Prevalence and morphology of middle mesial canals in mandibular first molars and their relationship with anatomical aspects of the mesial root: a CBCT analysis
The mandibular first molar (M1M) is the first permanent tooth to erupt in the oral cavity. Characterized by its complex anatomical morphology and relatively weak self-cleaning function, it is highly susceptible to pulp diseases and frequently needs root canal treatment (RCT) . In an ideal RCT procedure, root canals should be thoroughly cleaned and tightly filled. However, the omission of root canals is often a significant factor contributing to periapical inflammation post-RCT . This omission is typically attributed to the clinician’s inadequate understanding of the root canal system’s anatomy, as well as insufficient comprehensive preoperative examination. The typical anatomical structure of the M1M consists of two roots, with three root canals: the mesiobuccal canal, the mesiolingual canal, and the distal canal [ – ]. The root canal system of M1M is complex, with its morphology often exhibiting variability, making it prone to being overlooked by clinicians . One such variation is the presence of a middle mesial root canal (MMC). As early as 1981, Pomeranz HH et al. identified 12 cases of MMC among 100 treated mandibular molars and classified their morphologies into three types: fin, confluent, and independent . The reported prevalence of MMC in the literature varies widely, ranging from 0.26–53.8% [ – ]. This discrepancy may stem from differences in ethnicity or the methodologies adopted in the respective studies. In recent years, Cone Beam Computed Tomography (CBCT) has been widely used in the field of endodontics. In contrast to the limited two-dimensional images provided by periapical radiography, CBCT facilitates detailed three-dimensional visualization of anatomical structures in the horizontal, sagittal, and coronal planes . La et al. first pointed out in their case report that the presence of MMC can be accurately diagnosed utilizing CBCT . Under the premise of rational use, CBCT will serve as an important examination that can help clinicians make correct diagnosis and decisions at critical moments . The presence of the MMC holds significant clinical importance, and clinicians must be aware of this variation in order to thoroughly clean and seal all the canals during treatment, as missed canals have a notable correlation with the presence of apical periodontitis . It is noteworthy that the prevalence of MMC varies across different ethnicities, ages, and genders, and as a result, clinicians need to consider different factors when treating different populations . The prevalence of MMCs in M1Ms is a well-studied topic with multiple cross-sectional and meta-analyses exploring its anatomic correlations [ – ]. While this subject is not entirely novel, our study aims to contribute to the existing knowledge by delving deeper correlations, and provide further insights into the complex morphology of MMCs. Thus, the objectives of this study were to: (1) investigate the prevalence of MMC in M1M in northwestern China, based on CBCT image analysis; (2) evaluate the morphological characteristics of MMC and record their first appearance below the canal orifice; (3) ascertain whether the presence of MMCs is correlated with the distance between the mesiobuccal (MB) and mesiolingual (ML) canals, as well as with anatomical parameters of the mesial roots. 

