Pneumococcal Burden in Hospitalized Adults in Bogotá, Colombia. 


 Background: The incidence of invasive pneumococcal disease (IPD) varies depending on a number of factors, including vaccine uptake, in both children and adults, the geographic location, and local serotype prevalence. There are limited data about the burden of Streptococcus pneumoniae (Spn), serotype distribution, and clinical characteristics of adults hospitalized due to IPD in Colombia. Thus, this study will attempt to bridge this gap in the literature. Methods: This was an observational, retrospective, a citywide study conducted between 2012 and 2019 in Bogotá, Colombia. We analyzed reported positive cases of IPD from 55 hospitals in a governmental pneumococcal surveillance program. Pneumococcal strains were isolated in each hospital and typified in a centralized laboratory. The objectives of this study included assessment of Spn serotype distribution, clinical characteristics, mortality, ICU admission, and the need for mechanical ventilation. Results: A total of 310 patients with IPD were included, of whom 45.5% were female. The leading cause of IPD was pneumonia (60%, 186/310), followed by meningitis. The most frequent serotypes isolated were 19A (13.87%, 43/310) and 3 (11.94%, 37/310). The overall hospital mortality rate was 30.3% (94/310). Moreover, 52.6% (163/310 patients) were admitted to the ICU, 45.5% (141/310) required invasive mechanical ventilation and 5.1% (16/310) non-invasive mechanical ventilation. Conclusion: Pneumococcal pneumonia is the most prevalent cause of IPD, with serotypes 19A and 3 being the leading cause of IPD in Colombian adults. Mortality due to IPD in adults continues to be very high. 


Introduction
Invasive pneumococcal disease (IPD) is the leading cause of death in developing countries. According to the United States Active Bacterial Core Surveillance, in 2010, the incidence of IPD in ≥ 65 year old adults was 36.4 cases per 100,000 [1]. In Latin America, it is estimated that every year, IPD is responsible for up to 28,000 deaths, 182,000 hospitalizations, and 1.4 million outpatient consults [2]. Annually, more than 10 billion dollars are expended in healthcare-associated costs in the United States alone due to IPD. It is essential to highlight that pneumococcal infection is more frequent in children; however, its fatality rate is higher in older adults. Thus, pneumococcal infections have been considered as a public health care problem. Importantly, mortality due to IPD in adults has remained steady even after the extensive usage of broad-spectrum antibiotics and better vaccination coverage in children [2][3][4].
Over 99 serotypes of Streptococcus pneumoniae (Spn) have been described [5]. Each pneumococcal serotype has a different genetic and phenotypical composition; some of them are highly virulent to mammals [5]. Some Spn are more frequently associated with the development of IPD, depending on various factors that include vaccine undertake and geographic location. Importantly, pneumococcal vaccines are the most effective strategy to reduce IPD incidence [6]. There are two types of antipneumococcal vaccines: pneumococcal conjugated vaccines (PCV 7, PCV 10, and PCV13) and a pneumococcal polysaccharide vaccine (PPSV23). PCV13 includes puri ed capsular polysaccharide of 13 serotypes of Spn that frequently cause pneumococcal disease (1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 19A, 19F, 18C and 23F). This vaccine has polysaccharide antigens conjugated to a protein carrier (CRM197) to improve the immunogenicity [7]. The pneumococcal polysaccharide vaccine includes antigens from 23 serotypes of pneumococci. PPSV23 vaccine is composed of long chains of sugar molecules, polysaccharides, that are components of the pneumococcal serotypes contained in the vaccine, which stimulate the immune system to produce antibodies [8]. Different studies have shown that more than 80% of healthy adults who receive PPSV23 develop antibodies against the serotypes contained in the vaccine; the immune response usually occurs within 2 to 3 weeks after vaccination in adults. Overall, the vaccine is 60-70% effective in preventing invasive disease in infants [9][10][11]. The Centers for Disease Control and Prevention (CDC) recommends that PCV13 should be given to all children younger than two years old. On the other hand, one dose of PPSV23 is recommended in immunocompetent adults older than 65 years. Likewise, the CDC recommends PPSV23 in people 2 through 64 years old with certain medical conditions or in adults who smoke cigarettes [3,[12][13][14].
Since the introduction of the mandatory universal pneumococcal vaccination in children, IPD epidemiology has changed signi cantly worldwide [15]. Currently, studies are describing yearly increases in the incidence of IPD due to several non-vaccine serotypes [16]. For instance, in the United States, serotypes 19A, 6C, 23A, 15 B/15C, 31, and C7 are emerging as a frequent cause of IPD in adults [17]. In Colombia, some studies have reported serotypes 1 and 14 as the most prevalent serotypes causing IPD in children [18,19]. However, these ndings were obtained from small studies of children admitted to few hospitals. Another study identi ed the molecular characteristics of serotypes 11A, 23A, 15B/C, aiming at documenting the importance of capsular switching in the spread of non-vaccine serotypes causing IPD in children [20]. However, there is a lack of epidemiological data regarding IPD burden, serotype distribution, and clinical characteristics of adults hospitalized due to IPD in Colombia. Thus, this study was designed to ll that gap in the literature. We hypothesized that adult patients hospitalized due to IPD in Colombia would be infected, more frequently, with pneumococcal serotypes not included in the conjugate vaccines.
Moreover, we also hypothesized that these patients would develop community-acquired pneumonia (CAP) as the primary clinical syndrome causing IPD, with higher mortality in older adults. The current study was undertaken with the objectives of assessing the Spn serotype distribution, clinical diagnosis, mortality, ICU admission, length of hospital stay, and need for mechanical ventilation in adults admitted due to IPD.

