One single-center cross-sectional investigation on varicella antibody level of all age groups in Chinese people

ABSTRACT

Varicella outbreaks were mainly reported in developed regions with high vaccine coverage, but not in undeveloped areas. It is still not clear that whether the published data of varicella epidemiology could reflect the reality in China or not. In 2019, 657 subjects from People’s Hospital of Chongqing Youyang County were included. Anti-varicella-zoster virus (VZV) IgG antibodies were determined by ELISA. The anti-VZV IgG levels were categorized as positive when values were ≥100 mIU/mL. Our results showed that the rates of anti-VZV IgG seropositivity (χ² = 328.957, P < .0001) and geometric mean titers (P < .0001) were significantly influenced by age. The seropositivity declined dramatically from 84.5% in subjects ≤3 m of age, to 7.9% in subjects of >3 m-1 y (P < .0001). Then, the positivity rate increased slowly as age to 26.7% in >1-<3 y (P = .0006), and 34.5% in 3-<7 y (P = .4294). A steady rise (45.6%) in positivity was observed in subjects aged 7-<18 y. After then, the positivity began to increase robustly. A total of 87.8% of adults aged 18-<40 y had acquired VZV-specific immunity (P < .0001). The highest positivity rate was found in 40-<60 y  (98.3%) and ≥60 y (98.2%) group. In conclusion, most subjects of >3 m-<7 y age were susceptible to VZV. The proportion of subjects with natural infection-induced immunity increased with age. Nearly all subjects over 40 ages had positive anti-VZV IgG antibodies, which proved that they were infected by this virus in the past. These results suggested that VarV should be included in the national immunization program in China.

KEYWORDS:

• Varicella
• epidemiology
• Youyang
• anti-VZV IgG
• VarV

Introduction

Varicella, also known as chickenpox, is a highly communicable and vaccine-preventable respiratory disease mainly caused by the varicella-zoster virus (VZV).¹ It is usually a relatively a mild self-limiting disease in children after primary infection; however, it can cause morbidity, mortality, and cause severe complications including secondary skin and soft tissue infection, pneumonia, and central nervous system disease in adolescents, adults, and immunocompromised children.^2,3

Vaccination is the most successful and cost-effective intervention for the prevention of varicella. A live varicella attenuated live vaccine (VarV) has been available since 1974,⁴ and the vaccine was licensed for use in the United States in1995.⁵ As a consequence of the rise in vaccination rates, the disease was controlled better. Varicella immunization of children was started from 1998 in China, with single dose of varicella vaccination administrated at 12 months old.⁶ In China, vaccines are classified into two groups, CategoryⅠvaccines and CategoryⅡvaccines. CategoryⅠvaccines are vaccines which have been included in the national immunization, and it was mandatory and free to people. On the contrary, CategoryⅡvaccines were used in the private sector and people received the vaccine at their own expense, and they were not included in the national immunization. VarV is one of the CategoryⅡvaccines in China; therefore, the VarV coverage was lower than the CategoryⅠvaccines. Through the National Immunization Program (NIP) monitoring information system of China, the estimated coverage rate of VarV reached to 66.32% in 2014.⁷ However, the coverage of VarV showed a regional difference. Surveys showed that the number of immunized school-aged children in Beijing and Shanghai has exceeded 60% and 70%.⁸ However, the coverage of categoryⅡvaccine was relatively lower in the underdeveloped areas, such as Qinghai, Xinjiang, Sichuan, and Gansu.⁷ Meanwhile, the data on varicella epidemiology in these areas are limited.

Outbreaks of varicella are still seen in many developed countries, such as the USA, Germany, Australia, and Greece.⁹ Therefore, the second dose of VarV is recommended for preventing varicella in these countries. Recently, in China, outbreaks of varicella were also reported in developed cities and regions which had high coverage of VarV, such as Beijing,^10,11 Shanghai,¹² Jinan,¹³ and Shenzhen.¹⁴ The Beijing Center for Disease Control and Prevention (CDC) has introduced the routine two doses of inoculation procedure in 2012. After then, the strategy of two doses of varicella immunization was also recommended in Heilongjiang, Shandong, and Fujian province.⁸ However, varicella outbreaks are rarely reported in undeveloped areas which had low vaccination rates and low population density. It is still not clear that whether the published figures of varicella epidemiology could reflect the reality of varicella in China or not. Varicella was not a mandated reporting disease in China and the national varicella surveillance data through the China Information System for Disease Control and Prevention (CISDCP) – a web-based, are not available.¹⁵ Therefore, it is necessary to evaluate the population immunity and regional epidemiologic data, especially for remote regions or economically backward place.

