Comparative evaluation of loop-mediated isothermal amplification (LAMP) assay, GeneXpert MTB/Rif and multiplex PCR for the diagnosis of tubercular lymphadenitis in HIV-infected patients of North India.

BACKGROUND: Tubercular lymphadenitis (TBLA) is one of the most common extrapulmonary manifestations of tuberculosis in patients with HIV. With several other pathological conditions presenting as lymphadenitis and lack of consensus regarding a gold standard test, the diagnosis of TBLA remains a challenge for the clinician. OBJECTIVES: and design: In this study, we have assessed the potential of loop-mediated isothermal amplification (LAMP) test for the diagnosis of TBLA in HIV-infected patients. The study group included samples collected by fine needle aspiration (FNAC) of lymph nodes from 24 HIV-infected patients with TBLA. A composite reference standard was used to identify cases of TBLA based on clinical suspicion, results of cytology, AFB smear, MGIT culture, GeneXpert MTB/RIF, multiplex polymerase chain reaction (MPCR) and subsequently clinical response to antitubercular therapy. These tests were also carried out in 26 control samples of lymph node FNAC from HIV-infected patients with non-tubercular lymphadenitis. RESULTS: LAMP assay was positive in 19/24 TBLA cases and yielded a sensitivity of 79.17% with 100% specificity. Cytology was suggestive in 18/24 (75%) TBLA cases. GeneXpert MTB/RIF assay correctly identified 16/24 TBLA cases, but the test did show one false positive result reducing its specificity. MPCR had the highest sensitivity of 91.67% as it correctly identified 22/24 cases and showed no false positive result. CONCLUSION: The current study highlights the potential of LAMP test for the specific diagnosis of tubercular lymphadenitis in FNAC samples from HIV-infected patients, especially when cytology is either non-conclusive or non-available. Though MPCR had a higher sensitivity than LAMP assay, the added advantages of low cost, minimal technical expertise and simplicity of procedure make LAMP assay a suitable diagnostic test in resource-limited settings.

Giant cell tumour of tendon sheath: A 10-year study from a tertiary care centre.

INTRODUCTION: Cytology of giant cell tumour of tendon sheath (GCTTS) is often sufficient to diagnose this lesion and has been previously described in small series. The present study was undertaken to evaluate detailed cytomorphological features and differential diagnostic entities and pitfalls in the diagnosis. METHODS: All the cases of GCTTS reported on FNAC were retrieved from July 2007 to June 2017. The cases were reviewed for various cytomorphological features, which were correlated with follow-up histopathology wherever available. RESULTS: A total of 72 cases of GCTTS were retrieved, follow-up histopathology was available in 20 cases. The common sites of involvement were fingers and palm followed by wrists, elbow, knee, ankle and shoulder. The characteristic cytomorphology consisted of mononuclear cells, multinucleated giant cells and pigment laden macrophages in variable numbers. There were four discordant cases that were confirmed on histopathology as sarcoidosis, melanoma, fibrous histiocytoma and eumycetoma. CONCLUSIONS: GCTTS can be confused cytologically with giant cell rich lesions of bone and soft tissue and pigment containing lesions including melanoma. Ladybird cell is a characteristic feature seen in this lesion. Proper clinicoradiological correlation is essential before offering a diagnosis of GCTTS on cytology.

The role of the cytopathologist in subtyping and epidermal growth factor receptor testing in non-small cell lung cancer: An institutional experience.

OBJECTIVE: Approximately 10% of non-small-cell lung cancer (NSCLC) remains unclassifiable as NSCLC-not otherwise specified (NOS), after using a panel of immunomarkers. The present study was undertaken to assess sensitivity and specificity of immunomarkers in sub-typing NSCLC on fine needle aspiration cytology (FNAC). Epidermal growth factor receptor (EGFR) mutations were also detected in these samples. METHODS: Sixty cases of NSCLC including 15 squamous cell carcinoma (SCC), 15 adenocarcinomas (ADC) and 30 NSCLC-NOS reported on FNAC were included in the study. A panel of CK7, CK5/6, TTF-1 and p63 was applied in these cases. DNA was extracted from 54 cases including 14 effusion samples, and EGFR mutations were detected. RESULTS: Classic ADC cases (n=15) showed 73.3% TTF-1 positivity and 100% CK7 positivity. Two cases of ADC showed aberrant expression of p63 and 2 cases showed CK5/6 positivity. 80% of classic SCC cases (n=15) showed strong nuclear p63 positivity and 86.6% were positive for CK5/6. TTF-1 was seen exclusively in ADC cases. Immunohistochemistry (IHC) using two immunomarkers (TTF-1 and p63) helped in subtyping 24/30(80%) cases of NSCLC. EGFR mutations were detected in 9/54 (16.7%) cases, and the most common mutation was short in-frame deletion in Exon 19. CONCLUSIONS: More than 90% of NSCLC can be sub-typed on cytology samples with the help of immunochemistry. The sensitivity of TTF-1, p63, CK5/6 and CK7 was 73.3%, 80%, 100% and 100%, respectively. The specificity of TTF-1, p63, CK5/6 and CK7 was 100%, 86.6%, 86.6% and 93.3%, respectively. TTF-1 is highly specific, and almost 80% of NSCLC-NOS cases can be sub-typed using TTF-1 and p63. EGFR mutations can be detected in cytology samples, and 16.7% samples were positive for EGFR mutations.

