Prevalence And Mutations Analysis Of Hepatitis B With Coexistence Of Hbs Ag And HBsAb

BackgroundHepatitis B virus（HBV）is one of the most common chronic viral infections in the world.It is estimated that about 2 billion people have been infected,and more than 300 million people are chronic carriers of the virus.The appearance of the HBsAb indicates that the patient has been vaccinated anainst HBV,or has been infected with HBV,and is now in the recovery period of infection and has certain immunity.However,there were concurrent HBsAg and anti-HBs in chronic hepatitis B（CHB）carriers that had been reported in previous studies.The rate of simultaneous HBsAg and anti-HBs serological profile ranged from 2.43%to 8.90%in different patient groups^[1-7].Several reports showed that HBV mutations which might be attributed to the selection of immune escape mutations,could explain the coexistence of HBsAg and HBsAb in HBV infected patients^[4,6,8-11].However,some researchers rejected this hypothesis and instead suggested that the enigma of concurrent HBsAg and anti-HBs was related to the weak binding of anti-HBs to HBsAg^[12].So the mechanism underlying the coexistence of HBsAg and anti-HBs remains unclear and controversial.ObjectivesTo determine the prevalence and serological characteristics of the coexistence of HBsAg and HBsAb in Guangzhou between 2014-2016,and the influencing factors of the difference prevalence of concurrent HBsAg and anti-HBs were analyzed.Meanwhile,to assess the relationship between the main hydrophilic regional mutations of the S gene and the coexistence of HBsAg and HBsAb serological data in Chinese chronic HBV patients,and explore the possible mechanism and clinical significance of its occurrence.It is of great importance for the diagnosis,treatment and evaluation of chronic hepatitis B.Method1.All the HBV serological markers（HBsAg、HBsAb、HBeAg、HBeAb、HBcAb）were detected by chemiluminescent microparticle immunoassay（CMIA）technique with architect i2000 automatic analyzer（Illinois,USA）.The source of the specimen was chronic HBV infection patients treated at the First Affiliated Hospital of Guangzhou Medical University between 2014and 2016.The prevalence of the coexistence of HBsAg and HBsAb in Guangzhou area was obtained by using SPSS 22.0.2.The serum viral DNA of 132 patients with group I（44 cases）HBsAg（+）/HBsAb（+）and group II（88 cases）HBsAg（+）/HBsAb（-）in 2015 was amplified by Nested PCR,and the S gene sequences of hepatitis B virus was obtained by direct sequencing technology.The results of S gene mutation were analyzed by sequence comparison software MEGA6 and statistical software SPSS 22.0.Results1.A total of 13,369 patients with CHB were recruited from The First Affiliated Hospital of Guangzhou Medical University between 2014-2016 in this study,559 patients with the coexistence of HBsAg and HBsAb were discovered.The prevalence rates for the coexistence of HBsAg and HBsAb were 4.52%,4.02%and 4.04%,respectively,from 2014 to 2016.The proportion of male patients was higher than female,mainly with the mode dominated"HBsAg（+）/HBsAb（+）/HBeAg（-）/HBeAb（+）/HBcAb（+）".HBeAb positive patients was higher than HBeAg positive patients,accounting for 68.34%,and mainly with low concentrations of HBsAb 10.00¹00.00 mIU/mL,accounting for89.44%.There was statistically significant differences in age and gender（p<0.05）.While there is no significant difference with HBeAg positive by statistical analysis（p>0.05）.Among them,the average age in2015 was significantly higher than in 2014 and 2016,and the proportion of men in 2015 was significantly lower than in 2014 and 2016.2.In both groups,the HBV genotype mainly were B（56.82%）and C（43.18%）.There was no statistically significant difference in age,sex and HBV genotype distribution between the two groups（p>0.05）.However,the HbeAg positive rate of Group I patients was significantly higher than GroupⅡ,and the difference was statistically significant（p<0.001）.The Group I patients mainly with low concentrations of HBsAb 10.00^&lt;100.00 mIU/ml,accounting for 86.36%.The change of the major hydrophilic region（MHR）was59.09%.The number of residue changes per 100 residues within the MHR was7.1 times more frequent for group I than groupⅡ（2.50vs0.35,p<0.001）.At least two amino acid mutations occurred in 16 patients,of which 12patients（31.82%,12/44）came from Group I and only 2 patients（2.27%,2/88）came from Group II.Variation occurred mostly in the“a”determine region（3.41vs0.90,p<0.001）between groupⅠand groupⅡ.In addition,there were statistically significant differences in MHR1（2.81vs0.05,p<0.001）,MHR3（4.38vs1.06,p<0.001）,MHR4（2.05vs0.68,p<0.05）,and MHR5（1.24vs0,p<0.001）between the two groups.But there was no significant difference between the two groups at MHR2（2.27vs0.57,p=0.099）.There were also significant differences in point mutation prevalence between the two groups.The frequent escape variants in group I were located at positions s101Q,s114S,s120P,s126T/I,s129Q,s131T,s133M and s145G.While sQ101K,sT131N and sM133L were more frequently discovered in group I with significant difference（p<0.05）.Conclusions1.The prevalence with the coexistence of HBsAg and HBsAb in Guangzhou was 4.18%.The HBeAb positive patients were more likely to be present in the concurrent HBsAg and anti-HBs,however,the conversion of HBeAg to HBeAbhasnotdiminisheditsinfectivity.Simultaneously,low concentrations of HBsAb were more frequently discovered,It may indicate that HBsAb in patients with the coexistence of HBsAg and HBsAb was formed during the infection.2.In both groups,the HbeAg positive rate was higher in Group I,that means it was more contagious than groupⅡ.However,the low level of ALT suggests that the effect of both positive patients on the body is the result of long-term effects.The S gene mutation was more frequent in group I,this suggests that mutations in S region genes,especially in the"a"determine region,may caused the occurrence of the the coexistence of HBsAg and anti-HBs.HBsAg variants mainly located at the first loop of the"a"determine region（MHR3）,It may be that the HBV mutation are not affected by artificial active immunity and/or artificial passive immunity and/or treated with nucleoside analogues,but largely due to the long-term screening pressure of HBsAb on hepatitis B virus formed during the infection that causes HBV to choose immune escape mutations,and eventually led to the emergence of the coexistence of HBsAg and HBsAb.