Heart Disease and Arboviruses: A Systematic Review and Meta-Analysis

Dengue fever, chikungunya, and zika are highly prevalent arboviruses transmitted by hematophagous arthropods, with a widely neglected impact in developing countries. These diseases cause acute illness in diverse populations, as well as potential cardiovascular complications. A systematic review was carried out to investigate the burden of cardiac involvement related to these arboviruses. Multiple databases were searched for articles that investigated the association of cardiovascular diseases with arboviruses, published up to March 2022. Relevant articles were selected and rated by two independent reviewers. Proportion meta-analysis was applied to assess the frequency-weighted mean of the cardiovascular findings. A total of 42 articles were selected (n = 76,678 individuals), with 17 manuscripts on dengue and 6 manuscripts on chikungunya undergoing meta-analysis. The global pooled incidence of cardiac events in dengue fever using a meta-analysis was 27.21% (95% CI 20.21–34.83; I2 = 94%). The higher incidence of dengue-related myocarditis was found in the population younger than 20 years old (33.85%; 95% CI 0.00–89.20; I2 = 99%). Considering the studies on chikungunya (n = 372), the global pooled incidence of cardiac involvement using a meta-analysis was 32.81% (95% CI 09.58–61.49, I2 = 96%). Two Zika studies were included that examined cases of infection by vertical transmission in Brazil, finding everything from structural changes to changes in heart rate variability that increase the risk of sudden death. In conclusion, cardiac involvement in arboviruses is not uncommon, especially in dengue fever.


Introduction
Dengue fever, chikungunya, and Zika are a group of acute febrile viral diseases transmitted by hematophagous arthropods that have afflicted diverse populations for decades, with the first descriptions of isolated urban outbreaks before 1960 [1,2]. Throughout history, The Zika virus (ZIKV) is a flavivirus, with contemporary outbreaks first recorded in 2007 in Oceania [22]. With the appearance of microcephaly in neonates in 2016 in Brazil, it has become, according to the World Health Organization, a "public health emergency of international concern". By 2019, nearly 90 countries in various world regions had recorded autochthonous transmission of the Zika virus [23]. ZIKV commonly reported symptoms include rash, low grade fever (37.4 • C-38.0 • C), arthralgia, myalgia, fatigue, headache, and conjunctivitis. The most prominent clinical manifestation of ZIKV is microcephaly, a condition defined by an occipital-frontal head circumference (OFD) two standard deviations (SD) smaller than the average expected for age, gender, and population that was recently associated with a prenatal ZIKV infection [24]. Severe neurologic sequelae have also been described in adults, including meningitis, meningoencephalitis, and Guillain-Barre syndrome [25]. In addition, atypical and severe clinical manifestations may occur with myocarditis, pericarditis, heart failure, and arrhythmias such as atrial fibrillation, which are often underdiagnosed and increase mortality, especially in outbreak areas [26].
Despite international collaborative initiatives to contain neglected tropical diseases, there has been an apparent increase in the number of cases worldwide [27]. In Brazil alone, from January to May 2022, dengue, chikungunya, and Zika incidences increased more than 150%, 74%, and 214%, respectively, when compared to the same period in 2021 [28]. The increase in arbovirus infections generates a greater risk of complications or severe disease, especially in older populations and those with comorbidities [29,30]. Although reports exist showing severe cardiac abnormalities related to arbovirus infections, their actual frequency is still poorly known and varies by study [30][31][32]. This study aimed to carry out a systematic review of the literature regarding arbovirus-related cardiac complications, describing cardiac involvement in dengue, chikungunya, and Zika virus infections. In addition, a meta-analysis was performed to describe the incidence and estimate the risk of cardiac involvement in dengue fever.