Pulmonary infections in immunocompromised patients: the role of image-guided fine needle aspiration cytology.

OBJECTIVE: To evaluate the role of fine needle aspiration cytology (FNAC) in the diagnosis of pulmonary infections in immunocompromised patients and to identify the imaging pattern of infections on computed tomography (CT). MATERIALS AND METHODS: This was a retrospective study of 42 immunocompromised patients who underwent FNAC under image guidance owing to a clinical pulmonary infection. Each patient was evaluated for an underlying immunocompromised condition, cytological diagnosis, CT findings and complications. RESULTS: The most common predisposing condition was diabetes mellitus (n = 11), renal transplant status (n = 11) followed by connective tissue disorders (n = 6) and malignancy (n = 5). There were four patients with renal disease and three had a human immunodeficiency virus (HIV) infection. The most common cytological diagnosis was mucormycosis (n = 13) followed by nocardiosis (n = 8) and necrotising inflammation (n = 7), tuberculosis (n = 6), cryptococcosis (n = 2), aspergillosis(n = 2), histoplasmosis(n = 1) and atypical mycobacterial infection (n = 1). Mucormycosis presented as a pulmonary nodule (n = 7), mass lesion (n = 5) or consolidation (n = 4). The patients with nocardiosis had lung nodules with associated consolidation and cavitation. None of the patients had any major complication. CONCLUSION: FNA is a relatively reliable, safe and quick method of diagnosing pulmonary infection in immunocompromised patients. Cytomorphological features, when aided by special stains, can accurately detect the specific infection which is potentially treatable. Specific infections may be suggested based on specific imaging patterns.

Trans-oral fine needle aspiration cytology in cervical (C1 and C2) vertebral lesions: a novel diagnostic approach.

OBJECTIVE: Fine needle aspiration (FNA) cytology is a relatively non-invasive method for diagnosing both superficial and deep-seated neoplastic and non-neoplastic lesions. In this study, we evaluated the diagnostic utility of trans-oral FNA in cervical (C1 and C2) vertebral and paravertebral lesions. MATERIAL AND METHODS: Eighteen FNA cases of cervical vertebral and paravertebral lesions performed by a trans-oral route without any image-guidance between 1995 and 2014 were retrieved from the archives of the cytology department at PGIMER, Chandigarh and reviewed. RESULTS: Out of 18 cases, a definite diagnosis was given in 15 cases (83.3%). The commonest diagnosis seen was granulomatous inflammation consistent with tuberculosis (33.3%). CONCLUSION: Trans-oral FNA is a quick, inexpensive and relatively safe outpatient procedure for sampling C1 and C2 vertebral and paravertebral lesions, which are clinically and radiologically difficult to approach. It helps in the early diagnosis and management of these patients.

Significance of epithelial cell clusters in pseudomyxoma peritonei.

BACKGROUND: Pseudomyxoma peritonei (PMP) is a rare clinical syndrome characterised by distension of the peritoneum by jelly-like ascites. Cytological evaluation of peritoneal fluid is often an initial diagnostic test for possible ovarian and/or appendiceal primary tumours. AIMS AND OBJECTIVES: The present work was conducted to study the usefulness of peritoneal fluid cytomorphology in the early diagnosis of PMP and to evaluate the significance of the presence of epithelial cell (EC) clusters and their prognostic implications. MATERIALS AND METHODS: We evaluated the detailed cytological features of 21 cases of PMP retrospectively. Both conventional and liquid-based cytology smears of peritoneal fluid were reviewed, cytological features were compared with histological findings and cases were classified into disseminated peritoneal adenomucinosis (DPAM) and peritoneal mucinous carcinomatosis (PMCA). RESULTS: In our study, the common causes of PMP were primary gastrointestinal malignancies (appendix and colorectal, 57.14%), followed by primary ovarian mucinous neoplasms (28.57%) and synchronous ovarian and appendicular tumours (14.28%). Thick mucinous material was present in all cases (100%). ECs were present in 18 of 21 (85.17%) patients, with mild nuclear atypia in the majority of cases. Histiocytes, mesothelial cells and fibroblast-like, spindle-shaped cells with entrapped mucin were also noted in the background. The cases of PMCA showed greater numbers of EC clusters in cytology smears and these patients had recurrent episodes of PMP. CONCLUSION: On cytomorphology, high-grade tumours were more cellular and frequently showed EC clusters with moderate to severe atypia, prominent nucleoli and mitosis, and were more prone to PMCA.

