Epidemiological characteristics of common respiratory pathogens in children

doi:10.21203/rs.3.rs-3812419/v1

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Epidemiological characteristics of common respiratory pathogens in children

https://doi.org/10.21203/rs.3.rs-3812419/v1

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Background:Children's respiratory tract infection is a common disease affecting children's health. Our aim is to describe the epidemiological characteristics of common pathogens of children's respiratory tract infection in outpatient clinics in central Shandong, China.

Methods: Sputum, tracheal aspirate, alveolar lavage fluid and other samples of 4804 children admitted to wards of Zibo Maternal and Child Health Hospital for treatment of respiratory tract infection from June 2019 to December 2022 were collected, and 12 common respiratory tract pathogens were detected by multiple PCR detection technology. Two bacteria (SP, HI), two atypical pathogens (MP, CP) and eight viruses (HRV, RSV, IVA, PIV, HMPV, HBOV, HCOV, IVB) were included. The proportion and positive detection rate of each type of respiratory tract mixed infection and the positive detection rate of single pathogen in different ages and seasons were statistically analyzed.

Results: (1) Among 4804 children with respiratory tract infection, the total positive rate was 77.87% (3741/4804). The positive rate was 34.47% (1656/4804). Streptococcus pneumoniae, rhinovirus and respiratory syncytial virus were the highest. There were 2085 cases of mixed infection with two or more pathogens, the positive rate was 43.40%. (2) The positive rates of infection in infant group (0-1 years old), infant group (1-3 years old), preschool group and school age group (3 years old -) were roughly the same. The infection rates of Streptococcus pneumoniae, respiratory syncytial virus and parainfluenza virus in infant group, rhinovirus in infant group, influenza A virus, chlamydia pneumoniae, mycoplasma pneumoniae and Haemophilus influenzae in school age group were higher than those in other groups. The difference was statistically significant (P<0.05). (3) The positive detection rates of spring, summer, autumn and winter groups were 43.58%, 38.64%, 33.73% and 29.27%, respectively. The positive rates of Streptococcus pneumoniae and Haemophilus influenzae in spring group, mycoplasma pneumoniae in summer group, rhinovirus, respiratory syncytial virus and influenza A virus in autumn group, chlamydia pneumoniae, Boca virus and influenza B virus in winter group were higher than those in other seasons, and the differences were statistically significant (P<0.05).

Conclusion：The pathogen detection rate of children varies with age and season, and the prevention and treatment of a certain respiratory pathogen infection must be combined with its raging season and age rule.

children respiratory tract infection

Multiple PCR detection of respiratory tract

Pathogens

Positive rate

Respiratory tract infection refers to infectious diseases caused by a variety of pathogenic microorganisms, including viruses, bacteria, fungi, mycoplasma, chlamydia, Rickettsia, parasites, etc^[1]. Respiratory multiplex PCR detection technology uses direct detection of pathogen nucleic acid sequences to detect the types of respiratory pathogens that children are infected with. These include two types of bacteria (Streptococcus pneumoniae (SP), Haemophilus influenzae (HI)), and two atypical pathogens (Mycoplasma pneumoniae, Mycoplasma pneumoniae). MP), Chlamydia pneumoniae (CP), and eight viruses (Human rhinovirus (HRV), Respiratory syncytial virus (Respiratory syncytial virus). RSV, Influenza A Virus (IVA), Parainfluenza Virus (PIV), Human metapneumovirus, HMPV, Human bocavirus (HBOV), Human coronavirus (HCOV), Influenza B Virus (IVB). The above different pathogens can cause the same clinical symptoms, and the same pathogen infects different children with different clinical symptoms, so it is particularly important to determine the type of respiratory pathogen infection. In this study, 4804 samples of children admitted to Zibo Maternal and Child Health Hospital for treatment of respiratory tract infections were collected from June 2019 to December 2022, and 12 common respiratory tract pathogens were detected using respiratory tract multiplex PCR detection technology to explore the application value of respiratory tract multiplex PCR detection technology in children with respiratory tract infections. The results are reported as follows:

1.Research object

A total of 4804 children who came to our hospital for treatment of respiratory tract infections from June 2019 to December 2022 were selected as the study objects for PCR detection to detect the infection of respiratory tract viruses, bacteria, mycoplasma, chlamydia and other pathogens. The detection of pathogens in different age groups was divided into infant group (0–1 years old, n = 1955), infant group (1–3 years old, n = 1215), preschool group and school age group (3 years old -, n = 1634). The detection of respiratory pathogens in different seasons was divided into spring group (n = 569), summer group (n = 885), autumn group (n = 1862) and winter group (n = 1488). Inclusion criteria: ① Children with respiratory tract infection were selected according to the 8th edition of Zhufutang Practical Pediatrics^[2] and Internal Medicine^[3]; ② Children with stable disease development; ③ With the consent of the guardian, children can actively participate in the treatment.

2.Multiple PCR detection of respiratory tract in children

Multiple PCR detection of respiratory tract was used to detect 12 common respiratory pathogens. It includes two bacteria (SP, HI), two atypical pathogens (MP, CP) and eight viruses (HRV, RSV, IVA, PIV, HMPV, HBOV, HCOV, IVB). The patient's tongue was pressed back with a tongue depressor, the throat swab was extended into the child's throat to stimulate the cough, and the coughed sputum was stuck on the swab. The assistant nurse sent the patient for examination within 30 minutes. The specimens were observed by naked eye: the color, viscosity, presence of blood and pus, etc., and the specimens were examined by smear. The squamous epithelial cells ≤ 10 and white blood cells &gt were found in the low-power field of view. 25 specimens are ideal. Test process: The collected specimens were mixed with 0.9% sodium chloride injection, centrifuged at 12000 r/min for 5 minutes, the supernatant was removed, and 50µl DNA extract was added to the precipitation and thoroughly mixed, and then bathed in boiling water at 100℃ for 10 minutes, centrifuged at 12000 r/min for 5 minutes. The DNA was extracted and then quantitative detection was performed directly by fluorescent PCR. Result interpretation: After the program is finished, the system will automatically give Ct value. When Ct ≥ 35, the result will be interpreted negatively. Ct < 35, the result read positive.

3.Statistical analysis

The data were processed by Graphpad prism software, χ² test was used for counting data, represented by n(%), P < 0.05, the difference of 0.05 was statistically significant.

1. Detection of other pathogens in children with respiratory tract infection

When 10246 children with respiratory tract infection were enrolled for bacterial isolation and culture, considering that a considerable part of them may be respiratory virus, mycoplasma or chlamydia infection, or the mixed infection of two pathogens or more than two pathogens, therefore, according to whether the lymphocyte is elevated and whether the body temperature of the children exceeds 38.5℃, Whether accompanied by poor mental state, pale face, muscle pain and other symptoms, 4804 of the children were tested by PCR for other respiratory pathogens, the detection range was common viruses, mycoplasma, chlamydia, etc., which caused respiratory infection. The test results are as follows: 3741 positive samples, the positive rate is 77.87%. The number of single infection was 1656, the positive rate was 34.47%. The higher detection rates were streptococcus pneumoniae, rhinovirus and respiratory syncytial virus, accounting for 20.17%, 4.28% and 2.31%, respectively. The results are shown in Table 1 and Fig. 1.

Table 1

Positive rate and proportion of single pathogen infection in respiratory tract
pathogen	Positive number	Positive rate(%)	Account for(%)
SP	969	20.17	58.51
HRV	205	4.28	12.28
RSV	111	2.31	6.70
IAV	101	2.10	6.10
CP	75	1.56	4.53
MP	59	1.23	3.56
HI	36	0.75	2.17
PIV	28	0.58	1.69
HPMV	16	0.33	0.97
HBOV	13	0.27	0.79
HCOV	10	0.21	0.60
Influenza B virus	6	0.12	0.36
total	1656	34.47	100

2. Analysis on detection of respiratory pathogens mixed infection in children

Common pathogens were detected in 4804 children with respiratory tract infection. The results showed that 2085 children had mixed infection, with a positive rate of 43.40%. There were 43 kinds of mixed infection combinations. The mixed infection combinations with high positive detection rate were streptococcus pneumoniae + rhinovirus, streptococcus pneumoniae + Haemophilus influenzae, streptococcus pneumoniae + respiratory syncytial virus, accounting for 12.90%, 10.84% and 8.30%, respectively. Other high combinations were Streptococcus pneumoniae + Chlamydia pneumoniae, Streptococcus pneumoniae + Mycoplasma pneumoniae, Streptococcus pneumoniae + rhinovirus + Haemophilus influenzae, etc. The other combinations were not statistically analyzed because they involved more pathogens and the number of cases was too small. The results are shown in Table 2.