Chongqing is a municipality in the southwestern mountainous area of China, with 28 million permanent residents. Youyang Tujia and Miao Autonomous County, Youyang County for short, located in the southeast of Chongqing, is a state-level poverty-stricken county. The coverage of VarV in the whole Chongqing remains unclear. A study conducted in Sangpingba district reported that the coverage of VarV in preschool children (0–6y) was 76.28%.¹⁶ However, the reported incidence of varicella in Chongqing increased from 85.9 per million in 2007 to 300.2 per million in 2013.¹⁷ We conducted a seroprevalence study in children, adolescents, and adults (aged 0–89 years old) and measured varicella immunity to assess the current immune status of the Youyang population and dynamics of varicella epidemiology, to find susceptible age groups.

Materials and method

Serum samples

In 2019, 657 subjects from People’s Hospital of Chongqing Youyang County were included in this study. Young children visited the hospital for medical examination, adults and olds visited the hospital for medical examination, antenatal care, fracture, or chronic diseases (such as hypertension, heart disease, diabetes). Those subjects with any suspicious infection were not involved in the study. Subjects were stratified into the following age groups: ≤3 m, >3 m-1 y, >1-<3 y, 3-<7 y, 7-<18 y, 18-<40 y, 40-<60 y, and ≥60 y. The demographic characteristics about the subject’s gender, age or date of birth, and residence area (urban or rural) were also collected to assess its possible effect on varicella immunity. All serum samples were frozen at −20°C until analysis.

This study was reviewed and approved by the Ethics Committee of Youyang County People’s Hospital. Written informed consent was obtained from all participants or parents for the blood to be used for research on infectious diseases.

Serological testing

Anti-VZV IgG antibodies were determined by using a commercially available ELISA kit (Institut Virion/Serion GmbH, Germany), according to the manufacturer’s instructions. The antibody results were expressed in mIU/mL. According to the kit instructions, the serum antibody concentration ranged from 15 to 2000 mIU/mL, the samples with an antibody concentration >2000 mIU/mL were labeled as 2000 mIU/mL, and samples <15 mIU/ml were labeled as 7.5 mIU/mL. The anti-VZV IgG levels were categorized as negative, equivocal, and positive when values obtained were <50, 50–<100 and ≥100 mIU/mL, respectively. The anti-VZV IgG positive means past VZV infection, successful vaccination, or immunoglobulin recipient.

Data analysis

Serum anti-VZV IgG levels were expressed as geometric mean titers (GMT). The concentrations of anti-VZV IgG in different age groups were examined using one-way analysis of variance (ANOVA). The seroprevalence among different groups was compared with the chi-square test. Data were analyzed using the JMP (version 10.0) and SPSS (version 17.0). P ≤ 0.05 was considered statistically significant.

Results

Characteristics of study population

A total of 657 subjects were enrolled in the study. There were 379 males and 278 females, the median average age was 7 years old (range: 0–89 y). Three hundred eighty-four subjects lived in rural areas. There were 71 (10.8%), 89 (13.5%), 75 (11.4%), 87 (13.2%), 68 (10.4%), 74 (11.3%), 116 (17.7%) and 77 (11.7%) samples in the groups of ≤3 m, >3 m-1 y, >1-<3 y, 3-<7 y, 7-<18 y, 18-<40 y, 40-<60 y, and ≥60 y, respectively.

Seroprevalence of anti-VZV IgG

The proportions of negative, equivocal, and positive anti-VZV IgG among the total study subjects were 33.3%, 5.3%, and 61.3%, respectively. Compared with males (56.7%), the positivity rates of anti-VZV IgG were significantly higher in females (67.6%) (P = .0046). No significant difference was found between positivity rates and residence areas (Table 1).

