The Application of the Proposed Sydney System for Reporting Lymph Node Cytopathology: A Five-Year Experience of an Academic Institution in South India

Fine needle aspiration biopsy (FNAB) is one of the initial diagnostic tests in the evaluation of lymphadenopathy. It provides an immediate, cost-effective, and preliminary alternative to surgical biopsy. The role of FNAB varies at the primary or community level and secondary or specialist level. While at the community level, the aim of FNAB is to triage the cases of lymphadenopathy into those with a low or a high index of suspicion of significant disease, at the specialist level, its role is to arrive at a specific diagnosis with the use of ancillary studies [1]. The diagnosis and subcategorization of lymphomas rely on histopathological and immunohistochemical diagnoses. A wide variety of benign and malignant lesions can occur in lymph nodes. The clinical and imaging details are crucial to cytopathologists in arriving at a diagnosis [2, 3]. Nonetheless, there is a lack of uniformity in reporting lymph node cytopathology. The Bethesda system for thyroid cytopathology, the Milan system for reporting salivary gland cytopathology, and the Yokohama system for reporting breast cytopathology have been validated to be useful and are in routine practice in many institutions. In line with these, the Sydney system was proposed for the reporting of lymph node cytopathology [4]. There are five diagnostic categories, namely, L1 to L5, denoting nondiagnostic, benign, atypical cells of undetermined significance, suspicious of malignancy, and malignant, respectively. It also provides recommendations for management in each category. There are studies in the literature that had analyzed the utility of this system in routine reporting [5‒10]. The present study aimed to analyze the diagnostic utility of the proposed Sydney system for reporting lymph node cytopathology, establish the histopathological correlation of each category, and highlight the cytological features of rare and atypical lymphoid lesions.

This was a retrospective study conducted at an academic institute in South India. The study was reviewed and approved by the Institutional Human Ethics Committee of PSG IMSR (approval number PSG/IHEC/2022/Appr/Exp/132). All the FNAB aspirations from the lymph nodal masses performed during the study period of 5 years between January 2017 and December 2021 were included in the study.

Palpation-guided and ultrasound-guided FNAB, endobronchial ultrasound-guided transbronchial FNAB samples were included. FNAB was performed by pathologists or radiologists using 22–24-gauge needles. Onsite assessment of cellularity was performed by the cytotechnician or the pathologist in cases of palpation-guided FNAB and EBUS-guided FNAB.

The smears were stained by May-Grunwald-Giemsa, hematoxylin and eosin, and Papanicolaou stain. Ziehl-Neelsen staining was performed for all the lymph nodal aspirates.

Cell blocks were prepared from all the EBUS-guided FNAB samples, and immunohistochemistry was performed wherever necessary. The antibodies used were TTF1, napsin, CK5/6, p40, and synaptophysin. The clinico-demographic details of these patients including age, sex, nature of lymphadenopathy, and sonographic findings were collected from the hospital information system.

The cytopathology smears were retrieved and reviewed. The cases were reclassified according to the proposed Sydney system for reporting lymph node pathology as follows.

• L1; inadequate/nondiagnostic

• L2; benign

• L3; atypical cells of undetermined significance/atypical lymphoid cells of uncertain significance (AUS/ALUS)

• L4; suspicious

• L5; malignant.

The probable diagnostic possibilities were given for categories L4 and L5 wherever possible. The histopathology reports were retrieved from the laboratory information system. The cytopathological diagnoses were then correlated with histopathological diagnoses. The rate of malignancy (ROM) in each category was calculated based on the final histopathological diagnosis. For calculating the ROM, category L3 was considered benign. Atypical lymphoid hyperplasia (ALH) was not considered in the calculation of ROM.

Cross-tabulation of the cytopathological and histopathological diagnoses was done and sensitivity, specificity, and positive and negative predictive values of FNAB were calculated. Discordant cases were reviewed and analyzed to ascertain the probable cause for the error in cytopathology.

