European Journal of Cardiovascular Medicine

Myocarditis is defined as an inflammatory disease of the myocardium diagnosed by established histological, immunological, and immunohistochemical criteria [1]. Clinically, it represents a heterogeneous disorder with presentations ranging from subclinical disease to chest pain mimicking acute coronary syndrome, progressive heart failure, malignant arrhythmias, or sudden cardiac death [2,3]. The true incidence of myocarditis is difficult to determine because of diagnostic limitations, but autopsy series suggest it contributes to up to 12% of sudden deaths in young adults [4].

Endomyocardial biopsy (EMB) remains the diagnostic gold standard for myocarditis, providing the opportunity to evaluate tissue-level changes, identify histopathological subtypes, and perform viral and immunohistochemical testing [5,6]. The classical Dallas criteria, published in 1986, defined myocarditis as “the presence of an inflammatory infiltrate of the myocardium with associated myocyte necrosis not typical of ischemic damage” [7]. Although revolutionary at the time, the Dallas criteria were limited by inter-observer variability and insensitivity, as they did not incorporate molecular virology or immunohistochemistry [8]. Since then, refinements have been introduced, including immunohistochemical quantification of inflammatory cells and routine polymerase chain reaction (PCR)-based viral genome detection, which together have enhanced diagnostic yield [9,10].

Histopathologically, myocarditis exhibits diverse patterns that provide important diagnostic clues. Lymphocytic myocarditis is the most prevalent subtype, characterized by a predominance of T-lymphocyte infiltration and focal myocyte necrosis [11]. Eosinophilic myocarditis is typically associated with drug hypersensitivity, parasitic infection, or hypereosinophilic syndromes, and is defined by eosinophil-rich infiltrates [12]. Giant cell myocarditis is rare but fulminant, marked by widespread necrosis and multinucleated giant cells, and carries a grave prognosis if untreated [13]. Granulomatous myocarditis often reflects cardiac sarcoidosis or infectious granulomas such as tuberculosis, and is characterized by well-formed granulomas with or without necrosis [14]. Each histological subtype has distinct prognostic and therapeutic implications, highlighting the clinical importance of accurate classification.

Among etiologies, viral infection is recognized as the most common cause of myocarditis worldwide [15,16]. The spectrum of implicated viruses has shifted over time. In the 1980s and 1990s, enteroviruses (especially coxsackievirus B) and adenoviruses predominated [17]. Since the early 2000s, parvovirus B19 has emerged as the leading agent in Europe and North America, while human herpesvirus-6 (HHV-6), Epstein-Barr virus (EBV), and cytomegalovirus (CMV) have been increasingly detected [18,19]. Regional variation exists, with hepatitis C virus implicated in Japan and Chagas disease caused by Trypanosoma cruzi prevalent in South America [20]. Viral detection is usually performed by PCR, which is more sensitive than immunohistochemistry or in situ hybridization [21]. Importantly, the presence of viral genomes does not always prove causation, as latent infection or bystander viral persistence can confound interpretation [22].

The relationship between histopathological subtype and viral etiology remains incompletely understood. Lymphocytic myocarditis is strongly suspected to be viral in origin, particularly linked with parvovirus B19 and enteroviruses [23]. In contrast, eosinophilic and giant cell myocarditis are thought to be largely non-viral, often autoimmune or hypersensitivity-related [24]. Granulomatous myocarditis is typically associated with sarcoidosis or tuberculosis rather than viral infections [25]. However, individual studies have yielded inconsistent findings, with variable viral prevalence across histological patterns, reflecting differences in biopsy timing, sampling, and viral detection methods [26].

Understanding these associations has significant clinical implications. If specific histopathological patterns predict viral or non-viral etiologies, clinicians could better tailor diagnostic algorithms and therapeutic decisions. For instance, detection of viral genomes in lymphocytic myocarditis may support antiviral or immunomodulatory approaches, whereas identifying eosinophilic or giant cell myocarditis should prompt evaluation for non-viral triggers and consideration of immunosuppressive therapy [27,28].

Despite increasing recognition of this relationship, no prior synthesis has comprehensively quantified the strength of association between histopathological subtypes and viral detection in myocardial tissue.

Therefore, the objective of this systematic review and meta-analysis was to evaluate histopathological patterns of myocarditis and determine their association with viral etiologies detected by tissue-based molecular or immunohistochemical methods. By integrating evidence from multiple studies, this review aims to clarify diagnostic correlations, highlight regional and methodological differences, and provide guidance for future research and clinical management.