Search Strategy
The systematic search was conducted in MEDLINE/PubMed, LILACS, Embase, Scopus and Web of Science, using MeSH and Entrees terms for PubMed and Embase, and DeCS (Health Sciences Descriptors) for the other databases. Two independent reviewers (JMN and JRCFR) with expertise in the topic performed the search on 1 March 2022, with the following strategy: (heart OR "heart disease" OR "heart attack" OR "heart failure" OR "cardiovascular disease" OR heart OR cardiovascular OR "cardiovascular disease") AND (arboviruses OR dengue OR denv OR chikungunya OR chikv OR zika OR zikv). The Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement was used to conduct and report this systematic review (PRISMA, 2009). The studies were also selected using StArt (State of the Art through Systematic Review) computer software, version 2.3.4.2, from the Software Engineering Research Laboratory of the Federal University of São Carlos, São Paulo, Brazil (LAPES). This systematic review was registered in the International Prospective Register of Systematic Reviews (PROSPERO) as CRD42020219102.

Study Selection
Distinct study designs (case-control, cohort, prospective, and retrospective studies) involving cardiac complications in dengue, chikungunya, or Zika were selected. Nonoriginal studies, reviews, editorials, abstracts, case reports, and animal model studies were excluded. There was no language limitation.
For study selection, two phases were considered: the first consisted of reading the titles and abstracts. After this phase, the selected full manuscripts were obtained for reading. Two independent reviewers performed both phases. Discrepancies in both phases were resolved through discussion between the reviewers or by a third reviewer (SLPN). The original study investigators were contacted to clarify data when necessary. Any disagreements about study inclusion were resolved by consensus or arbitration by a third reviewer (SLPN).
The methodological quality instrument NOS was applied and cohort studies were classified according to their methodology. Three criteria are included in the NOS checklist: study selection (maximum 4 stars), comparability (maximum 2 stars), and assessment of outcome (maximum 3 stars), with researchers rating each criterion independently [33]. Manuscripts above six stars were considered good quality and low risk of bias; those with six stars were considered moderate quality; and those with five stars or less were considered low quality and, therefore, high risk of bias studies (Supplementary Table S1).

Data Collection and Extraction
After the final selection of the research articles, a form was created during a meeting between the researchers to extract the data from the selected studies for subsequent analysis of the results and discussion. Data were extracted based on patient characteristics, age group, sex, clinical classification of arboviruses, cardiac involvement, cardiac tests performed, comorbidities, study quality, and inclusion and exclusion criteria. The agreement was quantified by the kappa statistical method [34]. Levels of evidence were attributed according to the Oxford Centre for evidence-based medicine [35,36]. The quality of each study was assessed with the Newcastle-Ottawa Scale (NOS) [33]. The Meta-Analyses of Observational Studies in Epidemiology (MOOSE) checklist was applied to assess and reduce bias in the data analysis [35].

Statistical Analysis
Statistical analysis was performed using the software RStudio, version 4.2.0, 2022.02.2+485 (Boston, MA, USA), Open Source License, using the meta package (General Package for Meta-Analysis, 2022). Due to methodological limitations and the low number of included manuscripts, meta-analysis was not performed for the studies on chikungunya and Zika.
We conducted a meta-analysis of proportions to assess the global incidence of cardiac involvement in patients with dengue fever. The results were presented as pooled proportions (%) with a 95% confidence interval (CI). Possible publication bias regarding the incidence of cardiac involvement in patients with arboviruses was evaluated using Begg's and Egger's tests combined with a funnel plot [37].
To prevent misleading conclusions, the Freeman-Tukey double arcsine method was used to stabilize the variance [38]. Heterogeneity and consistency were evaluated using Cochran's Q test and I 2 statistics. The magnitude of heterogeneity was tested by the I 2 method, presented in percentage values of the variance, ranging from 0% to 100%, and observing the actual size effect from all studies considering values above 50% as substantial heterogeneity and values above 70% as high heterogeneity [39,40]. Subgroup meta-analysis was used to assess possible sources of heterogeneity: age group, continent, and study country. Meta-regression, when applicable, was performed using randomeffects analysis on covariates. In these analyses, p-values less than 0.05 were considered statistically significant.