Data entry was made in Microsoft Excel version 16.49. Statistical analysis was performed using IBM SPSS Statistics for Windows, version 20.0 NY.

ROM was calculated based on the histopathological diagnosis. Sensitivity, specificity, and positive and negative predictive values were calculated in two scenarios: one considering L3 as benign and the other considering L3 as malignant.

There were a total of 747 fine needle aspirations from the lymph nodes during the study period. The majority of the aspirations were from peripheral lymph nodes, performed percutaneously or under ultrasound guidance. EBUS-guided lymph node aspirations constituted 12.58%.

The distribution of cytological categories with diagnostic subcategories is outlined in Table 1. There were 79 (10.57%) cases of inadequate aspirates categorized as L1. The majority of these aspirates (n = 20) were nonrepresentative samples from EBUS-guided FNA.

The benign category (L1) was the most frequent constituting 54.6% (n = 408). Reactive lymphoid hyperplasia (n = 197, 48.2%) was the most common interpretation, followed by granulomatous (necrotizing/non-necrotizing) inflammation (n = 175, 42.9%), acute suppurative inflammation (n = 30, 7.35%), and histiocytosis (n = 6, 1.47%). 25 out of 175 cases of granulomatous inflammation were positive for acid-fast bacilli in cytopathology and confirmed as tuberculosis.

Category AUS/ALUS, L3, included 30 (4%) cases. Among these, 19 (63.3%) cases were reported as atypical lymphoid cells and 10 (33.3%) constituted atypical non-lymphoid cells. 28 (3.7%) cases were rendered a cytopathological diagnosis of suspicious for malignancy, category L4.

202 cases were categorized as L5. Among these, 181 (89.6%) cases were reported as metastatic malignancies and 21 (10.4%) cases were reported as lymphomas. Most lymphoid malignancies were non-Hodgkin lymphoma (NHL) (n = 20, 80.95%) followed by Hodgkin lymphoma (HL) (n = 1, 19.04%).

Among the metastatic malignancies (n = 181), the most frequent was metastatic carcinoma (n = 78, 43.09%) followed by squamous cell carcinoma (n = 48, 26.51%) and adenocarcinoma (n = 46, 25.41%). Histopathological samples were available for 289 (38.68%) cases. Of these, 27 were inadequate in histopathology. Hence, 262 cases with histopathological correlation were included in the calculation of ROM. The histopathological diagnoses of these cases and the calculated ROM are summarized in Table 2 and 3, respectively.

Histopathological diagnosis was available for 19 out of 79 cases in L1. Five of these cases turned out to be lymphomas (3 cases of NHL, 2 cases of HL) with an overall ROM of 26.3%.

In category L2, 125 cases had a histopathological follow-up. Of these, nine cases were rendered a malignant diagnosis (6 cases of lymphomas, 1 case each of carcinoma, melanoma, and follicular dendritic cell sarcoma [FDCS]). The calculated ROM in L2 was 7.2%.

In category L3, there were 13 cases where subsequent histopathology was available. Of these, 10 cases were reported as malignant (6 cases of metastatic malignancy, 2 cases of HL, 1 case each of NHL and plasmacytoma) accounting for an ROM of 76.9%. 14 out of 17 cases in L4 that had histopathological follow-up were malignant for an overall ROM of 82.35%. Three cases were rendered a benign diagnosis.

In category L5, 88 of 202 cases had a histopathological follow-up with all the cases proven malignant on histopathology. The calculated ROM in L5 was 100%. Two cases of NHL were rendered a cytological diagnosis of poorly differentiated carcinoma. One case of metastatic paraganglioma was reported as metastatic adenocarcinoma in cytopathology.

Immunocytochemistry on cell block was performed on 23 cases of EBUS-guided FNAB. There were 16 cases of adenocarcinoma, 4 cases of squamous cell carcinoma, and a single case each of adenosquamous and small cell carcinoma.

