3.1 Baseline characteristics
During the study period, 263 cases of IPD were diagnosed in children aged <18 years. Serotyping was not possible in 10 cases (3.8%), which were excluded. Of the 253 cases included, 151 (59.7%) were male; 87 (34.4%) were in children aged <2 years, 98 (38. 7%) in those aged 2–4 years, and 68 (26.9%) in those aged 5–17 years (Table 1).
There was a seasonal variation, with 72% of cases detected during the cool months (October to March; p<0.001).
Eighty-three patients (32.8%) had received ≥1 dose of PCV13, while 129 (51.0%) had received no dose of any conjugate vaccine.
Fifteen cases (6.0%) had a chronic disease, including congenital immunodeficiency (2 cases), congenital heart disease (3 cases), chronic pulmonary disease (1 case), chronic kidney disease (1 case), and cerebrospinal fluid leak (1 case), or had received immunosuppressive therapy in the previous 6 months (7 cases).
The most frequent clinical presentation was pneumonia (187 cases; 73.9%), of which 150 cases (80. 2%) were complicated pneumonia. It was followed by occult bacteraemia (25 cases; 9.9%) and meningitis (18 cases; 7.1%). The presentation in the remaining 23 cases (9.1%) was septic shock (7 cases; 2.8%), mastoiditis (7 cases; 2.8%), osteoarticular infection (6 cases; 2.3%), orbital cellulitis (2 cases; 0.8%) and pancreatitis (1 case; 0.4%). Figure 1 shows the distribution of clinical manifestations by age group. Complicated pneumonia was the most frequent clinical presentation in all age groups.
Forty–nine cases (19.4%) were admitted to the ICU: 7 of 7 (100%) with septic shock, 15 of 18 (83.3%) with meningitis, 23 of 150 (15.3%) with complicated pneumonia, 1 of 7 (14.3%) with mastoiditis, 2 of 37 (5.4%) with non-complicated pneumonia and 1 of 25 (4%) with occult bacteraemia (in this case ICU admission was caused by convulsions). Of the 49 ICU cases, 55.1% were caused by PCV13 serotypes, and 14.3% had an underlying disease.
Two patients died: an 18-month-old child with serotype–35F meningitis who had congenital immunodeficiency, and a 23-month-old child with septic shock caused by a non-vaccine serotype which was not identified.
3.2 Serotypes and clinical presentation
One hundred and sixty-three (64.4%) cases were due to PCV13 serotypes (Figure 2); It was 41.5% in patients who had received ≥ 1 dose of PCV13 (34/82 cases) and 75.2% in patients who had not received any conjugated vaccine (97/129; p<0.001).
The most frequent serotypes were: 3 (20.9 %), 1 (19.0%), 19A (6.7%) and 14 (5.9%) all included in PCV13.
Overall and complicated pneumonia were mainly caused by PCV13 serotypes (143/187 cases, 76.4% and 123/150 cases, 82.0%, respectively). PCV13 serotypes caused overall pneumonia in 87.7% of cases and non-PCV13 serotypes in 48.9% (OR: 7.47 [4.0–13.96]). Similarly, PCV13 serotypes led to complicated pneumonia in 75.5% of cases and non-PCV13 serotypes in 30% (OR: 7.2 [4.04–12.75]) (Table 2).
Serotype 3 caused overall pneumonia in 92.5% of cases, especially complicated pneumonia (83.0%), and was associated with both clinical presentations (OR:5.5 [1.9–15.92] and, OR:4.3 [2.01–9.35], respectively). Serotype 1 was also associated with overall pneumonia (OR: 21.82 [2.95–161.63]) and complicated pneumonia (OR: 5.16 [2.21–12.04]).
Meningitis was mainly caused by non-PCV13 serotypes (14/18, 77.8%). Although there was no predominant serotype, non-PCV13 serotypes caused meningitis in 15.6% of cases and PCV13 serotypes in 2.5% (OR: 7.32 [2.33–22.99]).
Occult bacteraemia was also caused mainly by non-PCV13 serotypes (16/25, 64%). Non-PCV13 serotypes caused occult bacteraemia in 17.8 % of cases and PCV13 serotypes in 5.5% (OR: 3.6 [1.56–8.76]). Some non-PCV13 serotypes were associated with occult bacteraemia namely 12F (OR: 19.73 [1.72–226.12]) and 10A (OR: 10.22 [1.94–53.74]). Also serotype 18C (PCV13 serotype), showed a slight association (OR: 48.61 [2.26–1043.02]).