Results
Of the 4209 publications identified using the search strategies described above, 42 articles were selected after meeting the inclusion criteria. The verification of cited articles and citation of included articles yielded nine relevant observational, prospective, and retrospective descriptive studies [41][42][43][44][45][46][47][48][49]. The study selection flowchart for the different phases of the systematic review is displayed in Figure 1.

Study Quality
A total of 52% of the studies (22 studies) presented acceptable methodological quality. Regarding the level of evidence, as these are cohort studies, the level of evidence was considered IIb [34]. Observational prospective studies on dengue that presented appropriate data (similar methodological designs) were included in the meta-analysis. It was impossible to perform a meta-analysis for the studies on Zika. No apparent asymmetry in the funnel plot was observed and the absence of evidence of suspected publication bias was supported by Begg's statistical test (p = 0.423). Asymmetry in the funnel plot was observed by Egger's statistical test (p = 0.001). A funnel plot was generated for the results with data from 34 studies on dengue, applying adjustment by the trim-and-fill method to assess the possible publication bias (Supplementary Figures S1 and S2).

Data Synthesis
A total of 42 articles were selected, resulting in 76,678 individuals exposed to arboviruses; 76,188 were infected by dengue, 372 by chikungunya, and 118 by Zika. The majority of the study participants (51%) were males. Only two manuscripts were published in Spanish, all others in English.
Among the 34 manuscripts on dengue, 32 were prospective studies and 2 were retrospective. As for the six articles on chikungunya, three was prospective and the other three were retrospective. There were only two Zika studies, one retrospective and one prospective. The majority of the studies came from the continent of Asia, with 13 from India, 6 from Thailand, 3 from Sri Lanka, 3 from Taiwan, and 1 from each of the following countries: China, Vietnam, Pakistan, Indonesia. There were also two manuscripts from Colombia, four from Brazil, and one article from each of the following countries: France, Paraguay, Cuba, French Guiana, Puerto Rico, and Guadalupe (Table 1). Of the 42 selected studies, 38 were single-center, with diagnoses by clinical-epidemiological criteria and laboratory confirmation. Of these, five included in their casuistic only severe dengue cases [41,58,61,77,78]. Thirty-eight studies used convenience sampling, two used sample size calculation [44,49], and one used the purposive sampling method [68]. Among the 42 selected studies, 11 included patients under 20 years old and children (1024 participants). Although 65% of the selected studies were published after 2015, most used clinical classification criteria for dengue severity before the 2014 update by the World Health Organization (Table 1).
Only 24% of the studies reported the prevalence of comorbidities and 20% of the studies did not include participants with comorbidities. The remaining 63% studies did not provide information on comorbidities. Regarding the clinical classification of arboviruses, 64% of the studies reported greater cardiac involvement in the more severe forms of the disease and 20% did not provide this information. It is important to note that the selected articles comprised a period from 1973 to 2022, involving different classifications for the severity of arboviruses. Only 8.3% of the manuscripts reported secondary dengue infection, without making clear its potential association with cardiac involvement (Table 1).
Among the papers on dengue, 72% identified electrocardiographic abnormalities in their samples, with sinus bradycardia and tachycardia being the most frequent findings. Left ventricular dysfunction identified by echocardiogram (ejection fraction < 50%) and changes in cardiac biomarkers (CK-MB, troponin, NT-pro BNP) were recorded in 199 and 1066 subjects, respectively.

Studies on Zika
Although there are several articles in the literature that address cardiac involvement related to ZKV (case report, editorial, abstract, cross-sectional study, reviews), we identified two articles in this review that met the inclusion and exclusion criteria up to the time of the search. These were observational studies (retrospective and prospective), evaluating cases of infection by vertical transmission in Brazil with a mean age between 58 days to 16 months. In one of the studies, a retrospective analysis of newborns in northeastern Brazil found that more than 13% of the sample had congenital cardiac abnormalities associated with the Zika virus (ostium secundum, a small apical muscular ventricular septal defect), although they have been little studied. Another prospective study showed a statistically significant difference between R-R values in patients with congenital Zika virus syndrome (24-h Holter monitoring), which may be associated with the risk of sudden infant death syndrome, suggesting early surveillance of these children [79,80]. The lack of more consistent primary studies on this scenario so far compromises the findings on cardiac outcomes in adults and children.