Table 1. The distribution of anti-VZV IgG in subjects aged 0–89 y from Youyang, Chongqing

Group	N	Anti-VZV distribution, N, Prevalence% (95% CI)
		Negative (<50 mIU/ml)	equivocal (50- < 100 mIU/ml)	Positive (≥100 mIU/ml)
Gender
Female	278	83, 29.9% (24.8%-35.5%)	7, 2.5% (1.2%-5.1%)	188, 67.6% (61.9%-72.9%)
Male	379	136, 35.9% (31.2%-40.8%)	28, 7.3% (5.2%-10.5%)	215, 56.7% (51.7%-61.6%)
χ^2 P		χ^2= 2.622, P = .1054	χ^2= 7.541, P = .0060	χ^2= 8.031, P = .0046
Residence
Rural	384	125, 32.6% (28.1%-37.4%)	18, 4.7% (3.0%-7.3%)	241, 62.8% (57.8%-67.4%)
Urban	273	94, 34.4% (29.0%-40.2%)	17, 6.2% (3.9%-9.7%)	162, 59.3% (53.4%-65.0%)
χ^2 P		χ^2= 0.254, P = .6144	χ^2= 0.750, P = .3865	χ^2= 0.787, P = .3750
Age
≤ 3 m	71	11, 15.5% (8.9%-25.7%)	0, 0 (0–5.1%)	60, 84.5% (74.3%-91.1%)
>3 m- 1 y	89	73, 82.0% (72.8%-88.6%)	9, 10.1% (5.4%-18.1%)	7, 7.9% (3.9%-15.4%)
>1- <3 y	75	44, 58.7% (47.4%-69.1%)	11, 14.7% (8.4%-24.4%)	20, 26.7% (18.0%-37.6%)
3- <7 y	87	46, 52.9% (42.5%-63.0%)	11, 12.6% (7.2%-21.2%)	30, 34.5% (25.3%-44.9%)
7-<18 y	68	36, 52.9% (41.2%-64.3%)	1, 1.5% (0.3%-7.9%)	31, 45.6% (34.3%-57.3%)
18-<40 y	74	8, 10.8% (5.6%-19.9%)	1, 1.4% (0.2%-7.3%)	65, 87.8% (78.5%-93.5%)
40-<60 y	116	1, 0.9% (0.2%-4.7%)	1, 0.9% (0.2%-4.7%)	114, 98.3% (93.9%-99.5%)
≥60 y	77	0, 0 (0–4.8%)	1, 1.3% (0.2% −7.0%)	76, 98.2% (93.0%-99.8%)
χ^2 P		χ^2= 263.917, P < .0001	χ^2= 41.629, P < .0001	χ^2= 328.957, P < .0001
Total	657	219, 33.3% (29.8%-37.0%)	35, 5.3% (3.9%-7.3%)	403, 61.3% (57.6%-65.0%)

The seropositivity of anti-VZV IgG was also analyzed by different age groups. The rates of anti-VZV IgG seropositivity were significantly influenced by age (P < .0001). The seropositivity declined dramatically from 84.5% in subjects ≤3 m of age, to 7.9% in subjects of >3 m-1 y (P < .0001). Then, the positivity rate increased slowly as age to 26.7% in >1-<3 y, and 34.5% in 3-<7 y. A steady rise in positivity was observed in school children and teenagers, leaving about half of the subjects of 7-<18 y age susceptible to varicella (P = .1602). After then, the positivity began to increase robustly. A total of 87.8% of adults aged 18-<40 y had acquired VZV-specific immunity (P < .0001). Almost all subjects over 40 ages had anti-VZV IgG antibodies (Table 1).

Titers of anti-VZV IgG

The anti-VZV IgG of subjects from Youyang ranged from 7.5 to 2855.2 mIU/mL. The GMT of anti-VZV IgG was 125.2 mIU/mL. Table 2 shows the anti-VZV titers according to gender, residence, and age. Compared with males, the GMT of anti-VZV IgG levels was significantly higher in females (151.8 versus 108.6 mIU/mL, P = .0312). There was no significant difference in IgG level between subjects who lived in rural areas and those who lived in urban areas (P = .1805).