This study was approved by the Medical Ethics Committee of Shaanxi Provincial People’s Hospital (No. SPPH-LLBG-17-3.2). Sample preparation The sample size in this study was calculated using the formula \documentclass[12pt]{minimal}
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				\begin{document}$$\:n=\frac{{z}_{\alpha\:/2}\:p(1-p)}{{\delta\:}^{2}}$$\end{document} , with a bilateral 95% confidence interval. The p -value was set at 7%, which was derived from a recently published cross-sectional meta-analysis . Utilizing these parameters, the calculated sample size is 107. In this study, CBCT data were selected from patients who visited the Department of Stomatology, Shaanxi Provincial People’s Hospital, between January 2021 and December 2022. These patients underwent CBCT scans due to the need for orthodontic treatment, tooth extraction, restoration, and other therapeutic purposes. All patients provided informed consent for the CBCT scan. A total of 449 patients with 898 mandibular first molars were included. The inclusion criteria were as follows: (1) the presence of bilateral M1Ms; (2) patients aged 18 years or older; (3) CBCT images being clear and free from artifacts. The following conditions were excluded: the affected tooth having undergone root canal treatment, post-and-core, or crown restoration; having immature apices; exhibiting periapical lesions; tooth deformity; severe calcification of the pulp cavity; root resorption; and root fracture. CBCT analysis CBCT scanning of the oral and maxillofacial region of the patient was performed using the Largev Smart3D (China), with a voxel size of 0.25 mm, a resolution of 2.0Ip/mm, a scanning time of 12.5s, an operating voltage of 100 kV, and a field of view (FOV) of 15 mm x 9 mm. Prevalence of middle mesial canals The software Smart V2 (Largev, version 2.0.9.4539, China) was used for the reconstruction of the CBCT images. Subsequently, the left and right M1Ms were observed from the axial, sagittal, and coronal planes by one attending physician (Wang DD) with 6 years of experience to evaluate the presence of MMCs. Meanwhile, the patient’s name, age, gender, and dental position were recorded. Two weeks after the evaluation, 20% of the samples were reassessed by the same physician under identical conditions to verify the intra-examiner consistency. Finally, the prevalence of MMC in the northwestern Chinese population was calculated. Morphological analysis and measurement Following image evaluation, a total of 65 M1Ms were identified to contain MMCs. Subsequently, the morphology of the MMCs was analyzed, and the vertical distance D, measured from the point where the MMCs first appeared to the root canal orifice, was recorded (Fig. A). The 65 teeth with MMCs constituted the experimental group. To establish a control group, we carefully matched each of these 65 teeth with a corresponding tooth from a different patient that did not have an MMC. Matching was based on gender, age (basically the same, with any difference not exceeding 5 years), and tooth position to ensure comparability. The following parameters were measured: (1) the distance between MB and ML root canal orifices (D1) (Fig. B); (2) the buccolingual width (L1) and mesiodistal width (L2) of the mesial roots (Fig. C); and (3) the degree of flattening of the mesial root, expressed as the ratio α = L1/L2. An analysis was conducted to determine whether these parameters differed significantly between the experimental and control groups. Statistical analysis Data management was conducted using Excel (Microsoft, Redmond, WA), statistical analysis was performed using SAS 9.4 (SAS Institute, Inc., Cary, NC, USA), and scatter plots were generated using GraphPad Prism 8. Quantitative data were described using mean ± standard deviation ( \documentclass[12pt]{minimal}
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				\begin{document}$$\:\stackrel{-}{x}\pm\:s$$\end{document} ), and comparisons between groups were conducted using paired t-test. Qualitative data were reported in the form of counts (percentages) and compared between groups, such as gender and age, using the χ² test. It should be noted that in this study, age was categorized into two groups: ≤40 years and > 40 years. The kappa test is used to assess the intra-examiner agreement of prevalence, and the intraclass correlation coefficient (ICC) is used to assess the measurements. Unless otherwise specified, the significance level α was set at 0.05. 

The sample size in this study was calculated using the formula \documentclass[12pt]{minimal}
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				\begin{document}$$\:n=\frac{{z}_{\alpha\:/2}\:p(1-p)}{{\delta\:}^{2}}$$\end{document} , with a bilateral 95% confidence interval. The p -value was set at 7%, which was derived from a recently published cross-sectional meta-analysis . Utilizing these parameters, the calculated sample size is 107. In this study, CBCT data were selected from patients who visited the Department of Stomatology, Shaanxi Provincial People’s Hospital, between January 2021 and December 2022. These patients underwent CBCT scans due to the need for orthodontic treatment, tooth extraction, restoration, and other therapeutic purposes. All patients provided informed consent for the CBCT scan. A total of 449 patients with 898 mandibular first molars were included. The inclusion criteria were as follows: (1) the presence of bilateral M1Ms; (2) patients aged 18 years or older; (3) CBCT images being clear and free from artifacts. The following conditions were excluded: the affected tooth having undergone root canal treatment, post-and-core, or crown restoration; having immature apices; exhibiting periapical lesions; tooth deformity; severe calcification of the pulp cavity; root resorption; and root fracture. 

CBCT scanning of the oral and maxillofacial region of the patient was performed using the Largev Smart3D (China), with a voxel size of 0.25 mm, a resolution of 2.0Ip/mm, a scanning time of 12.5s, an operating voltage of 100 kV, and a field of view (FOV) of 15 mm x 9 mm. 

The software Smart V2 (Largev, version 2.0.9.4539, China) was used for the reconstruction of the CBCT images. Subsequently, the left and right M1Ms were observed from the axial, sagittal, and coronal planes by one attending physician (Wang DD) with 6 years of experience to evaluate the presence of MMCs. Meanwhile, the patient’s name, age, gender, and dental position were recorded. Two weeks after the evaluation, 20% of the samples were reassessed by the same physician under identical conditions to verify the intra-examiner consistency. Finally, the prevalence of MMC in the northwestern Chinese population was calculated. 