Material And Methods
This was a retrospective multicenter observational study of patients from 55 hospitals, including public and private institutions, in Bogotá, Colombia, that provide health care to about 8 million people. The study was undertaken between January 2012 and January 2019. All patients older than 18 years of age who were diagnosed with IPD were identi ed and those cases with a fully available clinical history, and positive Spn cultures were included in the study. Patients in whom clinical data were incomplete or other bacteria were isolated (i.e., co-infections) were excluded. The ow chart indicating the selection process is part of the supplemental information in S1 g. The Translational Science in Infectious Diseases and Critical Care Medicine Research Group from the Universidad de La Sabana developed this study protocol in collaboration with The Public Health Secretary of Bogotá city.
We used the bacterial isolates reported to The Public Health Secretary under a Spn surveillance program. All pneumococcal isolates were con rmed and typed by the National Center of Microbiology (CNM) in a centralized laboratory following the Quellung reaction using polyclonal antisera [21]. This study was approved, and the use of informed consent was waived the Institutional Review Board of the Clinica Universidad de La Sabana (2020_MED-23221). The use of informed consent was waived because this was a retrospective study, without any intervention, and used only a chart review to obtain clinical data.
All methods were performed in accordance with the relevant guidelines and local and international regulation.

Study de nitions
We used international and well-accepted de nitions for each variable and clinical diagnosis. IPD was de ned as an infection con rmed by the isolation of Spn from a normally sterile site (e.g., blood, cerebrospinal uid, and pleural, joint, peritoneal uid and/or respiratory uid other than sputum) [22]. CAP was de ned according to the American Thoracic Society and Infectious Diseases Society of America (ATS/IDSA) guidelines as an acute infection of the pulmonary parenchyma acquired outside of the hospital setting evidenced by radiological ndings (e.g., chest radiograph, computed tomography or pulmonary ultrasound) compatible with alveolar in ltration and characteristic clinical presentation (e.g., acute onset of cough, fever, tachypnea, altered mental status, diaphoresis, etc.) [23]. Pneumococcal meningitis was de ned as the identi cation of Spn in blood cultures and/or CSF in association with the clinical syndrome of meningitis (i.e., fever, headache, meningismus, altered mental status, seizures, confusion or vomiting) According to the IDSA guidelines [24].
To reduce the risk of bias, only con rmed cases of IPD according to international clinical guidelines were included and all the clinical records were evaluated by trained personnel. Additionally, all missing information was not analyzed or inferred, and all clinical charts were reviewed blinded to the isolated pneumococcal serotypes to determine clinical outcomes.

