M13-IN5重组噬菌体对沙眼衣原体的抑制机制  

Mechanism of Inhibitory Effects of Recombinant M13-IN5 Phage on Chlamydia Trachomatis

作　　者：王江懿 尤聪[1] 赵乐然[1] 王梅 魏世娟[1] 练婷婷 邵丽丽[1] 刘全忠[1] WANG Jiangyi;YOU Cong;ZHAO Leran;WANG Mei;WEI Shijuan;LIAN Tingting;SHAO Lili;LIU Quanzhong(Department of Dermatology and Venereology,Tianjin Medical University General Hospital,Tianjin 300052,China;Department of Dermatology and Venereology,Tianjin First Central Hospital,School of Medicine,Nankai University,Tianjin 300192,China)

机构地区：[1]天津医科大学总医院皮肤性病科,天津300052 [2]天津市第一中心医院皮肤性病科,南开大学医学院,天津300192

出　　处：《中国皮肤性病学杂志》2022年第8期877-883,共7页The Chinese Journal of Dermatovenereology

基　　金：国家自然科学基金(31500157,31870169)。

摘　　要：目的探索重组M13-IN5噬菌体抑制沙眼衣原体(Chlamydia trachomatis,Ct)生长作用的机制。方法通过噬菌斑法测定M13-IN5噬菌体的滴度;CCK-8法检测M13-IN5噬菌体和青霉素对HeLa细胞的毒性作用;将不同滴度的M13-IN5噬菌体与50 U/mL的青霉素作用于沙眼衣原体后,通过免疫荧光计数包涵体数量;将滴度为10^(7)pfu/mL M13-IN5噬菌体与50 U/mL的青霉素作用于沙眼衣原体后进行子代感染力实验;采用定量逆转录PCR检测两组干预后CT_046、CT_111、CT_395、CT_443、CT_666、CT_823基因的转录变化,未干预沙眼衣原体感染组为阳性对照。两样本均数间比较采用t检验,多组样本均数间比较采用单因素ANOVA方法,多个样本间两两比较采用Bonferroni校正。结果滴度≤10^(9)pfu/mL的M13-IN5噬菌体以及50 U/mL的青霉素对HeLa细胞无毒性作用,10^(7)pfu/mL M13-IN5噬菌体与50 U/mL青霉素干预后,48 h形成的包涵体数量无明显区别,抑制率分别为19.62%和21.09%(P>0.05);M13-IN5噬菌体组干预形成的子代包涵体数量较青霉素干预组减少20.26%(P<0.05)。CT_046、CT_443在两个干预组转录水平均下降,M13-IN5噬菌体组较青霉素组下降明显(P<0.05);M13-IN5噬菌体干预组CT_666转录水平下降(P<0.05),青霉素干预组无变化(P>0.05);青霉素干预组压力应答基因CT_111、CT_395、CT_823转录水平增高(P<0.05),M13-IN5噬菌体干预组以上基因转录水平无变化(P>0.05)。结论重组M13-IN5噬菌体对沙眼衣原体的抑制作用强于青霉素,CT_046、CT_443、CT_666基因是重组噬菌体抑制沙眼衣原体生长的作用靶点。Objective To investigate the mechanism of inhibitory effects of recombinant M13-IN5 phage on Chlamydia trachomatis(Ct).Methods The titer of the M13-IN5 phage was determined by plaque forming assay.Cell counting kit-8 was performed to evaluate the effect of M13-IN5 phage and penicillin on the viability of HeLa cells.After C.trachomatis were treated with different titers of M13-IN5 phages and 50 U/mL penicillin,immunofluorescence microscopy was conducted for inclusion counting.Titer of 10^(7)pfu/mL M13-IN5 phage and 50 U/mL penicillin were employed in the progeny infectivity assay.Transcriptional levels of CT_046,CT_111,CT_395,CT_443,CT_666,CT_823 were measured by quantitative reverse transcriptional PCR.Ct infection alone was set up as the control group.Student's t-tests were used to analyze differences between two groups.One-way ANOVA followed by Bonferroni's multiple comparison test was used to determine differences between multiple groups.Results The viability of HeLa cells were not affected by M13-IN5 phage with titers≤10^(9)pfu/mL or 50 U/mL penicillin.The titer of 10^(7)pfu/mL M13-IN5 phage produced a comparable inhibitory rate of C.trachomatis as 50 U/mL penicillin,the inhibitory rates were 19.62%and 21.09%(P>0.05)at 48 hours post infection respectively;the progeny inclusion numbers in the M13-IN5 group decreased by 20.26%compared with the penicillin one(P<0.05).Compared with the control group,the transcriptional levels of CT_046,CT_443 decreased in both groups with a more significant reduction in the phage-treated group(P<0.05);the transcriptional level of CT_666 decreased in the phage-treated group(P<0.05),with no change in the penicillin group(P>0.05);the transcriptional levels of C.trachomatis pressure response genes such as CT_111,CT_395,CT_823 elevated in the penicillin group(P<0.05),while their transcriptional levels in the phage-treated group did not change(P>0.05).Conclusion The recombinant M13-IN5 phage had a better inhibitory effect on C.trachomatis growth than the penicillin.Recombinant M13-IN

关 键 词：沙眼衣原体 M13噬菌体 IN5 青霉素 

分 类 号：R374.1[医药卫生—病原生物学] R759[医药卫生—基础医学]