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ObjectiveTo assess the effectiveness of 23-valent pneumococcal polysaccharide vaccine (PPV23) against Streptococcus pneumonia (Spn)-associated respiratory tract infection in children with a test-negative design study. MethodsWe recruited totally 3 339 children at ages of 2 – 9 years hospitalized due to acute respiratory illness (ARI) and being detected for Spn isolates in sputum/bronchoalveolar lavage fluid (BALF) during 2017 – 2020 at the Soochow University Affiliated Children's Hospital for a test-negative case-control design study. The collected Spn isolates were serotyped with Quellung test. Of the all the participants, 1 264 being positive for PPV23 serotype Spn (VT-Spn) in sputum/BALF specimens were assigned into a case group; the two control groups were 485 being positive for non-PPV23 serotype (NVT-Spn) and 1 590 being negative for Spn (Spn –). The participants′ information on PPV23 vaccination were collected from the Vaccination Registry Database of Suzhou Municipal Center for Disease Control and Prevention. The adjusted odds ratio of PPV23 vaccination was calculated with logistic regression model to estimate the vaccine effectiveness (VE) of PPV23 against Spn-associated ARI. ResultsAmong the 1 749 Spn strains isolated, the most common serotypes were 19F, 6B, 23F, 6A and 19A; 76.56% and 72.27% were positive for 13-valent pneumococcal conjugate vaccine (PCV13) serotype and PPV23 serotype, respectively. For the participants of VT-Spn, NVT-Spn and Spn – group, the mean age were 3.88 ± 1.27, 4.02 ± 1.43, and 4.52 ± 1.92 years; the vaccination rate of PPV23 were 5.4%, 6.8%, and 5.8%, respectively; and the mean age at the vaccination was 2.53 ± 0.61 years for all the participants vaccinated., When using the NVT-Spn group as the control and after adjusting for age, clinical diagnosis, and other vaccination status, the logistic regression analysis revealed an overall PPV23 effectiveness of 18.3% (95% confidence interval [95%CI]: – 26.3% – 47.1%) for against Spn-associated ARI, an effectiveness of 15.7% for against community acquired pneumonia, and the effectiveness of 32.8, 16.5% and 57.5% for against serotype 19F, 6B and 15B Spn-associated ARI, respectively, and the effectiveness (26.0%) was better in the participants aged 2 – 5 years than that (7.6%) in those aged older than 5 year; while using the Spn – group as the control and after adjusting for age, household registration, other vaccination status, the year of seeking medication, season of seeking medication, and clinical diagnosis, the logistic regression analysis showed a VE of 29.9% (95% CI: – 22.0% – 59.7%) for the participants aged 2 – 5 years. Conclusion PPV23 is of certain protection against Spn-associated respiratory tract infections in 2 – 9 years old children and the effectiveness decreases with the prolonged time after receiving the vaccination