Variables
Data on gender, age, comorbidities, chronic medication, vaccination, physiological admission variables, blood count, bilirubin levels, liver enzymes, arterial blood gases, medical intervention, and antibiotic treatment were recorded for each patient. The initial severity of the disease was assessed using the Sequential Organ Failure Assessment (SOFA), quick Sequential Organ Failure Assesment (qSOFA) [25], and Acute Physiology and Chronic Health Evaluation II (APACHE II) [26]. During hospitalization, ICU admission, need for invasive mechanical ventilation or non-invasive mechanical ventilation, length of hospital and ICU stay were recorded and considered as secondary outcomes. All-cause hospital mortality was the primary outcome.

Data collection
Hospitals sent all isolated Spn strains to a centralized laboratory where each one was characterized. Clinical records were sent to the district health secretary of Bogotá city, and a retrospective review of all medical records was carried out to gather clinical information. Data was recorded onto an electronic case report form (eCRF) hosted in the servers of the health secretary of Bogota and then exported for statistical analysis.

Statistical analysis
For the statistical analysis, descriptive analysis was performed with measures of central tendency and dispersion for the quantitative variables, and frequencies with percentages for the qualitative variables. Numerical data are presented as mean (standard deviation, SD) or median (interquartile ranges, IQR) according to normality distribution. The serotype distribution of Spn was broken down by clinical diagnosis and age. Finally, mortality, ICU admission, need for invasive mechanical ventilation or noninvasive mechanical ventilation, and all-cause hospital mortality were calculated for the group as a whole as well as by diagnosis of IPD. Quantitative variables were contrasted by sher, and continuos variables were analyzed according to data distribution (i.e., normality) by t-test the Mann Witney. A p < 0,05 was considered statistically signi cant. All statistical analyses were performed using a statistical package SPSS 25 and Graph Pad Prism for MAC, licensed for the Universidad de La Sabana.
Similarly, in Colombia several studies have investigated serotype distribution before the introduction of PCV10 mass vaccination at the end of 2011 in Colombia [32]. Parra E.L. et al. [2]. concluded that serotype 14 was the most prevalent pneumococcal serotype in children and serotype 3 in older adults between 2005-2010. Later, in 2019 Camacho Moreno et al. [33]. found that serotype 19A was the leading cause of IPD in Colombian infants, being pneumonia its principal clinical presentation, followed by bacteremia and meningitis that is consistent with our results that evidenced that pneumonia and meningitis are the main clinical presentation in adults. Likewise, these ndings provided clinical data regarding serotype distribution in infants and evidenced the serotype replacement that followed the massive undertake of PCV10 in pediatric population. There has not been a prior study to document similar ndings in an adult population in Colombia. Our study presents for the rst time, that as described in children, strains 19A and 3 are the main cause of IPD in Colombian adults, probably due to indirect effects on pneumococcal transmission via herd protection. Interestingly, serotype 3 did not infect young adults (18-35 years old) but was the leading cause of IPD in patients ≥ 65 years old. Those ndings highlight the importance of prioritizing vaccination programs in adults as pneumococcal disease continues to be highly prevalent infection.
Our results show that the most prevalent pneumococcal serotypes are not included in the PCV10, which is part of the obligatory immunization program in Colombia for children < 2 years old. Paradoxically and despite the Colombian guideline recommendations for the use of PCV13 and PPV23 in adult population, serotypes 19A and 3 were the main cause of IPD in our cohort. This phenomenon could be explained by the low rate of vaccine PCV13 1.3% and PPV 23 0.6% (Table 1) undertake in this population and supported by what was previously mentioned, the PCV10 mandatory vaccination in children. Additionally, PCV 13 has been shown to have low immunogenicity for serotype 3 and 19A, which might also contribute to the high prevalence of these serotypes [34,35]. It has been well-described that low vaccine effectiveness against speci c Spn serotypes may explain some