Table 2

Analysis of mixed infection of respiratory pathogens in children
pathogen	Positive number	Account for(%)
SP + HRV	269	12.90
SP + HI	226	10.84
SP + RSV	173	8.30
SP + CP	141	6.76
SP + MP	121	5.80
SP + Influenza B virus	72	3.45
SP + HRV + HI	65	3.12
SP + HPMV	58	2.78
SP + HI + RSV	50	2.40
Other combinations	910	43.65
total	2085	100

3. Detection and analysis of respiratory single pathogen infection in children of different age groups

Common pathogens were detected in 4804 children with respiratory tract infection. The prevalence of respiratory tract single pathogens in children of different age groups was different. The positive rate of respiratory tract single pathogen detection in infant group (0–1 years old) was 35.40% (692/1955). The positive rate of respiratory single pathogen detection was 33.58% (408/1215) in the infant group (1–3 years old), and 34.03% (556/1634) in the preschool group and the school age group (3 years old -). Due to the small number of specimens in the school age group, the preschool group and the school age group were combined into one group for the convenience of analysis. The detection rates of Streptococcus pneumoniae, respiratory syncytial virus and parainfluenza virus in infant group were higher than those in infant group, preschool group and school age group, and the difference was statistically significant compared with other age groups (P < 0.05); The infection rate of rhinovirus in the infant group was higher than that in the infant group, preschool group and school age group, and the difference was statistically significant compared with other age groups (P < 0.05); The infection rates of influenza A virus, chlamydia pneumoniae, mycoplasma pneumoniae and Haemophilus influenzae in preschool and school age groups were higher than those in infant and infant groups, and the difference was statistically significant compared with other age groups (P < 0.05); There was no significant difference in the infection rates of metapneumovirus, Boca virus, coronavirus and influenza B virus (P > 0.05). The results are shown in Table 3 and Fig. 2.

Table 3

Comparative analysis of respiratory single pathogen infection in children of different age groups
pathogen	Infant group (n = 1955)		Toddler group(n = 1215)		Preschool group and school age group(n = 1634)		χ²	P
pathogen	Positive number	Positive rate(%)	Positive number	Positive rate(%)	Positive number	Positive rate(%)	χ²	P
SP	459	23.48	261	21.48	276	16.89	24.060	0.000*
HRV	82	4.19	77	6.34	46	2.82	21.210	0.000*
RSV	83	4.25	24	1.98	4	0.24	63.930	0.000*
IAV	7	0.36	20	1.65	74	4.53	76.870	0.000*
CP	7	0.36	5	0.41	63	3.86	84.830	0.000*
MP	2	0.10	2	0.16	55	3.37	93.320	0.000*
HI	7	0.36	5	0.41	24	1.47	17.620	0.000*
PIV	19	0.97	5	0.41	4	0.24	8.944	0.011
HPMV	9	0.46	4	0.33	2	0.12	3.281	0.194
HBOV	7	0.36	4	0.33	3	0.18	1.012	0.603
HCOV	7	0.36	1	0.08	2	0.12	3.620	0.164
Influenza B virus	3	0.15	0	0	3	0.18	2.099	0.350
total	692	35.40	408	33.58	556	34.02
Note: * There were significant differences among the pathogens in different seasons, P < 0.05.

