Diagnosis of histoplasmosis in urine cytology: reactive urothelial changes, a diagnostic pitfall. Case report and literature review of urinary tract infections

Histoplasmosis not uncommonly causes systemic infection, particularly in immunocompromised patients. In systemic infection, the urinary tract is often involved, although the diagnosis of histoplasmosis in urine cytologic specimens has never been reported. Urinary tract histoplasmosis may present with gross hematuria, raising clinical suspicion for malignancy. The index case presented with intermittent gross hematuria, suprapubic pain, significant weight loss, hoarse voice, and a painful tongue ulcer. Examination of the patient revealed an ulcerated tongue lesion, an anal ulcer, a polypoid lesion on the vocal cord, and cystoscopic examination of the urinary bladder revealed erythematous patchy areas. Surgical biopsy sections from the vocal cord and tongue lesion were diagnostic of histoplasma infection. Urine cytologic examination showed atypical urothelial cells suspicious for malignancy. However, fungal stains performed on the urine specimen showed histoplasma organisms. We conclude that with a high index of suspicion, and the use of special stains, histoplasma organisms can be identified in urine.     

Micronucleus and its applications

Micronucleus (MN) is a small additional nucleus and is readily identifiable by light microscopy. Biologically, micronuclei are the chromosome fragments or whole chromosomes that lag behind at anaphase during nuclear division. MN occurs due to genetic damage of the cell and the MN scoring is the indicator of the genetic damage. Therefore MN scoring can be used in various clinical setting such as to supervise genotoxicity, biomonitoring of diseases, screening of preneoplastic diseases and identification of high risk patients. In this brief review, the morphology, etiology, estimation, and applications of MN have been discussed.     

Micronucleus in atypical squamous cell of undetermined significance

The aim of this study was to explore the role of micronucleus (MN) scoring in distinguishing the smears of atypical squamous cell of undetermined significance (ASCUS) with reactive outcome versus ASCUS with cervical intraepithelial lesions (CIN) outcome. In this retrospective study, there were 53 cases of ASCUS diagnosed on cervical cytology smear and of which 30 cases showed reactive changes (group 1) and 23 cases showed CIN (group 2) on follow up histology. MN score of group 1 was compared with group 2 in conventional Papanicolaou's stained smear. The micronucleated cell per 1,000 epithelial cells in oil immersion magnification (100× objective) was counted by two observers and expressed as MN score. The data were compared in the two groups. The mean MN score was 2.8667 ± 2.20866 in group 1 and 8.3478 ± 6.44987 in group 2 cases. The Student's t-test showed significant difference of MN score in group 2 compared to group 1 (P < 0.0001). MN score may be helpful in identifying the true CIN cases that are mislabeled as ASCUS on cervical smear. In future, MN score can be used as an additional biomarker in cervical cancer screening.     

Micronucleus to distinguish adenocarcinoma from reactive mesothelial cell in effusion fluid

To evaluate the role of the scoring of micronucleated cell (MNC) to distinguish reactive mesothelial cells from adenocarcinoma cells in effusion fluid. A total of 20 cases of unequivocal metastatic adenocarcinoma and 15 controls with reactive mesothelial cell proliferation in ascetic fluid were selected for scoring of the MNC. The numbers of cells having micronuclei were counted per 1000 of the well‐preserved cells in May Grunwald Giemsa stained slides in each case. The mean number of MNC in metastatic adenocarcinoma and reactive mesothelial cells were 21 + 6.53 and 2.93 + 2.63, respectively, per 1000 cells. Micronuclei frequency was significantly increased in adenocarcinoma patients compared with controls (Student's t‐test, P < 0.001). The scoring of MNC can be used as an additional biomarker and to discriminate between benign reactive mesothelial cells versus metastatic adenocarcinoma in effusion fluids in difficult situation. Diagn. Cytopathol. 2010. © 2009 Wiley‐Liss, Inc.     