This systematic review and meta-analysis was conducted in accordance with the PRISMA 2020 guidelines [1].

Eligibility criteria- We included studies that met the following criteria: (i) original human research (cohort, case-control, cross-sectional, or case series with ≥5 patients); (ii) histopathological confirmation of myocarditis by endomyocardial biopsy (EMB) or autopsy; (iii) classification of myocarditis into at least one histological subtype (lymphocytic, eosinophilic, giant cell, granulomatous, or mixed); and (iv) myocardial viral testing by polymerase chain reaction (PCR), in situ hybridization, immunohistochemistry, or viral culture. Studies without histopathological data, without viral testing, or reporting insufficient outcomes were excluded. Non-English publications, single case reports, reviews, and animal studies were also excluded.

Search strategy- A comprehensive electronic search was conducted in PubMed/MEDLINE, Embase, Scopus, and Web of Science from database inception to March 31, 2025. The search combined controlled vocabulary (e.g., MeSH and tiab) and free-text terms related to myocarditis, histopathology, and viral etiologies. The PubMed strategy was:

Equivalent strategies were developed for Embase, Scopus, and Web of Science, with syntax adapted for each database. Search results were supplemented by manual screening of reference lists of included studies and relevant reviews. Conference abstracts were considered if they provided extractable data. Only English-language articles were included.

Study selection

All retrieved records were imported into EndNote X9 for de-duplication. Two reviewers independently screened titles and abstracts, followed by full-text review of potentially eligible articles. Discrepancies were resolved through discussion or adjudication by a third reviewer. A total of 2,418 records were identified across all databases (PubMed/MEDLINE, Embase, Scopus, and Web of Science). After removal of 612 duplicates, 1,806 records remained for screening. Following title and abstract screening, 1,521 records were excluded for not meeting inclusion criteria (irrelevant topic, no histopathology, no viral testing, or non-human studies). The full texts of 285 articles were assessed for eligibility, of which 253 were excluded (reasons: no extractable data [n=108], case report/small case series [n=72], review or editorial [n=41], non-English [n=21], and duplicate population [n=11]). Ultimately, 32 studies met eligibility criteria and were included in the qualitative synthesis, all of which provided extractable data for quantitative meta-analysis. The selection process is summarized in a PRISMA flow diagram (Figure 1).

Figure 1: Study selection Process PRISMA flow diagram

Data extraction

Data were independently extracted by two reviewers using a standardized form, including: study characteristics (author, year, country, design, sample size), patient demographics, type of biopsy (EMB or autopsy), histopathological classification, viral detection method, virus type identified, and clinical outcomes (e.g., mortality, heart failure, transplant). Disagreements were resolved by consensus.

Quality assessment

Risk of bias was assessed independently by two reviewers. The Newcastle-Ottawa Scale (NOS) was applied for cohort and case-control studies [2], while the NIH quality assessment tool was used for cross-sectional studies. For diagnostic accuracy studies, the QUADAS-2 tool was applied [3]. Studies were classified as low, moderate, or high risk of bias.

Data synthesis and statistical analysis

The primary outcome was the association between histological subtype and viral detection in myocardial tissue. For each study, 2×2 contingency tables were constructed (histological pattern present/absent vs viral detection positive/negative). Odds ratios (ORs) with 95% confidence intervals (CI) were calculated and pooled using a random-effects model (DerSimonian-Laird). Heterogeneity was assessed using the I² statistic (low <25%, moderate 25-50%, high >50%) [4]. Subgroup analyses were performed by virus type (parvovirus B19, enterovirus, adenovirus, HHV-6), detection method (PCR vs non-PCR), and biopsy type (EMB vs autopsy). Sensitivity analyses excluded high-risk-of-bias studies. Publication bias was evaluated using funnel plots and Egger’s regression test [5].

Statistical analyses were performed in R (metafor package) and Stata version 17. A p-value <0.05 was considered statistically significant.

A total of 32 studies were eligible for inclusion, comprising 4,256 patients with histologically confirmed myocarditis (Figure 1).

Study characteristics

The included studies were published between 1986 and 2024 and originated from Europe, North America, Asia, and South America, providing broad geographic representation [1-4]. Most were prospective or retrospective cohort studies, while five were cross-sectional and three were large case series. Endomyocardial biopsy (EMB) was the tissue source in 26 studies, while 6 utilized autopsy material. The mean age of patients was 42.1 years, and 64% were male. Viral detection was performed by PCR in 29 studies, immunohistochemistry in 11, and in situ hybridization in 6; several studies applied multiple methods. Follow-up periods ranged from acute hospitalization to over 5 years [5-8].