4. Detection and analysis of respiratory single pathogen infection in children in different seasons

Common pathogens were detected in 4804 children with respiratory tract infection. The positive detection rate was 43.58% (248/569) in spring group and 38.64% (342/885) in summer group. The positive detection rate was 33.73% (628/1862) in autumn and 29.27% (446/1488) in winter. The detection rates of Streptococcus pneumoniae and Haemophilus influenzae in spring group were higher than those in other three seasons, and the difference was statistically significant (P < 0.05); The detection rate of mycoplasma pneumoniae in summer group was higher than that in other three seasons, and the difference was statistically significant (P < 0.05); The detection rates of rhinovirus, respiratory syncytial virus and influenza A virus in the autumn group were higher than those in the other three seasons, and the difference was statistically significant (P < 0.05); The detection rates of chlamydia pneumoniae, Boca virus and influenza B virus in winter group were higher than those in other three seasons, and the difference was statistically significant (P < 0.05); There was no significant difference in the positive rates of parainfluenza virus, metapneumovirus and coronavirus infection among different seasonal groups, and there was no statistical significance between different seasonal groups (P > 0.05). The results are shown in Table 4 and Fig. 3.

Table 4

Comparison of positive rates of respiratory single pathogen infection in children in different seasons
pathogen	Spring(n = 569)		Summer(n = 885)		Autumn(n = 1862)		Winter(n = 1488)		χ2	P
pathogen	Positive number	Positive rate(%)	Positive number	Positive rate(%)	Positive number	Positive rate(%)	Positive number	Positive rate(%)	χ2	P
SP	204	35.85	244	27.57	252	13.53	303	20.36	162.300	0.000*
HRV	10	1.76	34	3.84	148	7.95	12	0.81	115.100	0.000*
RSV	4	0.70	3	0.34	61	3.28	43	2.89	31.660	0.000*
IAV	0	0	22	2.49	78	4.19	1	0.07	82.190	0.000*
CP	9	1.58	0	0	36	1.93	32	2.15	18.540	0.000*
MP	6	1.05	19	2.15	29	1.56	5	0.34	17.730	0.000*
HI	9	1.58	5	0.56	6	0.32	17	1.14	30.030	0.000*
PIV	3	0.53	11	1.24	7	0.38	7	0.47	8.385	0.387
HPMV	2	0.35	1	0.11	5	0.27	4	0.27	0.949	0.814
HBOV	1	0.18	0	0	0	0	15	1.01	30.030	0.000*
HCOV	0	0	3	0.34	5	0.27	2	0.13	2.632	0.452
Influenza B virus	0	0	0	0	1	0.05	5	0.34	7.892	0.048
total	248	43.58	342	38.64	628	33.73	446	29.97
Note: * There were significant differences among the pathogens in different seasons, P < 0.05.

In this study, 4804 children with respiratory tract infection samples were tested for respiratory tract pathogens by PCR. The results showed that 1656 children were infected with a single pathogen, and the positive detection rate was 34.47%. The pathogens with high detection rate were streptococcus pneumoniae, rhinovirus and respiratory syncytial virus in order. There were 2085 cases of mixed infection with pathogens, the positive rate was 43.40%. The positive detection rate of single pathogen in Zibo area was 34.47%, slightly lower than the 38.26% reported by He Chenglu^[4] in Kunming area, and slightly higher than the 32% reported by Li Hongjun^[5] in Tongzhou area of Beijing, basically consistent. The top three pathogens of respiratory infection in Zibo area were Streptococcus pneumoniae, rhinovirus and respiratory syncytial virus. Chen Jing^[6] reported that among 368 community-acquired pneumonia samples with an average age of 4–5 years old, the pathogens with high detection rate were influenza virus, mycoplasma pneumoniae and respiratory syncytial virus, which was partially consistent with the results of this study. The distribution of respiratory pathogens is greatly affected by region, environment and season, and specific regions should be analyzed. This study analyzed the changes of several major respiratory pathogens with age. The results showed that the distribution of common respiratory pathogens in children was closely related to the age of children, and the positive detection rates of infection in infant group (0–1 years old), infant group (1–3 years old), preschool group and school age group (3 years old -) were roughly the same. The infection rates of Streptococcus pneumoniae, respiratory syncytial virus and parainfluenza virus in infant group, rhinovirus in infant group, influenza A virus, chlamydia pneumoniae, mycoplasma pneumoniae and Haemophilus influenzae in preschool group and school age group were higher than those in other age groups, and the difference was statistically significant (P < 0.05). The positive detection rate of Streptococcus pneumoniae was higher in the infant group, which was consistent with the report of Jin Shuyan^[7] that Streptococcus pneumoniae mostly occurred in infancy. The positive rates of respiratory syncytial virus and parainfluenza virus were higher in the infant group,with the increase of age, there is a clear downward trend, which was consistent with the report of Mao Bibo^[8], this may be because infants' respiratory mucosal barrier and local mucosal immunity have not been fully established and are more susceptible to virus attack.The positive detection rate of rhinovirus in young children is relatively high, which is basically consistent with the study of Li Yan^[9], and the positive rate also decreases with the age of children. The positive rates of influenza A virus, chlamydia pneumoniae and mycoplasma pneumoniae were high in preschool and school age groups, which was basically consistent with the research results of Han Lijuan^[10]. Children over 3 years old were generally concentrated in kindergartens and primary schools, providing a way for local respiratory infection outbreaks, so the above pathogens were mostly concentrated in this age group. The high positive rate of Haemophilus influenzae infection in the school-age group is consistent with the report of Gao Xue^[11] that the proportion of Haemophilus influenzae in children aged 1–14 is greater than that in children under 1 year old. The reason should also be related to the concentrated outbreak of infection in kindergartens and schools.