Table 2. The GMT of anti-VZV IgG in subjects aged 0–89 y from Youyang, Chongqing

Group	N	Geometric mean titers (mIU/ml)
Gender
Female	278	151.8
Male	379	108.6
P	-	0.0312
Residence
Rural	384	134.1
Urban	273	113.6
P	-	0.1805
Age
≤ 3 m	71	235.2
>3 m- 1 y	89	16.2
>1- <3 y	75	34.0
3- <7 y	87	52.8
7-<18 y	68	60.2
18-<40 y	74	311.1
40-<60 y	116	530.8
≥60 y	77	629.7
P	-	<0.0001
Total	657	125.2

The overall GMT increased with age in parallel with the increase in seropositivity. The GMT decreased from 235.2 mIU/mL in subjects ≤3 m of age, to 16.2 IU/ml (P < .0001) in subjects of >3 m-1y. Then, the GMT increased slightly as age to 34.0 mIU/mL in >1-<3 y, 52.8 mIU/mL in 3-<7 y, 60.2 mIU/mL in 7-<18 y and 311.1 mIU/mL in 18-<40 y. The highest GMT was found in 40-<60 y (530.8 mIU/mL) and ≥60 y (629.7 mIU/mL) group (Table 2).

Discussion

In recent years, varicella outbreaks continue to occur in China. Although the national varicella epidemiologic data are not available, the data from the Immunization Information System and the Infectious Disease Reporting System in Beijing, Shanghai, and Jiangsu showed an increased trend of varicella cases. A total of 23,497 cases in Haidian district of Beijing were reported during 2007–2015.¹⁸ In the Fengtai District of Beijing, the varicella breakthrough rate increased from 0.7% in 2008 to 2.5% in 2012.¹⁹ A total of 13511 varicella cases in the Minhang District of Shanghai were reported during 2008–2014.¹² Therefore, it is important to investigate the population immunity to varicella, especially for the undeveloped region. The low immunization coverage of varicella in these areas may lead to an increased susceptible population and the possibility of varicella outbreaks.

The total positivity rate of anti-VZV IgG (61.3%) in Youyang was similar to the rates of Hangzhou (65.59%),⁸ Shenzhen (61.49%),¹⁴ and Taiwan (64.1%).²⁰ It is lower than the rates of Beijing (84.5%)²¹ and Harbin (71.49%).²² It is higher than that in another previous study in Jiangsu,²³ which reported that the positivity rate of anti-VZV IgG in subjects in children aged 1–9 years was 43.5%. The significant regional differences in positivity rate of anti-VZV IgG may be related to the different immunization coverage of VarV, local incidence of varicella, and different sample survey objects and periods.

The fact that higher seropositivity was detected in newborns ≤3 m of age (84.5%) than in the >3 m-1 y age group (7.9%), indicating that the maternal anti-VZV immunity wanes with time. The varicella vaccination was administrated at 12 months old in China. A previous study showed that most children received the first dose of varicella vaccination before 3 years old. Much to our surprise, the seropositivity rates in >1-<3 y group were only 26.7% in this study. The kids went to kindergarten at 3 years old, and most of them enter primary school at around 7 years old, who could receive a second dose of varicella vaccine. Meanwhile, preschool children who were not vaccinated at a younger age had an opportunity for vaccination. The seropositivity rates in preschool children (3-<7 y) were 34.5%. There is no good way to discriminate antibodies induced by natural infection or vaccination. Equivocal (22/35) antibody levels were found mostly in subjects of >1-<3 y and 3-<7 y age. Therefore, we speculated that the levels in these two groups may reflect the weaker immune status in the vaccinated population. However, the coverage rate of voluntarily self-paid VarV was actually low in Youyang region, even though the antibodies in these two groups came completely from vaccination.

The vaccination rate of VarV among primary school students is relatively lower than that of preschool children, while the positivity rate increased to 45.6% in the following age groups (7-<18 y). In addition, the lower equivocal rate was detected in this group. These results indicated that most of them received varicella immunity by natural infection or booster immunity due to VZV spreading. After then, the positivity began to increase robustly in the following non-immunized adults. Nearly all subjects over 40 ages had positive anti-VZV IgG antibodies. These results proved that the anti-VZV IgG in adults was induced by natural infection or booster immunity due to VZV spreading. Moreover, the higher positivity rate in adults and lower equivocal rate showed that acquired natural immunity is long-lasting.

