Epidemiological and clinical profile of infective endocarditis at a Brazilian tertiary care center: an eight-year prospective study

Introduction: Infective endocarditis (IE) is a systemic infectious disease requiring a multidisciplinary team for treatment. This study presents the epidemiological and clinical data of 73 cases of IE in Rio de Janeiro, Brazil. Methods: This observational prospective cohort study of endocarditis patients during an eight-year study period described 73 episodes of IE in 70 patients (three had IE twice). Community-associated (CAIE) and healthcare-acquired infective endocarditis (HAIE) were diagnosed according to the modified Duke criteria. The collected data included demographic, epidemiologic, and clinical characteristics, including results of blood cultures, echocardiographic findings, surgical interventions, and outcome. Results: Analysis of data from the eight-year study period and 73 cases (70 patients) of IE showed a mean age of 46 years (SD=2.5 years; 1–84 years) and that 65.7% were male patients. The prevalence of CAIE and HAIE was 32.9% and 67.1%, respectively. Staphylococcus aureus (30.1%), Enterococcus spp. (19.1%), and Streptococcus spp. (15.0%) were the prevalent microorganisms. The relevant signals and symptoms were fever (97.2%; mean 38.6 + 0.05°C) and heart murmur (87.6%). Vegetations were observed in the mitral (41.1%) and aortic (27.4%) valves. The mortality rate of the cases was 47.9%. Conclusions: In multivariate analysis, chronic renal failure (relative risk [RR]= 1.60; 95% confidence interval [CI] 1.01–2.55), septic shock (RR= 2.19; 95% CI 1.499–3.22), and age over 60 years (RR= 2.28; 95% CI 1.44–3.59) were indirectly associated with in-hospital mortality. The best prognosis was related to the performance of cardiovascular surgery (hazard ratio [HR]= 0.51; 95% CI 0.26–0.99).


INTRODUCTION
Infective endocarditis (IE) is a systemic infection for which a multidisciplinary team is required to approach treatment of this life-threatening disease [1][2][3][4] .In past years, the overall incidence of IE in the general population in developed nations has ranged between 3 and 10 cases per 100,000 individuals per year.However, recent epidemiological studies have suggested an increasing incidence of IE 5,6 .Despite trends toward earlier diagnosis and surgical intervention in IE patients in the 21 st century, in-hospital mortality rates have not improved over the last three decades 2,3,5 .In developed countries, IE is associated with high mortality rates of approximately 20%, despite medical and surgical advances 3,5,7 .In low-and middle-income countries, the mortality of IE ranges from 19 to 46% 8 .In these countries, the morbidity and mortality rates of IE are similar to those reported in the middle of the 20 th century in North America and European countries 7,8 .

Echocardiographic data
Transthoracic and transesophageal echocardiography was performed at the echocardiography service of HUPE.All echocardiography exams in which IE was suspected were reviewed by an expert in echocardiography.

Microbiological data
Bloodstream samples were collected, placed in at least three aerobic bottles (Plus Aerobic), and incubated for five days in a BACTEC 9240 blood culture instrument (BATEC/ALERT®, BioMérieux, Durham, NC, USA).Bacterial identification and antimicrobial susceptibility testing were performed using an automated VITEK 2 System (BioMérieux).Additionally, the vancomycin minimal inhibitory concentration (MIC) was determined by E-test and microdilution for Enterococcus sp. and methicillin-resistant Staphylococcus aureus (MRSA), respectively.
MecA and SCCmec typing were carried out by multiplex polymerase chain reaction (PCR) analysis that generated specific amplification patterns for SCCmec types I, II, III, IV, and V 10 ; in addition, molecular characterization based on multilocus sequence typing (MLST) was performed for all MRSA isolates 11 .
Microorganisms obtained from blood culture or after valve biopsies were identified by 16SrRNA sequencing or by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF).Serological and molecular Coxiella burnetii analyses were performed in all blood culture-negative endocarditis cases (BCNE) using an indirect immunofluorescence assay (Focus Diagnostics TM, Cypress, CA, USA) and PCR amplification of a fragment of the repetitive element IS1111 of a heat-shock protein gene 12 .