Meta-Analyses of Studies on Chikungunya
Because of the small number of studies identified for chikungunya in this review, we performed meta-analytic estimates and calculated the effect measure (weighted mean incidence) of all studies found, i.e., six studies, three prospective, and three retrospective, with 52% of the sample consisting of men. The global pooled incidence of cardiac events using the meta-analysis of the random-effects model was 32.81% (95% CI 09.58-61.49, I 2 = 96%, p < 0.01, 06 studies, 372 patients) (Figure 2). Subgroup analysis by primary study design showed a pooled incidence of 33.75% for prospective studies and 31.92% for retrospective studies, maintaining high heterogeneity (I 2 = 96%, I 2 = 98%, respectively). The main cardiac manifestation described was cardiovascular failure, which was associated with atypical and severe forms of CHIKV, especially severe sepsis or septic shock, in more than 80% of cases. Regarding myocarditis, the average incidence was low (2.38%; 95% CI 0.00-09.37, I 2 = 88%, p < 0.01, six studies, 372 patients); however, information on the type of cardiac events was lacking in the primary studies analyzed. In addition, comorbidities were reported in more than 90% of cases, with a predominance of hypertension, diabetes mellitus, kidney disease, ischemic heart disease, and chronic heart disease ( Table 1).

Meta-Analyses of Studies on Dengue
Only observational prospective single-center studies on dengue with NOS ≥ 6 were used to perform meta-analytic estimates and to calculate the effect measure (weighted mean incidence), i.e., 17 studies: 13 in patients 20 years of age or older and 4 in patients under 20 years. The global pooled incidence of cardiac events using the meta-analysis of the random-effects model was 27.21% (95% CI 20.21-34.83, I 2 = 94%, p < 0.01, 17 studies, 4616 patients). Subgroup analysis by age showed a pooled incidence of 28.32% for 20 years or older and 24.31% for under 20 years, maintaining high heterogeneity (I 2 = 80%, I 2 = 97%, respectively) ( Figure 3).

Meta-Regression
Due to the high heterogeneity already expected for this meta-analysis of observational studies, a random-effects meta-regression was performed, considering variables of interest (sex, year of study publication, clinical severity of dengue, death associated with cardiac event). The regression model showed a good fit (t 2 = 0.022). The heterogeneous performance accounted for 92.09% of the residual variance, showing that the year of publication of the study was significantly associated with a reduction in cardiac events (p = 0.004; Figure 6). However, there was no association with cardiac outcome according to sex, lethal outcome, and clinical severity (Supplementary Table S2).

Meta-Regression
Due to the high heterogeneity already expected for this meta-analysis of observational studies, a random-effects meta-regression was performed, considering variables of interest (sex, year of study publication, clinical severity of dengue, death associated with cardiac event). The regression model showed a good fit (t 2 = 0.022). The heterogeneous performance accounted for 92.09% of the residual variance, showing that the year of publication of the study was significantly associated with a reduction in cardiac events (p = 0.004; Figure 6). However, there was no association with cardiac outcome according to sex, lethal outcome, and clinical severity (Supplementary Table S2).