Distribution of histopathological patterns

Lymphocytic myocarditis was the most frequent pattern, observed in 61.8% of cases (n = 2,628). Eosinophilic myocarditis accounted for 9.2% (n = 392), granulomatous myocarditis for 6.5% (n = 276), and giant cell myocarditis for 3.8% (n = 161). Mixed or unclassified patterns comprised the remaining 18.7% (n = 799). Lymphocytic myocarditis predominated across all regions and study designs, though eosinophilic myocarditis was proportionally higher in autopsy series, consistent with fulminant or fatal presentations [9-11].

Viral detection

Overall, 53.4% (n = 2,268) of tissue samples tested positive for viral genomes or antigens. Parvovirus B19 was the most frequently detected virus, representing 41.2% of positive samples, followed by enteroviruses (28.5%), adenoviruses (12.6%), human herpesvirus-6 (7.9%), and others such as EBV, CMV, and influenza (9.8%) [12-15]. Viral detection rates were higher in autopsy series (61.2%) than in EMB studies (51.1%), likely reflecting the more extensive sampling available in autopsies [16].

Association between histopathological patterns and viral detection

Meta-analysis demonstrated a strong positive association between lymphocytic myocarditis and viral detection, with a pooled odds ratio (OR) of 3.12 (95% CI 2.25-4.32, I² = 28%) [17]. By contrast, eosinophilic myocarditis (OR 0.54, 95% CI 0.33-0.88, I² = 12%) and giant cell myocarditis (OR 0.41, 95% CI 0.19-0.88, I² = 0%) were inversely associated with viral positivity, indicating alternative etiologies [18,19]. Granulomatous myocarditis showed no significant association (OR 0.92, 95% CI 0.55-1.56, I² = 35%), consistent with its established links to sarcoidosis and tuberculosis [20].

Subgroup and sensitivity analyses

Subgroup analyses revealed that parvovirus B19 was most strongly linked with lymphocytic myocarditis (OR 3.78, 95% CI 2.33-6.12), while enteroviruses also showed significant association but with greater inter-study heterogeneity [21]. Viral detection rates were higher when PCR was used compared with immunohistochemistry or in situ hybridization. Sensitivity analyses excluding high-risk-of-bias studies yielded results consistent with the main analysis. Funnel plots were symmetrical, and Egger’s test did not suggest publication bias (p = 0.21) [22].

This systematic review and meta-analysis demonstrate that histopathological subtypes of myocarditis are closely linked with underlying etiologies, particularly viral infections. Across 32 included studies involving more than 4,000 patients, lymphocytic myocarditis emerged as the predominant subtype and was strongly associated with the detection of viral genomes, most notably parvovirus B19 and enteroviruses. In contrast, eosinophilic and giant cell myocarditis were rarely associated with viral detection, supporting the notion that these entities are largely autoimmune or hypersensitivity-mediated. Granulomatous myocarditis, commonly reflecting sarcoidosis or tuberculosis, showed no significant correlation with viral presence. These findings confirm the long-held suspicion that histological patterns carry important etiological implications and provide a foundation for integrating pathology with molecular virology in the diagnosis of myocarditis.

The predominance of viral genomes in lymphocytic myocarditis is consistent with previous mechanistic studies. Viruses such as enteroviruses and adenoviruses directly infect cardiomyocytes, leading to cytolysis and the recruitment of T lymphocytes, which produce the typical lymphocytic infiltrate and myocyte necrosis that define the condition [1,2]. Parvovirus B19, which has become the most frequently detected virus in Western cohorts, demonstrates endothelial tropism, causing microvascular dysfunction and ischemia that can mimic ischemic cardiomyopathy but is characterized histologically by lymphocyte-rich infiltrates [3,4]. These mechanisms explain the strong association observed between lymphocytic histology and viral detection. In contrast, eosinophilic myocarditis arises from immune-mediated hypersensitivity, often triggered by drugs, autoimmune disease, or parasitic infection, with tissue damage caused by the release of cytotoxic eosinophilic granules such as major basic protein and eosinophil cationic protein, independent of viral replication [5,6]. Similarly, giant cell myocarditis, a fulminant and often fatal entity, is driven by autoreactive T lymphocytes targeting cardiac antigens and is strongly associated with systemic autoimmune disorders rather than viral infection [7,8]. Granulomatous myocarditis, on the other hand, reflects delayed-type hypersensitivity reactions and is most often linked to systemic sarcoidosis or infectious agents such as Mycobacterium tuberculosis, again consistent with our finding of no significant viral association [9].

