Molecular epidemiological analysis of Influenza viruses in Influenza-like illness cases: a retrospective study in Chongqing Hi-Tech Zone, China (2021-2024)

doi:10.21203/rs.3.rs-5003876/v1

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Molecular epidemiological analysis of Influenza viruses in Influenza-like illness cases: a retrospective study in Chongqing Hi-Tech Zone, China (2021-2024)

https://doi.org/10.21203/rs.3.rs-5003876/v1

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The seasonal epidemiology and high variability of influenza viruses result in yearly differentiation of main pandemic strains. Effective prevention and control of influenza epidemics require enhanced surveillance, timely updates of vaccine components.Therefore, it is crucial to establish a continuous surveillance system to track epidemiological trends and develop appropriate vaccination and preventive measures.A retrospective analysis was conducted on the molecular epidemiological characteristics of influenza viruses in influenza-like cases at a hospital in Chongqing Hi-Tech Zone from 2021 to 2024. Among 39,986 ILI specimens tested using the immunocolloid gold method, 6,616 influenza viruses were detected, with a detection rate of 16.54%. Infections included 4,464 influenza A viruses (67.50%), 2,033 influenza B viruses (30.73%), and 119 co-infections of influenza A and B viruses (1.77%). From November 2021 to September 2022, the peak detection of influenza A virus occurred in January 2022, and influenza B virus peaked in April 2022. From July 2022 to March 2023, the influenza A virus peaked in December 2022, while the influenza B virus peaked in January 2023. Influenza B virus detection lagged behind influenza A virus detection. In this region, H3N2 was the predominant subtype of influenza A, and Victoria was the main subtype of influenza B. The "H1" subtype of influenza A did not appear throughout 2023 but accounted for 82% in January 2024 and 16% in February 2024. The "Yamagata" subtype of influenza B did not appear from 2023 to 2024. Influenza epidemics in the winter and spring seasons in Chongqing Hi-Tech Zone were predominantly caused by influenza A, with influenza B also circulating. Influenza A strains were mainly H3N2, while influenza B strains were primarily Victoria. This study provides essential data and a scientific basis for developing and optimizing influenza prevention and control strategies.

Molecular epidemiological analysis

Influenza viruses

Influenza-like illness cases

prevention and control

Influenza is a pandemic respiratory pathogen from the Orthomyxoviridae family, which can infect humans or other animals^[1]. Due to its frequent genetic variation, the antigenicity of pandemic strains often changes annually, posing challenges for vaccine preparation and influenza surveillance. Influenza viruses are classified into four types: A, B, C, and D, based on antigenic differences in their nucleocapsid and nucleoprotein antigens.^[2]. Among these, influenza A subtypes (H1N1 and H3N2) and influenza B are known as seasonal influenza because they can circulate among populations throughout the year^[3]. Influenza epidemics are often associated with significant economic and social burdens^[4], making networked surveillance a crucial tool for preventing and controlling pandemics^[5]. The World Health Organization analyzes laboratory surveillance data from global influenza surveillance networks annually and recommends vaccine strains for the subsequent year's vaccine preparation^[6–10].

Serum amyloid A (SAA) is an inflammation marker that has garnered significant attention recent^{ly [11–14]}. SAA levels can rapidly and markedly increase following infection, inflammation, or tissue injury, making it valuable for the early diagnosis of infectious diseases, risk assessment, evaluation of therapeutic efficacy, and prognosis determination. The elevation of SAA due to viral infections suggests its potential role in diagnosing influenza A and B^[15–19].

This study aims to explore the molecular epidemiological characteristics of influenza viruses in influenza-like cases in Chongqing Hi-Tech Zone from 2021 to 2024. By monitoring and analyzing these cases, the research assesses the potential of inflammatory markers, such as SAA and C-reactive protein (CRP), for use in influenza diagnosis. The findings provide a scientific basis for improving influenza prevention and control strategies in the region and across the country, ultimately helping to mitigate the impact of influenza on public health.

This study conducted a retrospective analysis of influenza-like illness (ILI) cases in a hospital in Chongqing Hi-Tech Zone, China, from 2021 to 2024. Inclusion criteria were based on the National Influenza Surveillance Programme (2017 version), requiring cases to meet the following conditions: body temperature ≥ 38°C with symptoms of acute respiratory infection such as cough, sore throat, or sputum. Some cases may exhibit mild symptoms or no fever; physical examinations typically revealed enlarged tonsils and pharyngeal congestion. All cases required signed informed consent and were reviewed and approved by the hospital ethics committee.