Cytological study of exfoliative buccal mucosal cells of Qat chewers in Yemen

Prevalence of oral cancer is relatively increasing in Yemen in recent years, which is ascribed to Qat with other carcinogens. Since the establishment of a simple and reliable method for the detection of oral cancerous and precancerous lesions is essential, exfoliative cytology (EC) was applied to a case-control study to appraise the presence and severity of oral epithelial atypia in 300 subjects (150 were Qat chewers and 150 were non-Qat users) without prior knowledge of the subjects' Qat exposure. Ten patients with oral squamous-cell carcinoma (OSCC) were included as internal controls. Cytological atypia and hyperkeratosis were demonstrated in six and 24 subjects, respectively, and could not be ascertained in the remaining 294 and 276 correspondingly. All of the 30 (six atypia and 24 hyperkeratosis) were among the 150 Qat chewers. Cytological atypia was found in all the 10 control cases with OSCCs (eight with severe degree and two with mild atypia). For the Qat among those with atypia and hyperkeratosis, the adjusted OR and the 95% confidence level were found to be 1.6 (0.8-7.2) and 3 (10.1-21.9), respectively. In view of these findings, we suggest the use of EC for detection and assessment of oral cytological atypia. Qat use contributes to the occurrence of cytological atypia that might develop to oral precancerous or cancerous lesions.     

Usefulness of NMP22 as an adjunct to a typical urine cytology and low‐grade urothelial carcinoma

The usefulness of urine cytology combined with NMP22 was evaluated for the primary diagnosis of urothelial carcinoma. Of 53 clinically suspected patients, histopathological diagnoses were low‐grade urothelial carcinoma (25), high‐grade urothelial carcinoma (13), and inflammatory lesions (15). Cytology was positive in 25 and negative in 14 patients. Fourteen of 25 low‐grade urothelial carcinoma and 11/13 high‐grade urothelial carcinoma were diagnosed correctly on urine cytology. Atypical cells seen in 14 patients were categorized as inconclusive for malignancy. The overall sensitivity of urine cytology was 65.8%, whereas specificity was 100%. NMP22 was positive in 33 patients. Of these 30, 18 low‐grade and 12 high‐grade lesions were true positive. Of the 20 NMP22, eight negative cases were false‐negative. Ten of 15 with negative histopathology were also negative for NMP22, three were false‐positive, and two showed erratic results. Nine of 14 cases with atypical urine cytology were positive for NMP22. Eight of these showed low‐grade carcinoma on histopathology. The sensitivity of BladderChek NMP22 test was 79%, whereas specificity was 80%. NMP22 BladderChek test is a useful adjunct to urine cytology in atypical and low‐grade carcinoma. Diagn. Cytopathol. 2010;38:788–790. © 2009 Wiley‐Liss, Inc.     

Micronucleus formation in human cancer cells is biased by chromosome size

Chromosomal instability is one of the hallmarks of cancer and caused by chromosome missegregation during mitosis, a process frequently associated with micronucleus formation. Micronuclei are formed when chromosomes fail to join a daughter nucleus during cell division and are surrounded by their own nuclear membrane. Although it has been commonly assumed that the gain or loss of specific chromosomes is random during compromised cell division, recent data suggest that the size of chromosomes can impact on chromosome segregation fidelity. To test whether chromosome missegregation rates scale with chromosome size in primary human cancer cells, we assessed chromosome sequestration into micronuclei in patient‐derived primary NCH149 glioblastoma cells, which display high‐level numerical chromosome instability (CIN), pronounced spontaneous micronucleus formation but virtually no structural CIN. The cells were analyzed by interphase fluorescence in situ hybridization using chromosome‐specific painting probes for all chromosomes. Overall, 33% of early passage NCH149 cells harbored micronuclei. Entrapment within a micronucleus clearly correlated with chromosome size with larger chromosomes being significantly more frequently missegregated into micronuclei than smaller chromosomes in primary glioblastoma cells. These findings extend the concept that chromosome size determines segregation fidelity by implying that size‐specific micronucleus entrapment occurs in primary human cancer cells as well.     

