A Comparative Study of Fine-Needle Aspiration and Nonaspiration Cytology Diagnosis in Thyroid Lesions.

BACKGROUND: Aspiration cytology is one of the first-line diagnostic tests in thyroid malignancies. Fine-needle aspiration cytology (FNAC) in thyroid lesions causes hemorrhagic smear and cell trauma, often leading to the repetition of smear and delay in diagnosis. This study was conducted to identify the diagnostically superior technique with regard to thyroid swelling and to assess the quality of smears obtained from FNAC and fine-needle nonaspiration cytology (FNNAC).
METHODOLOGY: This was a prospective diagnostic study carried out for 2 years in a tertiary care center from South India. All patients with complaints of thyroid swellings, after examination, underwent FNNAC, followed by FNAC of the lesion. They underwent thyroidectomy when indicated. The final postoperative biopsy reports were compared with the preoperative reports of these two techniques (FNNAC and FNAC). The quality of smears was compared using Mair's score.
RESULTS: The sensitivity, specificity, positive predictive value, negative predictive value, and accuracy in diagnosing malignancy were 93.4%, 100%, 100%, 98.78%, and 98.96% for FNNAC and 94.12%, 100%, 100%, 98.82%, and 99% for FNAC, respectively, which were comparable. Regarding the quality of smears, FNNAC had more smears with less blood in the background. FNAC had more smears with adequate cellularity. The difference in overall Mair's score between the two techniques was not significant (P = 0.28).
CONCLUSION: No difference was found in the accuracy of FNAC and FNNAC in diagnosing thyroid lesions. Furthermore, the smear quality of both techniques was comparable. Hence, either can be used based on the operator's preference and experience. Copyright:

INTRODUCTION

Fine-needle aspiration cytology (FNAC) is one of the first-line diagnostic tests in the evaluation of thyroid malignancies.[1] Application of suction in a highly vascular organ like thyroid leads to an increase in hemorrhagic smears and an increase in cell trauma.[2] Hence, it often results in nondiagnostic or indeterminate smears, leading to the repetition of smears, thus causing a delay in diagnosis and treatment. To overcome these difficulties, fine-needle nonaspiration cytology (FNNAC) was introduced, wherein suction pressure was not applied. Capillary action draws the cells into the hub of the needle, which is then smeared onto a slide and studied. Many studies were conducted comparing both techniques in various organs. Few studies favored FNNAC, and few studies concluded that the techniques were comparable.[123] This study was done to compare the efficacy of FNAC and FNNAC with respect to thyroid malignancies and also the quality of the smears.

METHODOLOGY

This was a prospective study done in a tertiary care center from South India for 2 years. The Institute Human Ethics Committee approval was obtained. All patients presenting to the general surgery outpatient department with complaints of thyroid swelling and age more than 18 years were included in the study after obtaining written consent. A thorough history was taken, and physical examination was done. Thyroid function tests and an ultrasound scan of the neck were done, followed by the cytodiagnostic study. If the patient had hyperthyroidism, FNAC and FNNAC were done after the treatment of hyperthyroidism.

The participants were made to lie supine with a wedge placed between the shoulder blades to extend the neck. The thyroid region was cleaned with spirit and let to air dry. The swelling was fixed with left hand and FNNAC was performed with a 23 G needle held between the forefinger and thumb of the right hand. The needle was inserted and multiple passages in different direction were done. The materials collected in the hub of the needle were expressed onto a slide and immediately fixed with Giemsa and Pap stain. FNAC was then performed similarly except that suction was applied with a 10-ml syringe attached to the needle on entering the swelling. Nonaspiration cytology was always performed first, followed by aspiration cytology. The slides were randomly labeled as A and B to blind the interpreting cytologist. The patients were worked up by the corresponding units in surgery and underwent thyroidectomy when indicated. The patients who did not undergo thyroidectomy were excluded from the analysis of the comparison of the accuracy of the techniques. However, they were included for comparison of Mair's scoring of the two techniques. The final histopathological reports of the operated patients were collated.

The smears were studied by the cytotechnologist and investigator. Cytotechnologists performed an initial and primary evaluation, and then, all the slides were referred to the investigator (cytopathologist) for final interpretation.

Based on Mair's total scores, the specimen was categorized as follows [Table 1]:[4]

Table 1

Mair’s scoring system to classify the quality of cytological aspirate

Criteria	Description	Score
Background blood or clot	Large amount Compromises diagnosis	0
	Moderate amount Diagnosis possible	1
	Minimal amount Diagnosis easy	2
Amount of cellular material	Minimal to absent Diagnosis not possible	0
	Moderate Sufficient for diagnosis	1
	Abundant Diagnosis of simple	2
Degree of cellular degeneration	Marked Diagnosis impossible	0
	Moderate Diagnosis possible	1
	Minimal Diagnosis of easy	2
Degree of cellular trauma	Marked Diagnosis not possible	0
	Moderate Diagnosis possible	1
	Minimal Diagnosis obvious	2
Retention of appropriate architecture	Minimal Nondiagnostic	0
	Moderate Some preservation of cell pattern	1
	Excellent Diagnosis obvious	2

Unsuitable for diagnosis (0–2)

Diagnostically adequate (3–6)

Diagnostically superior (7–10).