We screened cases from the hospital's electronic medical record system that met the ILI definition. Exclusion criteria included non-acute respiratory infectious diseases, known non-influenza viral respiratory diseases, and cases lacking necessary clinical and laboratory data. Additionally, basic patient information such as age, gender, occupation, and place of residence was collected to facilitate subsequent data analysis.

Detection methods

Pharyngeal swabs and sputum samples were collected using sterile swabs. These samples were promptly sent to the laboratory for testing immediately after collection. The colloidal gold method was used to detect viral antibodies, and the fluorescent PCR method was employed to detect nucleic acids. Gene sequencing was utilized to identify different subtypes.

Statistical processing

All data were analyzed utilizing SPSS version 22.0 statistical software. The research results were organized to ensure the logic and completeness of data screening. Descriptive statistics, such as mean ± standard deviation (x‾±s), were used for continuous data, while frequency and percentage were used for categorical data.

Comparisons between groups for count data were performed utilizing the χ² test or Fisher's exact probability method. Comparisons between groups for continuous data were performed utilizing the t-test or ANOVA. A two-sided P < 0.05 was considered statistically significant.

Multivariate logistic regression analysis was used to adjust for potential confounders among the independent variables. The correlation between inflammatory indicators, such as SAA and CRP, and influenza virus infection was analyzed to evaluate the sensitivity and specificity of these indicators for influenza. Further statistical analysis was conducted to provide a more accurate assessment of these relationships.

Influenza virus test results

A total of 39,986 ILI specimens were tested using the immunocolloid gold method, detecting 6,616 influenza viruses, resulting in a detection rate of 16.54%. Among these, 4,464 (67.50%) were influenza A viruses, 2,033 (30.73%) were influenza B viruses, and 119 (1.77%) were simultaneous detections of both influenza A and B viruses, as shown in Table 1.

From November 2021 to September 2022, the peak detection of the influenza A virus occurred in January 2022, while the peak detection of the influenza B virus occurred in April 2022. From July 2022 to March 2023, the peak detection of influenza A virus occurred in December 2022, and the peak detection of influenza B virus occurred in January 2023. Notably, the peak of influenza B virus detection lagged behind that of influenza A (Fig. 1).

Table 1

Influenza virus test results of ILI specimens
times	n	Influenza A virus (n = 4464)	Stream B virus (n = 2033)	Stream A + Stream B virus (n = 119)
November 2021	1	0	0	0
December 2021	0	0	0	0
January 2022	4365	1054	23	16
February 2022	3185	43	13	8
March 2022	3769	478	217	11
April 2022	3352	213	245	17
May 2022	2677	208	236	26
June 2022	2356	64	41	5
July 2022	782	53	12	1
August 2022	753	31	9	0
September 2022	1056	132	27	6
October 2022	1369	106	13	4
November 2022	3745	187	23	3
December 2022	6287	1236	541	14
January 2023	5428	647	625	6
February 2023	618	8	5	1
March 2023	243	4	3	1

H1N1 and H2N2 type test results

From April 2023 to April 2024, H3N2 was the predominant subtype of influenza A, while Victoria was the main subtype of influenza B. The H1 subtype of influenza A was absent throughout 2023 but accounted for 82% of detections in January 2024 and 16% in February 2024. The Yamagata subtype of influenza B was undetected from 2023 to 2024 (Table 2 and Fig. 2).

Table 2

Influenza virus test results of ILI specimens
times	influenza A virus		influenza A virus (e.g., virus that affects the immune system)
times	H1(%)	H3 (%)	Yamagata (%)	Victoria (%)
April 2023	0	100	0	100
May 2023	0	100	0	100
June 2023	0	0	0	0
July 2023	0	0	0	0
August 2023	0	0	0	0
September 2023	0	100	0	0
October 2023	0	100	0	100
November 2023	0	100	0	100
December 2023	0	100	0	100
January 2024	82	18	0	100
February 2024	16	84	0	100
March 2024	87	13	0	100
April 2024	87	13	0	100

Data analysis of SAA and CRP levels in patients with influenza A and B

From April 2023 to April 2024, our hospital recorded 241 influenza A and 207 cases of influenza B. The differences in SAA and CRP levels between these groups were statistically significant (p = 0.000 and p = 0.001, respectively). SAA and CRP levels were significantly higher in the influenza A group compared to the influenza B group. The increase in SAA levels in the influenza A group was greater than in the influenza B group and higher than in healthy adults. These findings indicate a correlation between influenza positivity and elevated SAA levels (Table 3 and Fig. 3).