The concordance between malignant cytopathology (L4 and L5) and histology was 97.1% The sensitivity, specificity, positive predictive value, and negative predictive value of FNAB when considering category L3 to represent benign cytology were 84.2%, 97.5%, 97.1%, and 86.2%, respectively. The sensitivity, specificity, positive predictive value, and negative predictive value of FNAB when considering category L3 to represent malignant cytology were 92.56%, 95.08%, 94.9%, and 92.8%, respectively.

The review of cytopathological smears was performed for the discrepant cases, and the review results are tabulated in Table 4. There were three cases of false positives in category L4 and nine cases of false negatives in category L2. Smears that were given a category of L1 on cytopathology were not reviewed. Cases that were rendered a cytopathological interpretation of L3 were reviewed but not included in the false-negative/false-positive cases.

There were no discordant cases in category L5 as all the cases that had histopathological follow-up were reported as malignant in cytopathology. However, the subtyping of malignancy was wrong in seven of these cases. Three cases that were diagnosed as NHL on cytopathology were poorly differentiated carcinomas on histopathology. Two cases that were given a diagnosis of carcinoma on cytopathology were reported as NHL on histopathology. One case of HL was reported as T cell-rich diffuse large B-cell lymphoma in histopathology. Another case interpreted as metastatic adenocarcinoma on cytopathology was diagnosed as metastatic paraganglioma.

FNAB continues to be a cost-effective first-line investigation in the evaluation of peripheral lymphadenopathies. With the myriad of diagnostic possibilities, cytopathological evaluation is extremely challenging [11, 12]. However, there exists a variation in the nomenclature used in the reporting of lymph node FNAB since there is a lack of guidelines and a reporting system. Similar to that proposed for reporting thyroid and breast cytopathology, which are proven to be a standardized reporting system, the Sydney system has been proposed to limit interobserver variability, enhance reproducibility, and enable effective communication of clinically relevant information with possible management recommendations [4].

It is vital to evaluate the application of the proposed system in the risk stratification and calculation of the ROM. The present study validates the utility of the proposed Sydney system for reporting lymph node FNAB in our population. The distribution of the diagnostic categories in cytopathology was comparable to other studies [5‒10].

The L1 category constituted 10.5%. This rate could be lowered by rapid onsite evaluation (ROSE) and guided aspirations. A significant proportion of these cases were EBUS-guided FNAB, which was inadequate despite rapid onsite evaluation. In such a scenario, where repeat FNAB is neither beneficial nor feasible, either core needle or excision biopsy is to be considered as proposed by the Sydney system in the management recommendation of L1 category lesions. In the other cases of superficial lymphadenopathies, the quality of the aspirate is largely dependent on the skill of the aspirator. The limitation of having different expertise is unavoidable in a teaching institution. Therefore, the repeat FNAB if mandated should be performed by an experienced aspirator to maximize the diagnostic yield.

The most frequent interpretation in category L2 was reactive lymphoid hyperplasia followed by granulomatous lymphadenitis (GLN), which is similar to that reported in other studies [5‒9]. There was a single case of IgG4-related lymphadenopathy on histopathology, which was rendered a diagnosis of GLN in cytopathology, Figure 1. Kaur et al. [13] in their analysis of cytopathological features of IgG4-related disease reported low cellularity, inflammatory background, conspicuous plasma cells, and eosinophils as the diagnostic clues to the disease. However, in the present case, there were no conspicuous plasma cells on the slide review. The granulomas were also observed in the histopathological sections. Granulomas, though unusual in the setting of IgG4-related lymphadenitis have been reported previously [14].