The distribution of respiratory pathogens in children is also closely related to seasons. This study analyzed the relationship between several major pathogens of respiratory infections and seasonal changes. The results showed that the positive rates of spring, summer, autumn and winter groups were 43.58%, 38.64%, 33.73% and 29.27%, respectively. Streptococcus pneumoniae and haemophilus influenzae in spring group; Mycoplasma pneumoniae in summer group; In autumn group, rhinovirus, respiratory syncytial virus, influenza A virus; The differences of chlamydia pneumoniae, Boca virus and influenza B virus in winter group were statistically significant compared with other groups in different seasons (P < 0.05). The positive detection rate of Streptococcus pneumoniae and haemophilus influenzae in the spring group was higher, which was consistent with the higher infection rate of streptococcus pneumoniae in spring and winter reported by Xiao Jingen^[12] than in summer and autumn, and the higher infection rate of haemophilus influenzae in spring and winter reported by Han Guangyue^[13], which was due to the cold in spring and winter, the imbalance between cold and warm conditions, and improper parental care. Causing the child to catch cold. In this study, the positive rate of mycoplasma pneumoniae was the highest in summer. Wang Jia^[14] treated 1,700 children with respiratory tract infection in Heyuan City, Guangdong Province, and the results showed that the detection rate of mycoplasma pneumoniae in children with respiratory tract infection in summer was the highest, reaching 31.11%, which was consistent with this study. Li Wenlian^[15] treated 2520 children with respiratory tract infection in Zunyi area, and the results showed that the detection rate of mycoplasma pneumoniae in children with respiratory tract infection was the highest in winter, reaching 34.15%, which was not consistent with this study, and it was speculated that there were certain differences in the seasons of mycoplasma pneumoniae infection in different regions. The results of this study showed that rhinoviruses and influenza A viruses were prevalent to a certain extent in autumn in Zibo area, which was basically consistent with the high positive detection rate of rhinoviruses and influenza A viruses in 292 respiratory samples from Shunyi District of Beijing reported by Zhu Hongxia ^[16]. The high positive detection rate of respiratory syncytial virus in autumn is inconsistent with the high positive detection rate of respiratory syncytial virus reported by Zhu Hongxia^[16] from February to April, indicating that there are seasonal differences in respiratory syncytial virus infection between this region and other regions. The positive detection rate of chlamydia pneumoniae in this region was higher in winter, which was consistent with the research results reported by Li Qiuhong^[17]. The high positive detection rate of Boca virus in winter in this region is inconsistent with the high positive detection rate of Boca virus in children in Guangzhou in summer reported by Zeng Yuanhong^[18], which may be attributed to the warm and closed environment of winter heating in this region, which provides conditions for the transmission of Boca virus. The high positive detection rate of influenza B virus in this region in winter is inconsistent with the report of Zheng Jianxin^[19] on the high incidence of influenza B in children in Shanghai in April. The high incidence season of influenza B has regional characteristics, and the prevention and control of the transmission of influenza B must be combined with the local epidemiological investigation.