Statistical analysis
Data were input to Excel 2007 and analyzed using STATA TM version 9.1.A descriptive analysis of the variables was performed.The relative risks (RRs) or hazard ratio (HRs) with 95% confidence intervals (CIs) were calculated for categorical variables, and v2 or Fisher's exact tests for expected values of <5 were also used to determine statistical significances.Logistic regression was used to examine the potential associations between variables and multivariate analysis was performed to identify risk factors independently associated with mortality.Variables with p >0.05 were excluded from the final model.P-values <0.05 were considered statistically significant.

Demographic and Epidemiologic findings
Demographic, clinical, microbiological, and complication data in the cohort of IE patients are shown in Table 1.
In the past eight years, 73 cases (70 patients) with IE were followed-up, with 0.75 episodes per 1,000 patients in the university hospital.The mean age of the patients was 46 (SD 2.5 years) (range: 1-84 years) and 65.7% of patients were male.
The mortality rate of all cases of IE was 47.9%.The results of two-dimensional transthoracic (TTE) or transesophageal echocardiography (TEE) of the 73 cases showed a mean diameter of valve vegetation of 1.3 ± 0.1 cm.In all IE cases, the mitral valve (41.1%) was the most frequent site of vegetation, followed by the aortic valve (27.4 %).Most patients developed native endocarditis (90.4%).

Clinical data
All 73 cases of IE fulfilled the Duke criteria.Only 38.4% of patients had classic clinical evidence of IE at hospital admission (Table 1).The clinical signs and symptoms that helped the physicians to diagnose IE upon admission to the university hospital are described.The median time between symptom onset and hospital admission was 27.5 ± 3.7 days.The most relevant classical signs and symptoms of IE were fever (97.2%; mean 38.6 ± 0.05°C) and heart murmur (87.6%).Change in heart murmur was observed in only 17.8% of patients and the following stigmas of IE were found: arterial embolic phenomenon (20.5%),Janeway lesion (5.5 %), petechiae (4.1%), and Roth spot (4.1%).The principal antecedent factors in our cohort were central venous catheterization (43.8%), hemodialysis (32.8%), antecedents of IE (16.4%), rheumatic fever (12.3%), previous dental manipulation (9.5%), and intravenous additive (1.3%) (Table 2).Our data also showed a higher number of patients with different types of comorbidities, including chronic renal failure in 27 patients (36.9%), diabetes mellitus in eight patients (10.9%),HIV in seven patients (9.5%), kidney transplants in seven patients (9.5%), and neoplasia six patients (8.2%) (Table 2).

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The prevalence of CAIE and HAIE was 32.9% and 67.1%, respectively.Of the 73 IE cases, 32.8% were classified as nonnosocomial HAIE and 34.3% as classical nosocomial IE, as previously described 1 .Only three patients developed a second episode of endocarditis during the study period (Table 1).The most frequent complications due to IE were sepsis (26.0%), acute renal failure (24.6%), and septic shock (19.1%) (Table 2).