Discussion
Arboviruses are associated with acute infections that are increasingly frequent in urban populations, with cyclical outbreaks affecting more vulnerable populations. The presence of the mosquito vector, climatic conditions, and the fragility of public policies favor their dissemination [28]. We showed that cardiac involvement in arboviruses is not a rare complication. Particularly in dengue and chikungunya, the incidence of cardiac complications was found in over a quarter of infected participants. However, it is important to emphasize that this result refers to hospitalized patients with dengue fever or chikungunya and does not apply to the general population exposed to this arbovirus.
Little is known about the pathophysiology of cardiac lesions, however, with regard to dengue, it is related to disease severity and correlates with the extent of plasma leakage [66]. However, other mechanisms have been described, such as tropism of the virus for

Discussion
Arboviruses are associated with acute infections that are increasingly frequent in urban populations, with cyclical outbreaks affecting more vulnerable populations. The presence of the mosquito vector, climatic conditions, and the fragility of public policies favor their dissemination [28]. We showed that cardiac involvement in arboviruses is not a rare complication. Particularly in dengue and chikungunya, the incidence of cardiac complications was found in over a quarter of infected participants. However, it is important to emphasize that this result refers to hospitalized patients with dengue fever or chikungunya and does not apply to the general population exposed to this arbovirus.
Little is known about the pathophysiology of cardiac lesions, however, with regard to dengue, it is related to disease severity and correlates with the extent of plasma leakage [66]. However, other mechanisms have been described, such as tropism of the virus for the myocardium, genetic factors, and a strong host inflammatory response leading to tissue destruction. Similar mechanisms of cardiac tropism have also been described in CHIKV and ZIKV, which trigger an intense inflammatory process with the release of proinflammatory cytokines (IL -18, TNF-α, IFN-γ) and damage cardiac tissue [26,66,79].
In this review, we found a high mean incidence of cardiac events (33%) in chikungunya patients, with more than 90% of these patients having at least one comorbidity (mainly hypertension and diabetes mellitus). Similarly, Alvarez et al., described cardiac involvement in 54.2% of cases in the populations of countries in the Americas, Asia, and Europe [82]. However, in contrast to other studies, we did not find a predominant myocarditis. One of the explanations for this finding would be the heterogeneity of the samples studied and, in particular, the lack of reporting of the nature of cardiac events (e.g., the causes of heart failure were not detailed), so it was not possible to extrapolate this result to the general population.
To justify the involvement of the heart in the context of these arboviruses, there is strong evidence that the tropism of CHIKV applies not only to fibroblasts and interstitial connective tissue but also to several other organs and tissues such as the spleen, skin, lung, bone, liver, and skeletal muscle. In the heart, a CHIKV antigen has been detected in fibroblasts, vascular endothelium, myocardium, and adipose tissue [75].
To date, there is a lack of prospective studies with acceptable methodological quality, especially concerning chikungunya and Zika, which compromises the most consistent epidemiological evaluation. Recent systematic reviews of cardiac complications in patients with dengue, chikungunya, and Zika have included primary studies of poor methodological quality as case reports in 30 to 50% of the sample [26,30,83]. Our review included only prospective and retrospective studies in order to bring more consistency of results.
Based on the methodological proposal of this review, and within the time period that the databases were searched, it was not possible to include studies with cardiac involvement in adults. Nevertheless, the two included articles address an understudied complication (congenital heart defects and changes in heart rate variability) that increases the risk of sudden death in these children. This is particularly important in countries where there is a high risk of an outbreak of this disease, such as Brazil, which increases exposure of pregnant women to ZKV and, consequently, vertical transplacental transmission, which can lead not only to microcephaly but also to cardiac involvement [26,28].
When assessing the pool of dengue-related cardiac abnormalities, myocarditis appears to be the most common potentially harmful finding. Although most of the myocarditis findings are subclinical with enzymatic and electrocardiographic changes, we found that about 5% of those with dengue fever had left ventricular dysfunction. Also associated with low left ventricular fraction, biomarkers (CK-MB, troponin) have been frequently described in observational studies and they are considered to have a poor prognosis when they appear together. In this scenario, recent studies have shown that elevated troponin is more sensitive in identifying minor myocardial injuries [54,66].