In the end, the following was noted.

Diagnosis of both the A and B slides based on the Bethesda category

Quality of the A and B slides based on Mair's scoring [Figures 1 and 2].

Figure 1

Fine-needle aspiration cytology smear from a case of conventional papillary thyroid carcinoma showing the well-preserved diagnostic papillary fragment with fibrovascular core (black arrowhead) (Pap, ×400)

Figure 2

Nonaspiration smear of the same case showing classic intranuclear cytoplasmic inclusions, with well-preserved morphology (black arrowhead) (Pap, ×400)

All the slides were reviewed again by the cytopathologist, and the reports were correlated with the final histopathological report.

Statistical analysis

The data were entered in Redcap (version 6.11.1) and exported in Excel sheet, and SPSS software (IBM Corp. Released 2016.IBM SPSS Statistics for Windows, Version 24.0. Armonk, NY: IBM Corp) was used for analyzing the data. FNAC and FNNAC reports were compared with the histopathological diagnosis to determine the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) in diagnosing malignancy. The quality of slides of both techniques was compared using Mair's scoring. Categorical variables such as FNNAC reports, FNAC reports, and Mair's scoring were calculated in terms of percentages. Pearson's Chi-square test was used to determine the association of Mair's scoring with histopathological findings. Statistical significance was set at P < 0.05.

RESULTS

A total of 125 patients who presented with thyroid swelling underwent both FNNAC and FNAC diagnostic tests. Among them, 104 patients were operated. Both FNAC and FNNAC details were taken into account for the diagnostic accuracy in malignancy specimens. Results of FNAC, FNNAC, and final biopsy reports were compared among the study patients [Table 2].

Table 2

Results of fine-needle aspiration cytology, fine-needle nonaspiration cytology, and final biopsy report among study patients

Bethesda system		FNNAC		FNAC
Category	Cytological diagnosis	Number of cases (n=125), n (%)	Operated cases with similar final report (n=104), n (%)	Number of cases (n=125), n (%)	Operated cases with similar final report (n=104), n (%)
1	Nondiagnostic/unsatisfactory	3	2 (1.9)	1	1 (0.9)
2	Benign-colloid/nodular goiter/lymphocytic/Hashimoto thyroiditis	100	82 (78.8)	103	85 (81.73)
3	Atypia/follicular lesion of undetermined significance	2	0	3	0
4	Follicular neoplasm	5	5 (4.8)	2	2 (1.9)
5	Suspicious for malignancy	5	5 (4.8)	6	6 (5.76)
6	Papillary carcinoma	8	8 (7.6)	8	8 (7.6)
	Medullary carcinoma	2	2 (1.9)	2	2 (1.9)

FNNAC: Fine-needle nonaspiration cytology, FNAC: Fine-needle aspiration cytology

All 125 cases who had both FNAC and FNNAC slides were included for comparison of Mair's scoring [Table 3].

Table 3

Results showing the diagnostic accuracy of both final needle cytology tests

Type	Sensitivity (%)	Specificity (%)	PPV (%)	NPV (%)	Accuracy (%)	FP (%)	FN (%)
FNNAC	93.75	100	100	98.78	98.96	0	1
FNAC	94.12	100	100	98.82	99	0	0.9

FNNAC: Fine-needle nonaspiration cytology, FNAC: Fine-needle aspiration cytology, PPV: Positive predictive value, NPV: Negative predictive value, FP: False positive, FN: False negative

Precise diagnosis of the nature of the lesion

FNAC and FNNAC reports were assessed in terms of the possibility of a precise diagnosis of the nature of the lesion. In this assessment, nondiagnostic and follicular neoplasm cases were taken as the absence of a precise diagnosis of the nature of the lesion. This was because the Bethesda-4 lesion does not explicitly confirm that such lesions are benign or malignant. Nondiagnostic cases required repeat cytology, as no opinion was possible on the same. Benign and malignant reports of FNNAC and FNAC were taken as a precise diagnosis of the nature of the lesion. Among FNNAC reports, seven reports (6.7%) did not give a precise diagnosis of the nature of the lesion. Among FNAC reports, in three cases (2.8%), a precise diagnosis of the nature of the lesion was not obtained.

Mair's scoring

All the 125 slides prepared using both the techniques were compared in terms of each component of Mair's score and categorized [Tables 4 and 5].