Following image evaluation, a total of 65 M1Ms were identified to contain MMCs. Subsequently, the morphology of the MMCs was analyzed, and the vertical distance D, measured from the point where the MMCs first appeared to the root canal orifice, was recorded (Fig. A). The 65 teeth with MMCs constituted the experimental group. To establish a control group, we carefully matched each of these 65 teeth with a corresponding tooth from a different patient that did not have an MMC. Matching was based on gender, age (basically the same, with any difference not exceeding 5 years), and tooth position to ensure comparability. The following parameters were measured: (1) the distance between MB and ML root canal orifices (D1) (Fig. B); (2) the buccolingual width (L1) and mesiodistal width (L2) of the mesial roots (Fig. C); and (3) the degree of flattening of the mesial root, expressed as the ratio α = L1/L2. An analysis was conducted to determine whether these parameters differed significantly between the experimental and control groups. 

Data management was conducted using Excel (Microsoft, Redmond, WA), statistical analysis was performed using SAS 9.4 (SAS Institute, Inc., Cary, NC, USA), and scatter plots were generated using GraphPad Prism 8. Quantitative data were described using mean ± standard deviation ( \documentclass[12pt]{minimal}
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				\begin{document}$$\:\stackrel{-}{x}\pm\:s$$\end{document} ), and comparisons between groups were conducted using paired t-test. Qualitative data were reported in the form of counts (percentages) and compared between groups, such as gender and age, using the χ² test. It should be noted that in this study, age was categorized into two groups: ≤40 years and > 40 years. The kappa test is used to assess the intra-examiner agreement of prevalence, and the intraclass correlation coefficient (ICC) is used to assess the measurements. Unless otherwise specified, the significance level α was set at 0.05. 

A total of 898 M1Ms from 449 patients were analyzed in the study. The intra-examiner consistency was high ( k = 0.81, ICC = 0.91, P <0.05). Among these, 65 M1Ms from 43 patients exhibited MMCs. The mean age of the patients was 34.89 ± 13.18 years (ranging from 18 to 94). The prevalence of MMC was 9.6% based on the number of individuals and 7.2% based on the number of teeth. No statistically significant differences were observed in the prevalence of MMC between genders or across age groups ( P > 0.05), as presented in Table . In this study, we summarized and classified the root canal morphology associated with MMC anomalies, identifying 19 distinct types (Fig. ). The most common configurations were 1-3-2, 2-3-2, and 2-3-2-1, with only one case exhibiting an entirely independent morphology. Illustrative examples of some of the typical cases are presented in Fig. . The first appearance of MMC was found to be 1.99 ± 1.60 mm below the canal orifice, and its distribution was shown in Fig. , which indicates that MMC mainly appeared within 4 mm below the canal orifice. After matching, both the experimental and control groups contained 65 M1Ms. A paired t-test was subsequently performed to compare the groups. The results revealed a statistically significant difference in the MB-ML distance (D1) between the groups, whereas no statistically significant differences were observed in the buccolingual width (L1), mesiodistal width (L2), or the flatness degree (α) of the mesial root ( P > 0.05). These findings are summarized in Table . 