Importantly, this review showed that younger people appear to be at a higher risk for dengue-related myocarditis. In general, myocarditis in children has been increasing in recent decades, with peaks in children under 2 years of age and adolescents, accounting for 5% of sudden cardiac deaths in this age group [84][85][86]. This phenomenon, although not well understood, is related to inflammatory factors, with the release of cytokines (TNF-α, interleukins 6, 13, and 18, and cytotoxic factor), greater susceptibility to viral infections with possible tissue infiltration, host genetic factors, and viral load [30,67,81,83]. Despite this, ventricular dysfunction associated with myocarditis and electrocardiographic changes are, in most cases, transient [55,62]. Therefore, it is clear that myocarditis is one of the most important complications in patients with dengue, especially in children and young adults. However, we lack controlled studies to understand the association of dengue and myocarditis better, allowing early diagnosis and treatment of this condition.
When using meta-regression to evaluate possible sources of heterogeneity, we did not observe a significant association of cardiac events with dengue classification. However, case series and observational studies often show this association, probably due to the involvement of an overactive immune response, which can lead to myocarditis with left ventricular dysfunction [62,69]. This lack of association in the meta-regression is probably related to conflicting clinical classifications of these arboviruses over time.
As a less potentially harmful finding, we report bradycardia as the most common arbovirus-related electrocardiographic abnormality. Previous data indicated a predominance of sinus tachycardia, especially in the acute phase [30]. This divergence may be related to the clinical phase of arboviruses, as some of the studies included in this review reported this finding during disease defervescence. The lack of information from the primary studies precluded analyzing this outcome in this meta-analysis.
Despite being well-known diseases, arboviruses challenge the health of populations mainly in metropolitan and peri-urban areas of tropical countries, due to reinfestation of their main vector (Aedes aegypti), causing increasingly severe clinical repercussions, as is the case of Zika in the Americas. In addition, these arboviruses (mainly dengue and chikungunya) have been reaching other ecological areas, such as temperate climate countries, through the uncontrolled expansion of another important vector (Aedes albopictus), generating large outbreaks and a sustained cycle of diseases, with serious health repercussions [87]. Therefore, a proper vector control policy is fundamental to minimize the impacts of this disease on populations.
Another important strategy to control symptomatic disease and to reduce its complications is to promote the training of healthcare teams for early detection and for monitoring of signs and symptoms. No less important, especially in endemic areas at a higher risk of outbreaks, is the development of vaccines. Although some vaccines are already commercially launched vaccines, their effectiveness is still considered low. Global collaboration is therefore needed to develop more effective vaccines, as was the case, for example, with COVID-19 [88].
This review has limitations due to the scarcity of primary studies with better methodological qualities, compromising the consistency of the effect size. There were information gaps, especially regarding comorbidities and secondary infections, in addition to the heterogeneity of the populations studied. Despite these limitations, this review brings an analysis of prospective primary studies with a better methodological quality, confirming that, through the estimation of grouped frequency, the cardiac involvement in arboviruses are not uncommon manifestations, being necessary preventive actions.

Conclusions
In conclusion, cardiac involvement is not uncommon as a complication of arboviruses, with higher quantity and quality data published for dengue fever. Myocarditis was the most frequent and potentially harmful cardiac complication in dengue fever, with indicatives of a higher burden in youth.

Supplementary Materials:
The following supporting information can be downloaded at: https: //www.mdpi.com/article/10.3390/v14091988/s1, Figure S1: funnel plot for evaluation of publication bias with pseudo 95% confidence limits (Begg's test); Figure S2: funnel plot with pseudo 95% confidence limits after trim fill adjustment (Egger's test); Figure S3: forest plot comparison of frequency of cardiac events in patients with dengue, grouped by country group; Table S1: methodological quality assessment by the NOS Score; Table S2: meta-regression of the variables gender and clinical severity of dengue in cardiac outcome. Institutional Review Board Statement: Ethical review and approval were waived for this study because it was a systematic review of previously published primary studies.
Informed Consent Statement: Patient consent was waived because this was a systematic review of previously published studies.

Data Availability Statement:
The data supporting the reported data can be accessed via the link https://github.com/jdinicacio/arbovirusis.git.

Conflicts of Interest:
The authors declare no conflict of interest.