The frequency of category L3 ranges from 0.8 to 8.3% [5, 7]. The current study reports a rate of 4%. The lower rate could be explained by the departmental protocol to obtain intradepartmental consultation before assigning an intermediate category. ALUS cases showed a heterogeneous population of cells with absent tingible body macrophages that could represent paracortical hyperplasia or NHL. AUS was used when there are scattered large atypical non-lymphoid cells. Atypical lymphoid cells were more common than non-lymphoid cells, which is comparable to the published studies [5‒10]. However, when considering the histopathological correlation, metastatic deposits were more frequent than lymphomas. When L3 category lesions are considered to represent malignancy, there were three discordant cases, of which one was inconclusive and the other two were reactive. Paracortical hyperplasia, a potential mimicker of low-grade NHL, has been reported previously [1].

In category L4, suspicious for lymphoid neoplasms (n = 21, 75%) were more frequent than non-lymphoid neoplasms, which contrasts with that reported by Gupta et al. (48.9%) [7]. In category L5, metastatic malignancies were more frequent, which is similar to that observed by Gupta et al. [7]. Poorly differentiated carcinoma was the most common interpretation followed by SCC and adenocarcinoma, while SCC was the most common in the study by Gupta et al. [7].

The calculated ROM was the least for L2 and the highest for L5. The study reports an ROM of 26.3% for L1, which is similar to that reported by Gupta et al. [7]. Five out of 19 cases that were reported as malignant showed predominantly necrosis and degenerated cells. ROSE by cytopathologists could have minimized the frequency of inadequate aspirates and thereby reduce the false negatives. ROSE has also been recommended by the proposed Sydney system to enhance yield and diagnostic accuracy. The higher ROM observed in the present study could be explained by the fact that those cases that are clinically suspicious were biopsied. Cases that fall under the L1 category should be followed up by a repeat FNAB or biopsy, especially when there is extensive necrosis.

Out of 125 cases with histopathological diagnosis in category L2, nine were found to be malignant. NHL (n = 4) was the most common false-negative diagnosis, which included 1 case each of diffuse large B-cell lymphoma and anaplastic large cell lymphoma and 2 cases of follicular lymphoma. The corresponding cytology smears of these cases were reported as reactive lymphadenitis (n = 3) and necrotizing GLN (n = 1). On review, three of these cases showed scattered large, atypical cells, one of which was pauci-cellular with a predominance of large cells. The remaining case did not show any large cells.

Two cases were diagnosed as HL in histopathology, Figure 2. The corresponding cytopathology diagnoses were reactive lymphadenitis and acute suppurative inflammation. A careful review of the cytopathology smears showed scattered mononuclear Reed-Sternberg (RS) cells in a rich polymorphous background that was poor in eosinophils.

The remaining two false-negative cases of carcinoma and melanoma were reported as reactive lymphoid hyperplasia. The review of these smears did not reveal any evidence of atypical cells. The histopathology also revealed partial effacement of these lymph nodes by metastasis. This could represent a sampling error.

The common causes for misinterpretation in the cases of false-negative diagnoses were scant cellularity, polymorphous lymphoid background, eosinophil poor reactive background, presence of scattered atypical cells, mononuclear RS cells, and paucity of classic RS cells. The RS cells being overlooked and masked by the dominance of polymorphous infiltrates has been reported previously [14]. Other studies have reported that the predominance of neutrophils in a necrotic background in rare cases of HL can mimic suppurative lymphadenitis, as observed in our case [15‒17]. Previous studies have also highlighted the difficulty in diagnosing lymphomas in cytopathology, especially in the presence of a reactive background [15‒17].

There was a single case of FDCS which was rendered a diagnosis of granulomatous inflammation in cytopathology, Figure 3. The smears reviewed showed loosely cohesive clusters of epithelioid cells resembling granulomas admixed with small lymphocytes, but nuclear grooves and prominent nucleoli were observed in some of the clusters. Previous studies have highlighted the difficulty in the accurate identification of FDCS in cytopathology [18]. These cases have overlapping features with entities like granulomas, undifferentiated carcinomas, and other mesenchymal neoplasms. The diagnosis of FDCS was made on histopathology based on S100 and CD23 expression in immunohistochemistry.