Our results are also in line with prior clinical observations and autopsy studies. In the 1980s and 1990s, enteroviruses and adenoviruses dominated as the major viral causes of myocarditis, but since the early 2000s, parvovirus B19 has emerged as the leading detected agent in both acute and chronic cases [10,11]. Human herpesvirus-6 and Epstein-Barr virus have also been implicated, though their pathogenic significance remains debated, as they may represent latent infections rather than causative pathogens [12]. Eosinophilic myocarditis has long been linked to drug-induced hypersensitivity reactions, particularly antibiotics, antipsychotics, and anti-inflammatory agents, and is only rarely viral in origin [13]. Giant cell myocarditis has been repeatedly reported in association with autoimmune thyroid disease, thymoma, and other autoimmune syndromes, further supporting a non-viral pathogenesis [14]. Granulomatous myocarditis has historically been equated with cardiac sarcoidosis in most Western cohorts, while in tuberculosis-endemic regions it is often due to Mycobacterium tuberculosis, a pattern that explains the absence of viral association [15,16].

Although the findings of this review reinforce biological plausibility, some limitations must be acknowledged. Histopathological classification is subject to inter-observer variability, especially when relying on the Dallas criteria, which are known to have limited sensitivity and reproducibility [17]. Sampling error also poses a challenge, as myocarditis is frequently patchy and EMB specimens may miss diagnostic foci [18]. Viral detection techniques varied across studies, with PCR generally more sensitive than immunohistochemistry or in situ hybridization, but PCR also risks detecting bystander viral genomes without active replication, raising the possibility of overestimating viral prevalence [19]. Furthermore, the timing of biopsy relative to symptom onset was inconsistently reported, yet it is well known that viral genomes may persist long after acute infection, complicating interpretation of causality [20]. Finally, most studies were observational and single-center, with heterogeneous patient populations, limiting the ability to control for confounding factors such as immunosuppressive therapy or comorbid conditions.

Despite these limitations, the clinical implications are significant. In patients with lymphocytic myocarditis, viral testing should be prioritized, as detection of a causative virus has prognostic implications and may influence therapeutic choices [21]. For example, persistent parvovirus B19 replication has been linked with adverse outcomes and progression to dilated cardiomyopathy [22]. Conversely, the absence of viral genomes in eosinophilic and giant cell myocarditis suggests that antiviral therapy would be ineffective, and instead, prompt initiation of immunosuppressive regimens may improve survival [23,24]. For granulomatous myocarditis, the findings highlight the importance of systemic evaluation for sarcoidosis or tuberculosis rather than focusing on viral diagnostics [25].

The integration of histopathology with molecular virology thus represents a key step toward precision medicine in myocarditis. Viral-positive lymphocytic myocarditis often portends worse outcomes and may benefit from antiviral or immunomodulatory strategies, while viral-negative immune-mediated forms may respond favorably to corticosteroids or other immunosuppressants [26]. Moving forward, prospective multicenter studies should employ standardized histological definitions and comprehensive viral panels to validate these associations. Advanced approaches such as RNA sequencing, viral transcriptomics, and spatial transcriptomics could further distinguish active viral replication from latent persistence and clarify causal mechanisms [27]. Randomized clinical trials stratified by histological and virological findings are also urgently needed to translate these diagnostic insights into evidence-based therapeutic strategies [28].

In inference, this systematic review establishes that histopathological patterns of myocarditis are strongly associated with underlying etiologies. Lymphocytic myocarditis is closely linked to viral infections, especially parvovirus B19 and enteroviruses, while eosinophilic and giant cell myocarditis are predominantly non-viral and granulomatous myocarditis reflects systemic immune or infectious disorders. These findings emphasize the importance of combining histopathological interpretation with molecular diagnostics to improve accuracy, refine prognosis, and guide therapy in patients with myocarditis.

Histopathological patterns of myocarditis are strongly associated with etiology. Lymphocytic myocarditis correlates with viral presence, especially parvovirus B19 and enteroviruses, whereas eosinophilic and giant cell myocarditis are typically non-viral. Integrating histology with viral diagnostics improves diagnostic accuracy and guides’ therapy.

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