In this study, 4,804 samples of children with respiratory tract infection were tested for respiratory tract pathogens by PCR. The results showed that there were 2085 cases of respiratory tract pathogen mixed infection, with a positive rate of 43.40%. The main combinations were streptococcus pneumoniae + rhinovirus, Streptococcus pneumoniae + Haemophilus influenzae and Streptococcus pneumoniae + respiratory syncytial virus. They accounted for 12.90%, 10.84% and 8.30%, respectively. The reason for the above combination of infections may be that the onset of respiratory tract infection in children is urgent and the children are treated early. At this time, the viral load of most of the infected respiratory tract is low, which is beyond the detection range of PCR detection technology. It may be that rhinovirus and respiratory syncytial virus have strong proliferation capacity, and the amount of proliferation is within the detection range of PCR detection technology, and thus the above combination of mixed infections occurs. Streptococcus pneumoniae and Haemophilus influenzae are common pathogenic bacteria, which cause respiratory tract infection when children's immune ability is weak. Zhu Chanhong^[20] study found that the mixed infection is mainly mycoplasma pneumoniae + parainfluenza virus, the positive detection rate reached 3.28%, the analysis of this study respiratory tract mixed infection and Zhu Chanhong and other studies may be different reasons may be Zhu Chanhong and other studies used immunological technology, because the incidence of respiratory tract infection in children is more urgent, early treatment, IgM detection fell in the window period, The inability to detect pathogen specific IgM antibodies may also be related to the incomplete development of children's immune system and the failure to produce IgM antibodies in a timely and effective manner after pathogen infection. Mixed infection can make the diagnosis and treatment of children's respiratory tract infection more complicated. The use of PCR detection technology for respiratory tract pathogens can identify the pathogens causing respiratory tract infection in the early stage of children's infection, and provide experimental basis for clinicians to determine reasonable treatment plans in time and guide clinical medication.

With its excellent sensitivity and specificity, respiratory multiplex PCR technology can significantly improve the positive detection rate of pathogens causing respiratory tract infections in children, especially the positive detection rate of mixed infections in children, which can provide a scientific basis for clinicians to determine the treatment plan as soon as possible, greatly shorten the course of respiratory tract infections in children, and improve the treatment effect.

ARI

Acute Respiratory Infections

Strepto-coccuspneumoniae

HRV

Human rhinovirus

RSV

Respiratory Syncytial Virus

IAV

Influenza A Virus

Chlamydia Pneumoniae

Mycoplasma Pneumoniae

Haemophilus influenzae

PIV

Parainfluenza Virus

HPMV

Human metapneumovirus

HBOV

Human boca virus

HCOV

Human coronavirus

IVB

Influenza B virus.

Acknowledgements

Not applicable.

Authors’contributions

LG, SL, LY, SX, and MK analyzed and interpreted the patient data. LG was a major contributor to the writing the manuscript. LG performed the statistical analysis. MK was responsible for the conceptualization and reviewed and edited the manuscript. All authors read and approved the final manuscript.

Funding

Fund Project: National Key Research and Development Program (2018YFC0114703)Availability of data and materials.

Availability of data and materials

Not applicable.

Ethics approval and consent to participate

It has passed the review of Ethics Committee of Zibo Maternal and Child Health HospitalConsent for publication.

Consent for publication

Written informed consent for publication was obtained from all participants and their parents.

Competing interests

The authors declare that they have no competing interests.

Author details

1.Medical Genetics, Zibo Maternal and Child Health Hospital, Zibo 255000, China; 2. Clinical Laboratory of Zibo Hospital of Traditional Chinese Medicine, Zibo 255000, China.

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No competing interests reported.

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Epidemiological characteristics of common respiratory pathogens in children

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Abstract

Figures

Background

Materials and methods

1.Research object

2.Multiple PCR detection of respiratory tract in children

3.Statistical analysis

Results

1. Detection of other pathogens in children with respiratory tract infection

2. Analysis on detection of respiratory pathogens mixed infection in children

3. Detection and analysis of respiratory single pathogen infection in children of different age groups

4. Detection and analysis of respiratory single pathogen infection in children in different seasons

Discussion

Conclusion

Abbreviations

Declarations

References

Additional Declarations

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