Table 3

Comparison of SAA and CRP levels [M (P25, P75), mg/L] in patients with influenza A and B from April 2023 to April 2024
groups	n	SAA	CRP
Influenza A group	241	86.5 (43.6, 128.5)^ab	18.7 (9.2, 33.7)^ab
ethereal group (physics)	207	25.6 (15.7, 75.6)^a	7.9 (2.4, 15.3)^a
P		0.000	0.001

Comparison of the distribution of influenza viruses by gender in various age groups

The differences in influenza A virus detection rates among genders aged 0–10 years and 60–70 years were statistically significant. In the 0–10 age group, the detection rate for males was 17.34% compared to 14.35% for females (P < 0.05). In the 60–70 age group, the detection rate for males was 3.52% compared to 6.13% for females (P < 0.05). For influenza B virus, the detection rate in the 0–10 years age group was 7.34% for males compared to 6.13% for females, also statistically significant (P < 0.05) (Table 4).

Table 4

Comparison of the distribution of influenza viruses by age group and sex
item	Age group	Number of influenza A virus detections (n)	Influenza A virus χ²	P	Number of detections of Stream B virus (n)	Stream B virus χ²	P
0 to 10 years	18752	3275	30.627	< 0.001	1307	5.249	0.036
male	10865	1856			738
women	7887	1419			569
> 10 ~ 20 years old	3875	545	0.387	0.413	325	0.062	0.629
male	1946	327			182
women	1929	218			143
> 20 ~ 30 years old	2637	236	0.253	0.721	206	0.136	0.658
male	1214	93			89
women	1423	143			117
> 30 ~ 40 years old	3276	258	0.003	0.899	125	0.385	0.472
male	1424	115			53
women	1852	143			72
> 40 ~ 50 years old	1321	57	0.005	0.953	23	0.394	0.437
male	589	23			8
women	732	34			15
> 50 ~ 60 years old	1629	123	0.936	0.547	13	0.649	0.352
male	757	42			4
women	872	81			9
> 60 ~ 70 years old	1753	106	9.978	0.002	21	1.525	0.136
male	697	49			12
women	1056	57			9
> 70 years	1928	47	0.659	0.346	11	0.178	0.459
male	745	21			9
women	1183	26			2

Comparison of the distribution of influenza viruses by sex

Among the 32,930 ILI cases, 15,610 were male and 17,320 were female. The detection rates of influenza A and B viruses were higher in males (12.01% and 4.89%, respectively) compared to females (7.83% and 3.69%, respectively). These differences were statistically significant (P < 0.05)(Table 5).

Table 5

Comparison of the distribution of influenza viruses by sex
distinguishing between the sexes	n	influenza A virus		influenza A virus (e.g. virus that affects the immune system)
distinguishing between the sexes	n	Number of detections (n)	Detection rate (%)	Number of detections (n)	Detection rate (%)
male	15610	1874	12.01	763	4.89
women	17320	1356	7.83	639	3.69
χ²		35.623		13.261
P		< 0.001		< 0.001

Colloidal gold and fluorescent PCR results

A random selection of 300 cases underwent fluorescent PCR detection for influenza virus nucleic acid, resulting in 73 influenza virus cases, yielding a detection rate of 24.33%. Among these, 52 cases were influenza A virus (71.23%), and 21 cases were influenza B (28.77%). Using the immunocolloidal gold method to detect influenza virus antigens, 49 cases were detected, with a detection rate of 16.34%. The detection rate of influenza virus using the fluorescent PCR method was significantly higher than that of the immunocolloidal gold method (χ² = 5.736, P = 0.004), as shown in Table 6.

Table 6

Results of colloidal gold and fluorescent PCR assays
immunocolloid gold method	Fluorescent PCR method		add up the total
immunocolloid gold method	positive	negatives	add up the total
masculine	42	7	49
negatives	31	220	251
add up the total	73	227	300

Surveillance of influenza samples in Chongqing High-Tech Zone revealed the molecular epidemiological characteristics of influenza virus strains, including detection rates and temporal variations of major genotypes. These findings contribute to understanding influenza virus transmission patterns and trends, support the iterative updating of influenza virus strains, and help predict transmission and pathogenicity, thus providing a scientific basis for public health decision-making^[20–23].