Other clinical presentations were caused mainly by non-PCV13 serotypes (16/23; 69.6%). Non-PCV13 serotypes caused these presentations in 17.8% of cases and PCV13 in 4.3% (OR: 4.81 [1.90–12.22]).
3.3 Serotypes and clinical presentation according to age group
PCV13 serotypes were most frequent in the 5–17 years age group (77.9%; OR: 2.41 [1.27–4.59]) and non-PCV13 in <5 years, mainly in the <2 years age group (55.2% OR: 3.63 [2.10–6.29]. Serotype 19A was more frequent in the <2 years age group (58.8%; OR: 2.95 [1.08–8.05]), serotype 3 in the 2–4 years age group (58.5%; OR: 2.79 [1.50–5.20]) and serotype 1 in the 5–17 years age group (60.4%; OR: 6.50 [3.31 –12.77]) (Figure 2).
Table 3 and figure 3 show the serotype distribution by age groups and the association to clinical presentation.
In children aged <2 years, PCV13 serotypes were associated with overall pneumonia (OR: 5.29 [2.08–13.41]) and complicated pneumonia (OR: 4.84 [1.91–12.22]). Serotype 3 caused overall pneumonia in 91.7% of cases (OR: 12.57 [1.54–102.33]) and complicated pneumonia in 83.3% (OR: 10.62 [2.16–52.3]). Serotype 19A was associated with complicated pneumonia (OR: 4.32 [1.03–18.07]) in 70% of cases.
In the 2–4 years age group, PCV13 serotypes were associated with overall pneumonia (OR: 5.42 [1.70–17.23]) and complicated pneumonia (OR: 6.92 [2.61–18.36]), while non-PCV13 serotypes were associated with meningitis (OR: 27.38 [1.42–527.7]). Serotype 3 was associated with complicated pneumonia (OR: 6.7 [1.85–24.24]) and serotype 9V with occult bacteraemia (OR: 102.7 [4.27–2472.3]).
In the 5–17 years age group, PCV13 serotypes were associated with overall pneumonia (OR: 8.17 [1.91–34.86]) and complicated pneumonia (OR: 6.05 [1.81–20.18]), whereas non-PCV13 serotypes were associated with other clinical forms [OR: 9.3 [1.51–57.12]]. Serotype 1 was associated with complicated pneumonia (OR: 10.43 [2.17–50.12])
3.4 Serotypes and method of diagnosis
The diagnosis was made by culture in 62 cases (24.5%), culture and RT-PCR in 69 (27.3%) and by RT-PCR in 122 (48.2%).
PCV13 and non-PCV13 serotypes showed no significant difference in the percentage of cases diagnosed exclusively by PCR (52.1% versus 41.1%).
There were significant differences between serotypes in the method of diagnosis: 86.8% of cases due to serotype 3 were diagnosed only by PCR, versus 38% in the other serotypes (p<0.001). By contrast, other serotypes, such as serotype 14 and 24F were diagnosed less frequently by PCR (13.3%, 2/15 and 0%, 0/6) than the other serotypes (p = 0.005 and p = 0.030, respectively).
3.5 Non-susceptible antibiotics by serotypes
Antimicrobial susceptibility was studied in 122/131 (93.1%) strains isolated. Forty-four cases (36.1%) were non-susceptible to penicillin, and 4 cases (3.3%) were penicillin resistant. Twenty cases (16.4%) were also non-susceptible to cefotaxime (Table 4).
There were no significant differences in the percentage of non-susceptible penicillin strains between PCV13 serotypes (36.1%) and non-PCV13 serotypes (36.0%). PCV13 serotypes were associated with strains not susceptible to cefotaxime (p = 0.010) and with isolates not susceptible to both penicillin and cefotaxime (p = 0.010).
Serotype 19A showed a penicillin MIC >0.06 mg/L in 90.9% of cases studied, (p<0.001) and MIC>2 mg/L in 27.3% (p = 0.002). In addition, 54.5% of serotype 19A strains were non-susceptible to cefotaxime (p = 0.003) and also non-susceptible to penicillin (p = 0.003).
All serotype 14 strains (100%) showed a penicillin MIC >0.06 mg/L (p<0.001), and 81.8% of cases were non-susceptible to both cefotaxime (p<0.001) and penicillin (p<0.001).
Serotype 11A showed non-susceptibility to cefotaxime in 60% of strains compared with 14.5% for other serotypes (p = 0.031). These strains also were non-susceptible to penicillin (p = 0.031).
Other serotypes showed significant differences in antimicrobial susceptibility, namely 24F which showed penicillin MIC >0.06 mg/L in 83.3% (p = 0.023), and 23B in 100% (p = 0.045).