Table 4

Mair’s score in fine-needle nonaspiration cytology and fine-needle aspiration cytology in study patients

Mair’s score	Background blood or clot, n (%)		Cellular material, n (%)		Cellular degeneration, n (%)		Cellular trauma, n (%)		Retention of appropriate architecture, n (%)
	FNNAC	FNAC	FNNAC	FNAC	FNNAC	FNAC	FNNAC	FNAC	FNNAC	FNAC
Score 0	3 (2.4)	1 (0.8)	3 (2.4)	1 (0.8)	3 (2.4)	1 (0.8)	3 (2.4)	1 (0.8)	3 (2.4)	1 (0.8)
Score 1	17 (13.6)	39 (31.2)	46 (36.8)	19 (15.2)	12 (9.6)	9 (7.2)	22 (17.6)	21 (16.8)	0	0
Score 2	105 (84)	85 (68)	76 (60.8)	105 (84)	110 (88)	115 (92)	100 (80)	103 (82.4)	122 (97.6)	124 (99.2)
P	0.003		<0.001		0.4		0.627		0.622

FNNAC: Fine-needle nonaspiration cytology, FNAC: Fine-needle aspiration cytology

Table 5

Mair’s category in fine-needle nonaspiration cytology and fine-needle aspiration cytology in study patients

Mair’s category	FNNAC (%)	FNAC (%)
Unsuitable for diagnosis (0-2)	3 (2.4)	1 (0.8)
Diagnostically adequate (3-6)	2 (1.6)	1 (0.8)
Diagnostically superior (7-10)	120 (96)	123 (98.4)

FNNAC: Fine-needle nonaspiration cytology, FNAC: Fine-needle aspiration cytology

The amount of background blood or clots was significantly less in FNNAC than FNAC. The amount of cellular material was significantly more in FNAC than FNNAC. Among other parameters, no significant difference was found. This study had used a scoring method which was purely subjective analysis.

DISCUSSION

In the present study, FNAC diagnosed more benign cases. Follicular neoplasm and nondiagnostic cases were more in FNNAC. Compared to nonaspiration cytology, follicular neoplasm and nondiagnostic cases were less in aspiration cytology (4.8% and 1.9% vs. 1.9% and 0.9%). This is similar to findings observed in studies conducted by Sinna and Ezzat, Kasper et al., and Hirachand et al.[123] In a study conducted by Carvalho et al., they found that FNAC and FNNAC provided similar cytological diagnosis, respectively (malignant: 3.8% vs. 3.8%, P = 0.871; suspicious: 10.4% vs. 10.8%, P = 0.913; and nondiagnostic: 10.0% vs. 11.2%, P = 0.598).[5] However, as in our study, the follicular neoplasm and nondiagnostic cases were more in nonaspiration cytology than in aspiration cytology. In a study conducted by Hirachand et al., the nondiagnostic smears were more (2.6%) in FNAC, but the percentage of colloid goiter (72.53%) and papillary carcinoma (4.76%) was similar to this study.[3]

The most common diagnosis in aspiration and nonaspiration cytology was colloid and nodular goiter (82% and 77%), respectively. The most common malignancy was papillary carcinoma. This is similar to the study conducted by Hirachand et al., according to which the most common diagnosis in thyroid lesions was colloid goiter (72.53%) and the most common malignancy was papillary carcinoma (4.76%).[3]

Nondiagnostic cases

Among the nondiagnostic cases in FNNAC, one was benign and another was malignant by FNAC. Nondiagnostic smear could be due to sampling from a nonrepresentative area, or it could also result from a poor technique. To reduce the incidence of sampling from the nonrepresentative area, the technique could have been done under ultrasound (USG) guidance. If we had performed only FNNAC, a malignancy case could have been missed, or the surgery plan would have been delayed, as we had to repeat FNAC. In FNAC, one case was nondiagnostic which was follicular neoplasm by FNNAC. This was due to increased blood in the smear, which obscured the diagnosis.

Follicular neoplasm cases

There were five cases diagnosed as follicular neoplasm by FNNAC. Among them, three cases were multinodular goiter, one was follicular adenoma, and one was follicular carcinoma by histology. Follicular carcinoma could not be diagnosed by cytology, as it needs the presence of capsular and vascular invasion, which can only be demonstrated in thyroidectomy specimens. Differentiating colloid goiter and follicular neoplasm cytologically is often difficult. As we aspirate from the hypercellular areas at times, it leads to an overestimation of the diagnosis.[1] Again, this can be corrected by doing a USG-guided aspiration. Furthermore, multiple punctures can be done from various parts of the swelling. Aspiration cytology picks up colloid goiter more than nonaspiration cytology, as aspiration helps in sucking in colloid, which gets smeared liberally on the slide and helps in diagnosis. In FNAC, two cases were follicular neoplasm and the corresponding FNNAC reports were the same.