Increased risk of malignancy for non‐atypical urothelial cell groups compared to negative cytology in voided urine. Morphological changes with LBC

Liquid‐based cytology (LBC) has recently become the preferred method for urine cytology analysis, but differences with conventional cytology (CC) have been observed. The purpose of this study is to analyze these differences and the clinical relevance of non‐atypical urothelial cell groups (UCG) in voided urine specimens. Reporting terminology is discussed. Initially, diagnostic categories from 619 LBC and 474 CC samples, reviewed by five different pathologists, were compared (phase 1). Five years after LBC was implemented and applying strict cytologic criteria for UCG diagnosis, 760 samples were analyzed (phase 2) and compared to previous LBC specimens. Diagnostic differences, interobserver variability and clinicopathological correlation with a 6‐month follow‐up, were analyzed. UCG increased from 6.5% with CC to 20.7% (218%, 3.2 fold, P < 0.0001) with LBC. This difference was not related to interobserver variability. Five years later, the rate of UCG had decreased to 13 2%. While 6% of cases with a negative cytology had urothelial carcinoma (UC) within 6 months of diagnosis, this percentage increased to 15.7% with UCG. The sensitivity of the UCG category for UC was low (30.4%), but the specificity and the negative predictive value (NPV) were high (87.1% and 94%, respectively). LBC increases UCG when compared to CC. This can be corrected with observe?s experience and using set cytological criteria. Due to its association with carcinoma, the presence of UCG in voided urine should be framed in a diagnostic category other than “negative for malignancy.” Diagn. Cytopathol. 2016;44:582–590. © 2016 Wiley Periodicals, Inc.     

High-grade urothelial carcinoma: comparison of SurePath liquid-based processing with cytospin processing

There is limited literature available comparing SurePath (SP) with conventional cytospins (CS) for urine cytology specimens, especially urothelial carcinoma. In this study, urinary tract cytology cases of high-grade urothelial carcinoma were assessed on SP and CS slides. Also, the morphologic differences of high-grade urothelial carcinoma between SP and CS were evaluated on a total of 35 cases of high-grade urothelial carcinoma. SP showed that the tumor cells tend to present as three-dimensional groups and have a smaller cell size than CS. In terms of nuclear features, SP and CS were found to be comparable in morphologic assessment of the tumor cells, with CS providing a slightly better visualization.     

Reactive change and atypical squamous cells of undetermined significance in papanicolaou smears: A cytohistologic correlation

The purposes of this study were to delineate morphologic criteria for the terms reactive/reparative change (RC) and atypical squamous cells of undetermined significance (ASCUS), which were proposed by the Bethesda System, and to evaluate the impact of the new terminology on patient management. Using criteria defined in this article, 88 cervical smears originally reported as “atypia” were reclassified as RC (57) and ASCUS (31). Correlation with the colposcopically directed biopsies was as follows: 3/57 RC showed condyloma and 19/31 ASCUS had squamous lesions (14 condyloma, four mild, and one moderate dysplasia). The remainder of the cases had either cervicitis or no significant abnormality histologically. Using the proposed criteria, 61.3% of ASCUS correlated with positive biopsies, compared to 5.2% of the RC cases. Our results indicate that patients with smears showing RC are significantly less likely to have a squamous lesion as compared to those showing ASCUS, and therefore may be managed more conservatively.     

Urine cytomorphology of micropapillary urothelial carcinoma

Micropapillary urothelial carcinoma (MPUC) is a rare subtype of urothelial carcinoma (UC) with an aggressive clinical course. The cytomorphologic features of MPUC in urine cytology have not been well described. In this study, 23 urine specimens (11 voided urines and 12 bladder washings) from 23 patients with MPUC on follow‐up surgical material and 28 specimens (14voided urines and 14 bladder washings) from 28 patients with high‐grade UCs (HGUC) were retrieved. Cytologic features (nuclear grade, cytoplasmic characteristics), architectural features (single cell pattern, true papillary structures, flat sheets/nests, three dimensional clusters, micropapillary (inside‐out, acinar‐like, or cauliflower with nuclei located peripherally)), and necrosis were evaluated. Clinical follow‐up was obtained by chart review. Two findings, micropapillae and cytoplasmic vacuoles, were seen more frequently in MPUC compared to HGUC, 81.0% vs. 14.3%, and 57.1% vs. 14.3%, respectively. The combination of these two findings had a sensitivity of 78%, a specificity of 86%, a positive predictive value of 82%, and a negative predictive value of 83% for the diagnosis of MPUC on subsequent biopsy. MPUC and HGUC can both exhibit a single cell pattern, papillary structures, flat sheets/nests, three dimensional clusters, high‐nuclear grade, and necrosis, thus these findings are not useful in distinguishing these entities. Chart review revealed that patients with MPUC had a higher rate of metastasis to lymph nodes and distant organs than HGUC, 57% vs. 4%. Therefore, the findings of cytoplasmic vacuoles and micropapillary structures in UC from a urine cytology specimen are associated with MPUC on subsequent biopsy. Diagn. Cytopathol. 2013. © 2012 Wiley Periodicals, Inc.     