Chongqing Hi-tech Zone has a strong population mobility, which increases the likelihood of influenza generation, prevalence, and dissemination. Therefore, monitoring the pathogenetic and epidemiological patterns of influenza in this area is particularly significant. This study analyzed all ILIs reported at Chongqing Hi-Tech Zone University Hospital from November 2021 to April 2024. Both influenza A and B peaked during the winter and spring seasons, with influenza B peaking later than influenza A. This finding is consistent with previous literature reports^[24][25][26]. It can be concluded that influenza A predominantly dominates the epidemic season.

From April 2023 to April 2024, there were 241 positive cases of influenza A and 207 positive cases of influenza B. Among the influenza A cases, H3N2 was the predominant subtype, while Victoria was the main subtype for influenza B. The H1 subtype of influenza A was absent throughout 2023 but accounted for 82% of the cases in January 2024 and 16% in February 2024. The Yamagata subtype of influenza B did not appear from 2023 to 2024^[27]. The study found that children aged 0–10 years were affected by influenza. There was a statistically significant difference in the detection rates of influenza A and B viruses between males and females in this age group (p < 0.05)^[28–30].

The SAA and CRP levels of 241 influenza A positive cases and 207 influenza B positive cases were analyzed, showing statistically significant differences between the two groups (p = 0.000 for SAA and p = 0.001 for CRP). The CRP levels were 7.9 mg/L in the influenza A positive group and 18.7 mg/L in the influenza B positive group. The SAA levels in the influenza A positive group were significantly higher than in the influenza B positive group. The CRP level in the influenza A positive group was significantly higher than in the influenza B positive group. A correlation was found between influenza A and B positivity and elevated SAA levels, with influenza A positivity showing a stronger correlation with elevated SAA^[31–33].

The correlation between influenza A and B positivity and SAA elevation was notably strong. In summary, winter and spring influenza in Chongqing's High-Tech Zone is predominantly caused by influenza A, though influenza B is also prevalent. Among the positive cases, H3N2 is the primary subtype for influenza A, while Victoria isthe primary subtype for influenza B. Elevated SAA levels are correlated with both influenza A and B positivity.

Although the study demonstrates the epidemiological characteristics of influenza virus infection and the potential application of inflammatory markers, several limitations exist. The limited sample size may restrict the generalizability of the findings. Additionally, the study did not analyze the effect of different influenza A virus strains on the levels of 14 inflammatory markers or explore the potential influence of other respiratory pathogens on these markers.

Future studies will explore the mechanisms behind these epidemiological features to obtain more comprehensive and objective findings. Additionally, further research is needed on the expression patterns of inflammatory markers in different populations, such as children, the elderly, and patients with chronic diseases, and their role in the preventive mechanisms of influenza vaccination. As research deepens, a clearer understanding of the function and importance of the influenza vaccine and inflammatory markers will provide a solid theoretical basis for influenza prevention and treatment.

The winter and spring influenza virus epidemics in the High-Tech Zone of Chongqing Municipality, China, were dominated by influenza A, with influenza B circulating. The predominant strains were H3N2 for influenza A and Victoria for influenza B. This study provides important data and a scientific basis for developing and optimizing influenza prevention and control strategies in the region.

Acknowledgements

Not applicable.

Funding

This study is supported by the Scientific and Technological Research Program of Chongqing Municipal Education Commission (KJQN202000443 );the China Postdoctoral Science Foundation (2023MD734129).

Author Contributions

W.C. and Y. H.D. Conceived the manuscript. W.C. and Y. H.D. wrote the draft manuscript. P.X. curated the data and prepared the figures and tables. J.J.X. critically read and edited the manuscript. P.X. curated the data and prepared the figures and tables. J.J.X. critically read and edited the manuscript.

Data availability

No datasets were generated or analysed during the current study.

Ethics approval and consent to participate

The study was approved by the Ethics Committee of the University Town Hospital of Chongqing Medical University(LL-202339).

Consent for publication

All authors have read and agreed to the published version of the manuscript.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work.

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You are reading this latest preprint version

Molecular epidemiological analysis of Influenza viruses in Influenza-like illness cases: a retrospective study in Chongqing Hi-Tech Zone, China (2021-2024)

Status:

Version 1

Abstract

Figures

Introduction

Material and Methods

Detection methods

Statistical processing

Results

Influenza virus test results

H1N1 and H2N2 type test results

Data analysis of SAA and CRP levels in patients with influenza A and B

Comparison of the distribution of influenza viruses by gender in various age groups

Comparison of the distribution of influenza viruses by sex

Colloidal gold and fluorescent PCR results

Discussion

Conclusion

Declarations

References

Additional Declarations

Status:

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