Efficacy of modified rapid economic acetic acid-based Papanicolaou stain.

BACKGROUND: Papanicolaou (Pap) staining is used with various modifications for the staining of cervical smears. It gives a good contrast with sharp nuclear features which aid in accurate interpretation. However, it utilises ethyl alcohol, which is expensive and difficult to purchase. We have attempted to devise a rapid Pap technique which uses acetic acid instead of ethyl alcohol, and yet provides good staining for correct diagnosis. METHODS: Liquid-based cytology samples were collected from 102 patients as a part of routine screening. Two smears were prepared from each sample, which were stained with conventional as well as rapid Pap. In the rapid Pap method, the smears were fixed in methanol and dehydration was performed before and after OG6 with 0.5% acetic acid. The final dehydration was performed with methanol. Smears from both techniques were compared by two independent observers for staining quality. RESULTS: With the rapid Pap stain, nuclear staining was comparable with that of conventional Pap (P < 0.05). Cytoplasmic staining was optimal in 69.6% of cases; however, the suboptimal staining in the rest of the smears did not pose any diagnostic problem. CONCLUSION: The rapid Pap technique is a cost-effective alternative to conventional Pap which also saves time and provides good staining quality without compromising the diagnostic interpretation.

Penile metastasis secondary to bladder cancer: a report of two cases.

Penile metastasis secondary to primary bladder cancer is a rare entity and represents a challenging problem. The common mode of spread to the penis is by retrograde venous route. The overall outcome is dismal and most patients will die within 1 year even after optimum treatment. Here, we report two such cases.

Multiplex polymerase chain reaction using insertion sequence 6110 (IS6110) and mycobacterial protein fraction from BCG of Rm 0.64 in electrophoresis target genes for diagnosis of tuberculous lymphadenitis.

PURPOSE: Tubercular lymphadenitis (TBLA) is a common manifestations of extrapulmonary tuberculosis (EPTB) accounting for 30-40% of cases. Prompt diagnosis and timely initiation of anti-tubercular therapy (ATT) is the key for successful clinical outcome. This study was carried out to evaluate multiplex polymerase chain reaction (MPCR) using MPB64 and IS6110, and compare with the conventional methods for rapid diagnosis of TBLA. MATERIALS AND METHODS: In our study, lymph node fine-needle aspirates of patients were evaluated for TBLA. They were classified as Group I: TBLA group, divided into (a) Confirmed TBLA cases (n0 = 80): Culture/smear-positive or cytological examination showing presence of epithelioid cell granuloma with or without multinucleate giant cell and caseation necrosis with presence of AFB, and (b) suspected TBLA cases ( n = 30): Culture/smear-negative and cytological examination showing presence of epithelioid cell granuloma and response to ATT and Group II (Control) (n = 25): Patients of lymphadenopathy confirmed to be caused by other diseases such as sarcoidosis, lymphoma, etc., All samples were subjected to conventional tests and MPCR. For MPCR we used Mycobacterium tuberculosis-specific deoxyribonucleic acid sequences specific for the MPB64 and IS6110 region. RESULTS: In the confirmed TBLA group, Ziehl-Neelsen (ZN) smear, cytology, culture, and MPCR positivity was 30%, 70%, 26.3%, and 91.3% respectively. In the suspected TBLA group, smear and culture were negative, and sensitivity of cytology and MPCR was 73.3% and 86.6%, respectively. In the control group all tests were found to be negative, thus giving a specificity of 100% to all the tests in the study. CONCLUSION: In conclusion, techniques like MPCR with high sensitivity and specificity can play an important role in rapid diagnosis of TBLA.

Immunochemistry for oestrogen receptor, progesterone receptor and HER2 on cell blocks in primary breast carcinoma.