False-positive and false-negative cases

The false-positive (FP) rate in the present study is 0%. Similar FP rates were obtained in the meta-analysis conducted by Gharib and Goellner which pooled data from seven series of similar studies in thyroid lesions.[6] In a study conducted by Sinna and Ezzat and Thanigaimai et al., the false positive rate for aspiration cytology in thyroid lesions (7.2% and 66.6%) was higher than the present study.[17] Zero FP was achieved because all the slides were reviewed by the experienced investigator and as such malignancy cases were also very less in this study.

The false-negative (FN) rate indicated benign cytology, which turns out to be malignant on histopathology. There was one FN case in both techniques. The FN rate was 1% in FNNAC and 0.9% in FNAC. In the case of lymphocytic thyroiditis, postsurgery was found to have lymphoma. The error was due to missing of lymphomatous focus while sampling. Again, a USG-guided FNAC/FNNAC would have helped to prevent the FN rate. Furthermore, in cases where there is suspicion of lymphoma, flow cytometry on the smears would have helped in diagnosis, without resorting to surgery.

Similar rates were obtained for aspiration cytology in thyroid lesions in studies conducted by Gharib and Goellner and Sinna and Ezzat.[16] In contrast to this study, the study conducted by Thanigaimai et al. found that the FN rate of aspiration cytology in thyroid lesions was higher than in the present study.[6] From the present study, it can be concluded that FP and FN rates were comparable in aspiration and nonaspiration cytology.

Sensitivity and specificity

In a few studies like Sinna and Ezzat, indeterminate (Bethesda 3 and 4) and unsatisfactory smears were not included in the calculation of the efficacy of the technique.[1] However, in the study conducted by Kasper et al., smears with indeterminate cells were included in the benign category and smears with suspicious for malignancy were included in the malignant category.[2] This leads to an underestimation of the accuracy of the technique. In the present study, follicular neoplasm and nondiagnostic cases were excluded from analysis for estimation of accuracy of the technique.

In this study, the specificity and PPV of FNNAC and FNAC are comparable (100%). The sensitivity and NPV varied marginally. A similar result was obtained in a study conducted by Rodrigues and Sindhu, wherein no difference was found in the diagnostic accuracy of the two techniques in thyroid lesions.[8]

This is, in contrast, to study conducted by Carvalho et al., comparing FNAC and FNNAC among thyroid lesion in 58 patients who underwent surgery.[5] They found that the sensitivity of FNAC was less than FNNAC. However, there was no statistically significant difference, and they concluded that either test could be used based on the operator's preference and experience. In contrast to present study, Mahajan and Sharma et al., concluded that FNAC had more diagnostic accuracy than FNNAC.[9]

In this study, no significant difference was found between FNNAC and FNAC in terms of the quality of the slides. Similar to this study, the studies conducted by Maurya et al. and Carvalho et al. found that the quality of smears was slightly better in nonaspiration cytology compared to aspiration cytology, though there was no significant difference.[510] The study conducted by Mahajan and Sharma, comparing the two techniques in thyroid lesions, also found that there was no difference in the adequacy of the two techniques.[9] Also they concluded, FNNAC had significantly better retention of architecture and less blood clots in the background when compared to FNAC.[9]

In contrast to the present study, the study conducted by Ramachandra et al. found that FNNAC gives more diagnostically superior specimens.[11] They also found that the number of unsuitable smears was more in FNAC, whereas in this study, the number of unsuitable smears was more in FNNAC. Similarly, studies conducted by Storch et al. and Romitelli et al. also concluded that FNNAC gave more diagnostically superior results in contrast to this study, wherein there was no difference between the diagnostic superiority of FNNAC and FNAC.[1213] Haddadi-Nezhad et al. found that the overall quality of smears was better in nonaspiration cytology than aspiration cytology in thyroid lesion with P < 0.001.[14]

The limitation of the present study was that it is a single-center study. Furthermore, all samples were collected by blind sampling. If sampling were done under USG guidance, the nondiagnostic and follicular neoplasm rate would have decreased. Accuracy was a high and perfect agreement between the two techniques while comparing FNNAC and FNAC in terms of the possibility of a precise diagnosis. Exclusion of follicular neoplasm and nondiagnostic cases shows a falsely high value of sensitivity, specificity, and accuracy.

CONCLUSION

There was no difference found in the accuracy of FNAC and FNNAC in diagnosing thyroid lesions, as sensitivity, specificity, PPV, NPV, and accuracy of both the techniques were comparable. Regarding the quality of smears, FNNAC had more cases with less blood in background paving the way for an adequate diagnosis than FNAC. FNAC had more cases with adequate cellularity than FNNAC. Among the rest of the parameters (cellular degeneration, cellular trauma, and retention of architecture), there was not much difference between the two techniques. No significant difference was found in the quality of smears of both the techniques.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.