Prevalence of identified organisms in IE
At least three blood cultures were collected from each patient, resulting in 89.0%positive blood cultures, whereas only 9.6% remained without an identified etiology.In total, an isolate was obtained in 90.4% of cases (89.0% of cases of IE with positive blood culture and one with a positive valve culture) (Table 1).
The classic agents of IE included S. aureus (30.1% of cases), Enterococcus spp.(19.1%) and Streptococcus spp.(15.0%).Six of the S. aureus isolates were methicillin resistant and contained the mecA gene.The MIC for vancomycin by microdilution ranged from 0.5 to 2.0 µg mL -1 .Analysis of the MRSA strains by MLST revealed five different sequence types (ST) (1, 5, 25, 105, and 188).Among MRSA strains, the SCCmec types included I, II, and IV.PVL toxin genes were detected in 33.3% of all MRSA strains (ST105 and ST1).In addition, two of six MRSA isolates belonged to known epidemic lineages (USA400 and USA800).
Four cases (5.4%) of IE due to vancomycin-resistant Enterococcus faecium (VRE) were found.MRSA and VRE strains were only diagnosed in HAIE patients.
As mentioned, Streptococcus spp. was identified in 15.0% of patients with IE in Rio de Janeiro.The viridans group was isolated most frequently (10.9% of all IE cases).Nonhemolytic S. mutans was isolated from two positive cases and one β-hemolytic S. agalactie strain was also isolated.
The prevalence of IE caused by Gram-negative bacteria was 8.2%, corresponding to six patients.Five patients developed HAIE after Gram-negative bacteremia in our hospital, in which vascular catheters were the major source of these microorganisms.Three bacteremia cases were due to Serratia marcescens, one to Burkholderia cepacea, one to Acinetobacter baumannii complex, and one to Escherichia coli.
One patient developed CAIE due to E. coli bacteremia from a urinary infection.E. coli analysis by whole genome sequencing (WGS) identified the isolate as ST-69.The isolate was resistant to ampicillin, trimethoprim, and trimethoprim/sulfamethoxazole and had different virulence genes associated with iron uptake systems (aer, chuA, fepA, fhuA, fhuE, fyuA, irp2, and sitA) and adhesion (fimH and fimA).
In our cohort, five patients had ERFP, four patients of which had HAIE.Two developed IE after candidemia.The other two cases of HAIE are described hereafter.The first case involved a woman with a kidney transplant who was investigated for fever of unknown origin (FUO), from which we isolated Rhodotorula mucilaginosa in aortic valve culture.In the second case, Corynebacterium spp. was initially found in the blood culture.However, 16S rRNA sequencing identified Microbacterium testaceum.Only one patient developed CAIE due to Corynebacterium spp.
Serological and molecular analyses of Coxiella burnetii were performed in all blood culture-negative endocarditis cases (BCNE) using an indirect immunofluorescence assay.One case showed serological evidence for C. burnetii but Q fever was excluded because the male patient was diagnosed with systemic lupus erythematosus with Libman-Sacks endocarditis and antiphospholipid syndrome (Table 2).