The middle mesial canal (MMC) is a variation found in mandibular molars, and its prevalence varies among different populations, exhibiting a wide range in the literature. For instance, Barros-Costa M et al. reported a prevalence of 11.1% in a Brazilian population , Tahmasbi et al. found a rate of 26% in an American subpopulation , and Liu J et al. documented a rate of 15.2% in a northern Chinese population . In a transnational cross-sectional meta-analysis conducted in 2023, the worldwide prevalence of MMC in mandibular first molars was approximately 7% . Another global meta-analysis yielded a prevalence of 4.15% . All of these conclusions were drawn from observations using CBCT. Recently, Nosrat A. et al. conducted a large-scale clinical study on mandibular molars and reported a prevalence of 8.8% . The present study included 449 patients and 898 M1Ms from northwest China, with a prevalence of 9.6% based on the number of patients and 7.2% based on the number of teeth. These findings were also analyzed through CBCT, and the results fall within the aforementioned range. Pertek Hatipoğlu F noted that variations in MMC prevalence are not only related to ethnicity but also influenced by factors such as sample size, CBCT parameters, inter-examiner differences, and interference of isthmus . In this study, there was no significant difference in the prevalence of MMC between genders, consistent with the results reported in previous extensive studies [ – , − ]. This underscores the importance of conducting a thorough examination for the presence of MMCs in the M1Ms of both males and females during treatment. Similarly, in this study, there was no significant difference in the prevalence of MMC across age groups, consistent with some previous research findings [ – , − ]. However, some studies have indicated that the prevalence of MMC is higher in younger patients and decreases with age , which may be related to secondary dentin deposition. Conversely, other studies have reported that the prevalence of MMC peaks in the 41–60 age group , possibly due to dentin formation at the isthmus between the MB and ML root canals, allowing previously connected root canals to be separated by dentin, thereby forming another root canal. The relationship between the prevalence of MMC and age remains controversial and requires further investigations. Professor Pomeranz initially proposed three classifications of MMC morphology based on apical radiographs. Compared with CBCT, two-dimensional images have certain limitations in some cases . Barros-Costa M et al. used CBCT to analyze the morphology of MMC and divided it into four types: independent, confluent, mesiolingual confluent, and mesiobuccal confluent. They pointed out that the prevalence of the independent type was the highest, accounting for 60.7% . However, this is inconsistent with the results of a recent systematic review, which indicated that the most common form of MMC is the confluent type, which is more frequently merged with MB canals . Versiani MA et al. employed microCT to observe 48 M1Ms with MMC and also noted that the most common morphology is the confluent type. Furthermore, 3D images in this literature demonstrated that the morphology of MMC is significantly diverse . Among the 65 teeth in this study, 19 different root canal types were identified, with the most common type being 1-3-2, and only one independent type was found. The varying results of studies on MMC morphology may be related to racial differences and require further investigation. In clinical practice, it is essential to conduct a thorough interpretation of radiological images. to ensure that the diameter and taper of root canal preparation can be rationally decided based on the morphology of MMC . In addition, this study measured that the first appearance of MMC was 1.99 ± 1.60 mm below the canal orifice, and MMC mainly appeared within 4 mm beneath the orifice. This is basically consistent with the research findings reported by Alroomy R et al. . However, there is little literature reporting this measurement, and more data are expected to be reported in the future. During clinical procedures aimed at identifying the MMCs, practitioners are advised to thoroughly inspect the area situated approximately 3 to 4 millimeters beneath the canal orifice. Regarding the relationship between the presence of MMC and the MB-ML distance, there remains controversy in the literature. Akbarzadeh et al. evaluated 210 CBCT data and found that the MB-ML distance was negatively correlated with the presence of MMC . Conversely, studies by Alroomy R, Barros-Costa M, et al. have shown that the average distance between MB and ML is greater when MMC is present . Our study also concurs with this finding, demonstrating that the MB-ML distance is larger at 3.84 mm when MMC is present, compared to 3.62 mm when it is absent. Although we cannot determine a specific MB-ML distance that definitively indicates the presence of MMC, we can infer that a larger space between MB and ML is necessary to accommodate MMC. In clinical practice, a distance close to 4 mm can serve as a reference point to raise awareness of the potential presence of MMC. With regard to the relationship between the presence of MMC and the buccolingual width, mesiodistal width, as well as the degree of flattening of the mesial root, there are only a handful of related studies. One study has pointed out that teeth with MMC exhibit a larger buccolingual distance (8.79 mm compared to 8.44 mm), whereas no significant difference is observed in the mesiodistal distance . However, our study failed to uncover a significant correlation between these factors. We argue that when detecting MMC, greater emphasis should be placed on the distance between MB and ML, rather than focusing on the anatomical morphology of the mesial root. Nevertheless, this issue necessitates further research to provide definitive proof. This study has several limitations. First, the 250 μm voxel size used in the CBCT device may reduce sensitivity in detecting fine anatomical structures like the MMC; literature suggests that voxel sizes of 200 μm or smaller yield more accurate results [ – ]. The study sample comprises a relatively young population with an average age of 34.89 years, which limits the assessment of potential differences in MMC prevalence across age groups. Secondary dentin deposition is a well-documented phenomenon, potentially leading to the occlusion or disappearance of MMCs. In this study, there is a large variability in age, ranging from a minimum of 18 to a maximum of 94, which may affect the results. Including only patients requiring orthodontic or restorative treatment may reduce the generalizability of the results to the general population. Additionally, using only CBCT means that the advantages of higher-resolution techniques, such as micro-CT, could not be leveraged. The assessments being conducted by a single observer may lead to a lack of inter- and intra-observer reliability, which could impact reproducibility. Finally, as the study is based on a Northwest Chinese population, its generalizability regarding MMC prevalence to other ethnic groups or geographic regions is limited. 

In summary, the complexity of MMC morphology, with prevalent confluent canals, necessitates careful assessment during endodontic treatments. To effectively detect MMC, it is advisable to carefully examine the area within 4 mm below the canal orifice. In instances where the MB-ML distance is notably large, it is recommended to exercise meticulousness during the detection process to ensure the assessment of MMC. 

Below is the link to the electronic supplementary material. Supplementary Material 1 