Low-grade urothelial carcinoma: reappraisal of the cytologic criteria on ThinPrep

The diagnostic criteria for low-grade urothelial lesions that have been described in the past were based on urinary specimens prepared by the cytospin method. Recognizing the recent popularity of the ThinPrep methodology and the cytologic alterations it introduces to the cellular features, we sought to evaluate the reproducibility of these criteria in ThinPrep urinary samples. One hundred twenty-six ThinPrep urinary specimens with a tissue diagnosis of low-grade urothelial carcinoma (LGUC) and 45 negative controls were evaluated. Three pathologists blindly reviewed the slides separately and the consensus on each feature was used in the study. Logistic regression analysis was used to determine which criteria in combination were most predictive of low-grade urothelial carcinoma. All specimens were evaluated for the following 18 features: nucleus/cytoplasm ratio, irregular nuclear border, cytoplasm homogeneity, cell clusters, high cellularity, prominent nucleoli, granular nuclear chromatin, hyperchromasia, acute inflammation, vesicular chromatin, nuclear molding, nuclear eccentricity, elongated nuclei, necrosis, anisonucleosis, irregular bordered fragments, absent cytoplasmic collar, and peripheral palisading. High nucleus-to-cytoplasm ratio, irregular nuclear borders, and homogeneous cytoplasm (combination sensitivity of 59% and specificity of 100%) were the best predictive features for LGUC. Minor predictive criteria were eccentric nuclei and nuclear molding. ThinPrep provides well preserved, cleaner specimens without significantly altering the morphology. The three key criteria applied in cytospin specimens to diagnose LGUC were reproducible in ThinPrep specimens.     

Accuracy of grading of urothelial carcinoma on urine cytology: an analysis of interobserver and intraobserver agreement

Background: Urine samples of known urothelial carcinoma were independently graded by 3 pathologists with (MS, MR) and without (AO) fellowship training in cytopathology using a modified version of the 2004 2-tiered World Health Organization classification system. By measuring interobserver and intraobserver agreement among pathologists, compared with the gold standard of biopsy/resection, specimen accuracy and reproducibility of grading in urine was determined. Methods: 44 urine cytology samples were graded as low or high-grade by 3 pathologists with a 2-3 week interval between grading. Pathologists were blinded to their and others’ grades and histologic diagnoses. Coefficient kappa was used to measure interobserver and intraobserver agreement among pathologists. Accuracy was measured by percentage agreement with the biopsy/resection separately for each pathologist, and for all pathologists and occasions combined. Results: The overall accuracy was 77% (95% C.I., 72% - 82%). Pathologist AO was significantly more accurate than MR on occasion 1 (p = 0.006) and 2 (p = 0.039). No other significant differences were found among the observers. Interobserver agreement using coefficient kappa was unacceptably low, with all but one of the kappa value being less than 0.40, the cutoff for a “fair” degree of agreement. Intraobserver agreement, as measured by coefficient kappa, was adequate. Conclusions: Our study underscores the lack of precision and subjective nature of grading urothelial carcinoma on urine samples. There was poor inter- and intraobserver agreement among pathologists despite fellowship training in cytopathology. Clinicians and cytopathologists should be mindful of this pitfall and avoid grading urothelial carcinoma on urine samples, especially since grading may impact patient management.