of the reasons for the distribution of serotypes. Serotype 3, which was the most prevalent serotype identi ed in older adults in our cohort, is hyper-encapsulated as it has a rather simple capsule con guration, and its massive production is metabolically inexpensive when compared to other serotypes. Additionally, this capsule is not covalently linked to its cell wall and is continuously shed into the bacterial environment and acts distally to the bacterium as a decoy for antibodies generated by the vaccine. More importantly, antibodies and complement factors that become deposited on bacteria-associated capsules would also be shed. As a result, the vaccine does not introduce su ciently high antibody titers to promote adequate clearance of serotype [5]. Taking these potential explanations into account, it is important to promote public health programs adult vaccination with PCV13 and PPV 23 to evaluate its coverage.
Our study is subjected to limitations and strengths that are important to acknowledge. First, we analyzed a relatively small cohort of patients; however, it is a multicentre study that carriages a valuable number of cases. Moreover, the data presented in this manuscript might serve as starting point to carry out a bigger study that can truly represent Colombian IPD. Furthermore, being a study in Colombia alone, it may not be replicable to other countries, although the major ndings were similar to that of those previously published international studies. Moreover, as it is a retrospective study, some information was missing from medical records, leading to an abnormal data distribution of some of the physiological variables and exclusion of several patients. Additionally, due to the characteristics of our health system, there is several discrepancies about the number of days patients are treated for IPD; thus, readers should only extrapolate these results to countries with similar health care systems. We did not assess ICU mortality rate because not all patients were admitted to a hospital with ICU capabilities. Another limitation is that we did not follow the patients over time; thus, we cannot analyze long-term outcomes. Nevertheless, the results appear to re ect the overall trend of changes in the Spn serotype distribution in Colombia, with the inclusion of around 50% of isolates during a period of 8 years from the capital city. Importantly, we studied only adult patients, which is an essential strength of our study because even though children develop IPD more frequently than adults, the older population has a higher mortality rate. The surveillance isolation program is critical to identify the replacement serotypes and redirect vaccine development. Despite the limitations, our study shows that Spn continues to cause severe IPD in Colombian adults. Continuous monitoring of serotypes and pneumococcal disease is necessary for a better understanding of the pneumococcal burden trends and epidemiology.
In conclusion, this study provides information on the current circulating pneumococcal serotypes in adults hospitalized due to IPD in Bogota, Colombia. Pneumococcal pneumonia continues to be the most prevalent cause of IPD in this population. Comorbidities and smoking are commonly found in these patients therefore should be carefully observed to achieve early diagnosis is and treatment. Further research is needed in order to develop more immunogenic vaccines against the most prevalent serotypes strains of Spn 19A, 3, and 6A that still occur in certain countries. Likewise, there is a necessity in the Colombian adult population to promote pneumococcal vaccination as recommended by the national guidelines. More extensive prospective studies are necessary to assess future changes in the epidemiology and to make the best decisions regarding immunization strategies.

Declarations
Ethics approval and consent to participate: This study was approved, and the use of informed consent was waived the Institutional Review Board of the Clinica Universidad de La Sabana (2020_MED-23221). The use of informed consent was waived because this was a retrospective study, without any intervention, and used only a chart review to obtain clinical data. Local ethical guidelines were followed to carry out this project.
Consent for publication: Not applicable.
Availability of data and materials: The datasets used and/or analysed during the current study available from the corresponding author on reasonable request.

Supplementary Files
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