DISCUSSION
IE is a life-threatening systemic infectious disease, in which a multidisciplinary group of specialists is required for the case treatment and follow-ups 2 .Despite IE being a rare pathology, its incidence has increased in developed countries and few studies are available in developing countries 2,6 .Endocarditis is the fourth most important life-threating infectious syndrome after urosepsis, pneumonia, and intra-abdominal sepsis in medical departments in developed countries 13 .
In this IE cohort, the prevalence of CAIE and HAIE was 32.9% and 67.1%, respectively.In addition, 32.8% were episodes of non-nosocomial HAIE and 34.3% were nosocomial IE.Therefore, a higher prevalence of HAIE was observed than those reported in recent epidemiological studies of IE in lowand middle-income countries 7,8 .
All patients with MRSA bacteremia as the cause of IE developed HAIE and we did not observe any case of MRSA due to CAIE.Among the six MRSA IE cases, five different STs were found, in which two isolates were related to known epidemic lineages (USA400/ST1/SCCmec IV and USA 800/ ST105/SCCmec IV).Some epidemic clones with worldwide distribution can be identified by the characterization of SCCmec, ST, and PVL, as we presented in our results.
The in-hospital mortality in six patients with MRSA IE was 50%.In the present cohort, 100% (n=4) of the patients with vancomycin-resistant E. faecium HAIE died during treatment.The overall in-hospital mortality was 47.9% and 10.9% and 36.9% in the CAIE and HAIE patient groups, respectively.We observed the best outcome in the group of patients that received clinical and surgical treatment for the IE episode, according to the guidelines for the management of IE 2 .
In most developing countries, the epidemiology of IE remains similar to the rates reported in North America and Europe in the twentieth century; the disease often affects a younger age group, is associated with rheumatic heart disease, and is predominantly caused by streptococci 6,8 .The present study showed a different scenario in Rio de Janeiro, where, in 73 cases of IE, S. aureus was the most frequent cause of IE, followed by Enterococcus spp and Streptococcus spp.The current epidemiology of IE in Brazil may be more similar to that of North America, where S. aureus and Enterococcus spp are the most significant agents of IE 16,18 .
In general, the prevalence of positive blood cultures in IE patients was 87.6%. Figure 1 reviews 21 IE studies performed in low-and middle-income countries during the same period of our investigation.The authors specified the frequency of positive and negative blood cultures and only elicited papers included studies presenting the data for etiology, prevalence of CAIE and HAIE, and outcome of disease.This resulted in 3,592 patients with IE, as defined by the Duke criteria, of which 2,362 (65.7%) and 1,109 (30.8%) had positive and negative blood culture results, respectively (Figure 1).Among the positive blood cultures, Streptococcus spp. was isolated in 852 cases (23.7%),S.aureus in 687 (19.1%), and Enterococcus spp. in 102 (2.8%).The prevalence of CAIE and HAIE were 46-94% and 9-56.3%, respectively.The mortality rate of inpatients was 7.3-46.4%.
The difference in the etiology of IE and outcome in our study may have been due to the high proportion of patients with many comorbidities and the low prevalence of rheumatic heart disease.The principal comorbidities in our patients with IE were chronic renal insufficiency (36.9%), diabetes mellitus (10.9%), kidney transplantation, and neoplasia (8.2%), similar to those observed in the ICE cohort study 18 .In our series, one reason for the high prevalence of S.aureus and Enterococcus spp.IE may have been due to the fact that many patients had vascular devices (Table 2) compared to the proportions in other studies of IE in developing countries.
The most relevant classical signs and symptoms of IE were fever (97.2%), and heart murmur (87.6%), similar to those found in other studies 19 .The classic Oslerian manifestations of endocarditis were present only in 38.4% of patients with IE at admission, which corroborates to the indication for echocardiography in cases of bacteremia due to Staphylococcus spp., Enterococcus spp., and Streptococcus spp. in teaching hospitals [2][3][4][5]19 .
In our multivariate analysis, chronic renal failure (RR = 1.60; 95% CI 1.01-2.55;p <0.05), septic shock (RR 2.19; 95% CI 1.499-3.22;p <0.01), and age over 60 years (RR 2.28; 95% CI 1.44-3.59;p <0.01) were indirectly association with in-hospital mortality.As expected, the best prognosis was for valve and cardiovascular surgery (HR = 0.51; 95% CI 0.26-0.99;p <0.05).In our review of IE in developed countries, we had access to only 12 studies that included data from multivariate analysis of in-mortality rates in IE patients.These studies showed statistically significant relationships for age over 45 years, dialysis, chronic renal failure, septic shock, heart failure, prosthetic dysfunction, nosocomial IE, neoplasia, mobile vegetation, mental alteration, central nervous system emboli, coronary artery disease, aortic vegetation, and large vegetation.
The experience from a single teaching hospital and failure to investigate all patients by transesophageal echocardiography may have been limiting factors in the results of the present study.However, this eight-year prospective study of endocarditis highlights the epidemiologic and microbiologic features in Brazilian hospitals.In the near future, further investigation will be developed in different hospital centers.
In conclusion, among 73 episodes of IE reported in the last eight years in our hospital, located in the metropolitan area of Rio de Janeiro, S. aureus, Enterococcus spp., and Streptococcus spp.were the main causes of infection.Enterococcus spp. was also identified as an emerging etiology agent of HAIE in our country.

TABLE 1 :
Epidemiology, etiology, and outcomes of infective endocarditis (IE) among inpatients from an urban university hospital.

TABLE 2 :
Echocardiographic, risk factors and clinical findings of 73 cases of infective endocarditis (IE).

TABLE 3 :
Univariate analysis of the risk of mortality in 73 cases with infective endocarditis (IE).

TABLE 4 :
Multivariate analysis of the risk of mortality in 73 patients with infective endocarditis.