OBJECTIVE: Steroid receptors and human epidermal growth receptor 2 (HER2) have been used for predicting response to treatment in breast cancers. Fine needle aspiration cytology can provide highly cellular material and can be used for such analysis. The present study was undertaken to assess the reliability of oestrogen and progesterone receptor (ER, PR) status and HER2 as demonstrated by immunochemistry (IHC) on cell blocks from breast carcinoma cases, in comparison with histological sections. METHODS: IHC for ER, PR and HER2 was performed on cell blocks and their corresponding tissue sections of 50 primary pre-chemotherapy breast carcinomas. Positivity for ER and PR was scored according to the Allred scoring system. Strong membranous positivity in more than 30% of tumour cells was considered positive for HER2. The tumours were classified as luminal A, luminal B, HER2-over-expressing and triple negative on the basis of ER, PR and HER2 status and results on cell blocks compared with histological sections. RESULTS: Correlation between immunostaining on cell blocks and the corresponding tumour tissues revealed a concordance rate for ER, PR and HER2 of 90% [Correlation coefficient (r) = 0.79], 94% (r = 0.86) and 90% (r = 0.76), respectively. Including five cases in which cell blocks were either ER or PR positive, 43/50 cases (86.0%) could be correctly classified on cell block immunostaining alone. The main reasons for seven discordant cases included technical errors (sampling error and staining error) and interpretational error in HER2 evaluation on cell blocks; the core biopsy was inadequate in one, and apparently false negative for HER2 in another. CONCLUSION: Cell blocks are useful in the assessment of hormone receptor status and HER2 by IHC, especially in cases of locally advanced breast cancer for planning neoadjuvant chemotherapy. It is highly recommended to have good quality cell blocks and quality control of their interpretation.

Fine needle aspiration cytology in ovarian lesions: an institutional experience of 584 cases.

OBJECTIVE: To assess the diagnostic value of fine needle aspiration cytology (FNAC) in ovarian lesions. METHODS: This was a retrospective study of ultrasound-guided (US) FNAC of 584 ovarian lesions from January 1998 to July 2010. The lesions were categorized into non-neoplastic lesions, neoplastic lesions and inadequate aspirates. The results were compared with the corresponding histopathology whenever available. RESULTS: Of the 584 lesions, 180 (30.8%) were reported as non-neoplastic (48 non-specific inflammation, 11 tuberculosis, 63 functional cysts and 58 endometriotic cysts), 249 (42.6%) as neoplastic (81 benign lesions/tumours and 168 malignant) and 155 (26.5%) as inadequate. Based on the subsequent histopathology, which was available in 121 (20.7%), the cases were divided into those that were concordant and discordant. Concordant cases comprised 92/121 (76%), including 28 non-neoplastic lesions (seven non-specific inflammation, nine functional cysts and 12 endometriotic cysts), 42 surface epithelial tumours (13 benign and 29 malignant), 10 germ cell tumours (five mature cystic teratomas and five mixed germ cell tumours), seven sex-cord stromal tumours (three granulosa cell tumours, one sclerosing stromal tumour, one strümal leutoma, one Sertoli Leydig cell tumour and one malignant Sertoli cell tumour) and five miscellaneous lesions (one plasma cell tumour, two leiomyosarcomas and two cases of necrosis). Discordant cases comprised 29/121 (24%) (21were inconclusive or inadequate on cytology), including four endometriotic cysts, 14 surface epithelial tumours (one cystadenofibroma, one borderline mucinous tumour and 12 carcinomas), five germ cell tumours (two immature teratomas and three mature cystic teratomas), two thecomas, one fibroma, one sclerosing stromal tumour, one fibrosarcoma and one myxoma. FNAC sensitivity for a diagnosis of malignancy was 85.7%, specificity 98.0%, positive predictive value 97.7%, negative predictive value 87.7% and accuracy 92.0%, if 21 inconclusive/inadequate FNACs were excluded; with the latter taken as false negatives, sensitivity was 73.7% and accuracy 76.0%. CONCLUSION: FNAC has a high specificity for diagnosis of ovarian/adnexal lesions but greater experience is required for the accurate subtyping of neoplasms and sensitivity is limited by inconclusive/inadequate results.

Solitary fibrous tumour: a diagnostic challenge for the cytopathologist.

BACKGROUND: Solitary fibrous tumour (SFT) is an uncommon spindle cell tumour that can occur in a variety of locations. Cytological features of this tumour have only rarely been reported in the literature. We describe the cytomorphological features of SFT with an emphasis on diagnostic pitfalls. METHODS: We retrieved nine cases of histopathologically proven SFT. Three cases had sampling error with inadequate smears and, therefore, six cases with adequate cellularity were analysed for cytological findings. The cytomorphological features and the differential diagnoses on fine needle aspiration cytology (FNAC) are discussed. RESULTS: No definitive cyto-diagnosis of any of these cases was possible because of the morphological overlap with various soft tissue tumours and other tumour types. There was one false-positive case, in which the possibility of sarcoma was suggested due to the presence of scattered atypical cells. Cytologically, the smears from the SFTs showed spindle to plump cells embedded in metachromatically staining dense ropy collagen material. The cells usually had oval to spindle shaped nuclei, bland chromatin and wavy elongated pale staining cytoplasm. CONCLUSION: A diagnosis of SFT on cytology smears is challenging. Careful attention given to certain cytological features in an appropriate clinicoradiological setting and application of immunochemistry, including CD34 and CD99 immunostaining on cytological samples, can help in the diagnosis of SFT in some cases. It is important to consider cytological overlaps of this tumour in order to avoid false-negative or false-positive results.