The significant regional differences of the positivity rate in young children were correlated with inequalities of immunization coverage. The seropositivity rates in >1-4 y group were only 25.4% (95% CI: 18.5%-33.8%) in this study. It is lower than that in a previous study in Korea,²⁴ which reported that the positivity rate of anti-VZV IgG in children aged 1–4 y was 67.3%. A recent study performed by Zhang et al.²³ reported that coverage of VarV in children aged 1–4 y was 47.1% in Jiangsu. Moreover, a previous study in Sangpingba district of Chongqing has reported that the positivity rate of that the coverage of VarV in preschool children (0–6y) was 76.28%.¹⁶ Therefore, even in the same city, the immunization coverage of VarV could show significant regional differences. Up to 2009, 19 countries introduced VarV for national routine immunization for children or selected vaccination for high-risk groups. The World Health Organization (WHO) recommends routine childhood immunization against varicella to be considered in countries where varicella is a relatively important public health and socioeconomic problem.²⁵ To control the spread and outbreaks of varicella, VarV should be included in the routine immunization program in China. It is important to eliminate regional differences in immunization coverage of VarV and local incidence of varicella.

Several studies in developed cities and regions had indicated that high single-dose varicella vaccination coverage is effective in reducing varicella incidence, but not sufficient to prevent outbreak.^26-29 However, while outbreaks of varicella were reported in developed cities and regions, they were rarely reported in the undeveloped areas with low vaccination rates. The reason for this finding may be complicated. The immunity to varicella in children from developed cities and regions was mainly acquired from vaccination; thus, the weaker immune status occurs. On the contrary, considering the seropositivity changes in our study, we speculated that a portion of children (7-<18 y) from undeveloped or rural regions acquired immunity to varicella by natural infection. Therefore, about half of the children in primary school after seven from undeveloped or rural regions got immunity to varicella, and it was long-lasting and more effective to prevent varicella outbreaks. Another possibility is that outbreaks are not being notified. It may be also explained by the mild manifestations and low hospital-visiting rates in undeveloped or rural regions. Lots of children (especially for mild manifestations) in these areas received treatment in a private clinic, thus resulting in the occurrence of varicella could not be detected by CDC.

This study found that the positivity rates and GMT of anti-VZV IgG were significantly influenced by gender. Compared with male, the positivity rates (χ² = 8.031, P = .0046) and GMT (P = .0312) of anti-VZV IgG were significantly higher in female. It is consistent with two previous investigations in Jiangsu (47.3% vs 40.1%) and Hangzhou (72.4% vs 63.79%), which revealed that seroprevalence is significantly higher in girls than in boys. The reason for this finding is unclear. It may include the different vaccination coverage, incidence of varicella, contact rate, exposure levels, and asymptomatic infection rate of these children.

There were certain limitations in this study. First, the history of vaccination was not collected, it cannot be elucidated whether the antibodies of the study subjects are due to infection or immunization. The situation of varicella immunization of children was speculated by age groups; thus, the findings in this study can only suggest causation, but cannot prove it. This vaccination information has important implications, since the WHO warns that populations with childhood vaccination coverage of 20–80% could change the age pattern of presentation of chickenpox toward adult ages in which there is an increased risk of severe chickenpox and associated mortality.³⁰ The seropositivity rates in >1-<3 y and 3-<7 y group in our study were maintained in this range, we must be alert to the possibility. The VarV coverage rate of healthy children in Youyang should be collected to analyze. Second, it was a hospital-based study of the entire population. The extensive inference capacity could be obtained from the outcome based on community populations. Meanwhile, the samples from more remote regions, economically backward places, and the whole country were required. Multi-center studies should be performed in the future to give evidence for the national improvement of varicella immunization strategies. Third, the collected demographic characteristics were limited in our study, there are a number of other factors that can influence anti-VZV IgG seropositivity, such as immigration status, education level, household income, and number of children. Fourth, the recruitment of these subjects was voluntary, it would potentially induce selective bias. We will use purposive sampling methods to select provinces, counties (districts), and streets, and obtain the sample based on the Immunization Information Management System (IIMS) in further study.

In conclusion, this study summarizes the varicella immunity in subjects aged 0–89 years from Youyang, Chongqing. Most of the subjects of >3 m-<7 y age were susceptible to VZV, the positivity rate increased steadily as age to 45.6% in the following age groups (7-<18 y). Nearly all subjects over 40 ages had positive anti-VZV IgG antibodies. The coverage rate of voluntarily self-paid VarV may be low, and varicella virus widely circulated in Youyang region. To prevent the varicella outbreaks, VarV should be included in the national immunization program in China.

Disclosure of potential conflicts of interest

No potential conflicts of interest were disclosed.

Funding

This work was supported by the Medical Research Project of Chongqing Health and Family Planning Commission (2016ZDXM041).