimp tissues.The detection limit of the DNA probe is 6.8pg of purified PcNOBV DNA.No hybridization signals were observed using DNA from healthy shrimp as template.Healthy P.chinensis,artificially infected P.chinensis and pond reared adult P.chinensis were screened for PcNOBV infection by both PCR and the hybridization assay.The results showed a good relationship between PCR and the hybridization assay.These findings demonstrate that the DIG labeled probe can be used as a sensitive,specific and cost effective reagent for detection of PcNOBV.