There were ten cases of malignancy reported in L3, accounting for an ROM of 76.9% which is higher than that reported in previous studies. These included 6 cases of metastatic carcinomas, 2 cases of NHL, and 1 case each of HL and plasma cell myeloma. On review of smears, scattered large, atypical cells were observed in 3 out of the 6 cases of metastasis, whereas no atypical cells were identified in the remaining 2 cases. The corresponding histopathology of these lymph nodes showed partial involvement by metastasis. The two cases of NHL showed features of paracortical hyperplasia, a mimicker of low-grade lymphoma which has been reported previously [1].

Three false-positive cases were observed in category L4. These cases were reported as suspicious of NHL; however, they were diagnosed as an autoimmune lymphoproliferative syndrome (ALPS) (n = 1), ALH (n = 1), and GLN (n = 1) on subsequent histopathology.

The cytopathological features in ALPS and ALH showed sheets of large cells with prominent nucleoli, scanty reactive cells, and an absence of tingible body macrophages. These cases were challenging diagnoses in histopathology (Fig. 4-6). The diagnosis was made only after correlating with serology, IHC, other ancillary studies, and clinical findings. Infectious mononucleosis, as a potential mimicker of lymphoma, has been reported previously [1]. The diagnosis of ALPS was made after the demonstration of double-negative T cells by flow cytometry. AHLs as observed in the cases of infectious mononucleosis and ALPS are the causes of false positives in cytology. However, these cases require histopathological diagnosis with ancillary techniques and hence excision is mandated [19].

The sensitivity and negative predictive value of FNAB is high when the L3 category is considered to represent malignant lesions. The atypical category L3 reduces the false-negative rates.

There was a single discordant case in L5. This was a case of metastatic paraganglioma, wherein the high cellularity with acinar formations of the oval to round pleomorphic cells in a hemorrhagic background was misinterpreted as metastatic carcinoma, Figure 7. A few studies have highlighted the difficulty in distinguishing retroperitoneal paraganglioma from adrenal cortical carcinomas, renal cell carcinoma, and metastatic adenocarcinoma [20]. The sensitivity and specificity of FNAB of lymph nodes fall well within that reported by other studies which vary from 75% to 99% and 87% to 100%, respectively, Table 5.

The limitations of the study include its retrospective design and lack of histopathological correlation in all cases. However, the present study discusses a variety of lymph node pathologies.

The application of the newly proposed Sydney system for reporting lymph node cytopathology will aid in better communication between pathologists and clinicians. The specificity and positive predictive value of FNAB are high when category L3 (atypical) is considered negative for malignancy.

The authors believe that the application of the newly proposed Sydney system for reporting lymph node cytopathology will aid in better communication between pathologists and clinicians. The use of uniform terminology will also enhance better reproducibility in cytopathological diagnosis. The fact that FNAB is still the primary modality of investigation in the evaluation of lymph nodes in rural settings needs to be acknowledged. Though there was a good correlation between malignant cytopathology and histopathology, further studies are needed to ascertain the cytopathological categorization of rare lesions, thereby aiding in better risk stratification.

The study was approved by the Institutional Human Ethics Committee PSG IMSR (approval number PSG/IHEC/2022/Appr/Exp/132). A waiver of consent was obtained from the Institutional Human Ethics Committee of PSG IMSR since it is a retrospective study conducted on archived slides.

The authors have no conflicts of interest to declare.

No source of funding was applied in this retrospective study.

Sakthisankari Shanmugasundaram and Nandhini Bala Balasubramanian: conceptualization, data curation, formal analysis, software, and writing – original draft, writing – review and editing. Abinaya Sundari Amirthakatesan: methodology, resources, analysis, and writing – review and editing. Sakthisankari Shanmugasundaram and Nandhini Bala Balasubramanian should be considered the joint first authors.

All data generated or analyzed during this study are included in this article. Further inquiries can be directed to the corresponding author.