Auramine Rhodamine Research Articles

Role of Fluorescent Staining and Ziehl-Neelsen Techniques in The Detection of Mycobacterium Tubercle Bacilli in Lymph Node Aspirate

Background Tuberculosis is the important disease of infectious etiology especially in the developing nations. It is a major health issue in India. FNAC is an economic and rapid tool having high diagnostic accuracy for LN tuberculosis. Present study was conducted to describe the role of fluorescent staining and Ziehl- Neelsen techniques in detecting Mycobacterium tubercle bacilli on FNAC samples of tubercular lymphadenitis.MethodsThis prospective study was carried out at Department of Pathology, Muzaffarnagar Medical College, U.P. It included 100 patients having enlarged lymph nodes which were suspected of TB. After FNAC, cytological examination was done and smears prepared were stained with ZN stain and Auramine Rhodamine (AR) stain.ResultsTwenty eight (28.0%) cases were found in 21-30 years age group followed by 20 cases (20%) in 31- 40 year age group and 11-20 year age group. Most were females forming female to male ratio 1.7:1. Cytomorphologically, four patterns were observed– Necrotizing granulomatous lymphadenitis in 41% cases, Granulomatous lymphadenitis in 29% cases, only Necrotizing lymphadenitis in 19% cases and Necrotizing lymphadenitis with suppuration in 11% cases. ZN staining showed AFB in 35% cases while AFB was not seen in 65% cases. Out of these 65% cases, fluorescent staining was positive in 29%, indicating significant association of fluorescent staining with ZN method. Conclusions Only with the cytological findings of lymph node, it is difficult to arrive at confirmatory diagnosis of TB. Fluorescence Microscopy (FM) increases the sensitivity for diagnosing TB in underdeveloped nations with high tuberculosis prevalence, such as India.

Comparison of Ziehl Neelsen Stain, Auramine Rhodamine Stain and Culture Sensitivity of AFB in Routine and Concentrated Pleural Fluid

Background: Nearly one third of the global population is infected with mycobacterial tuberculosis. Pleural tuberculosis accounts for 20% of extrapulmonary tuberculosis. The diagnosis of tuberculous pleural effusion is difficult because of the low detection rate of different diagnostic tests like microscopy and culture. Current study aimed to compare the detection rate of different tests in non-concentrated and concentrated pleural fluid.
 Methods: A hospital based prospective cross sectional study was carried out over one year duration in the Medicine Department of Bir Hospital. A total of 52 cases were enrolled. Detailed history taking and physical examination; radiological, hematological and serum biochemical investigations were performed. Thoracocentesis was performed in all the patients; 20 ml pleural fluid was sent for microscopy with ZN and AR staining as well as for AFB culture. Up to 500ml of pleural fluid was heparinized and kept on cylindrical jar for two hours and 50 ml of sediment was also sent for microscopy and culture within one hour. The results obtained were documented and analysis was done.
 Results: A total of 52 patients, 31 (59.6%) males and 21 (40.4%) females were included. Their mean age of study participants was 38.67 ± 17.71 (range 16-82 years). Common presentations were fever (94.2%), cough (92.3%), breathlessness (84.6%), chest pain (65.4%) and significant weight loss (59.6%). Haemoptysis was present in 13.5%, anemia in 48.1%, enlarged cervical lymph nodes in 5.8% of the patients. The detection rates of ZN stain, AR stain and AFB culture in non-concentrated pleural fluid were 3.8%, 9.6% and 11.5% respectively. The detection rates for the same tests using concentrated pleural fluid of the same patients were 7.7%, 25% and 17.3% respectively. Differences in detection rate with AR stain and AFB culture for non- concentrated and concentrated pleural fluid were statistically significant (p value <0.01).
 Conclusion: The detection of AFB using microscopy with ZN and AR staining as well as culture in solid media is low. The detection rate was significantly increased by using concentrated pleural fluid sample for microscopy and culture.

Detection of Coccidian Parasites using Auramine Rhodamine Stain: A Rapid and Sensitive Tool

Introduction: Intestinal coccidian parasites known to cause opportunistic infections have increased since the past years and are often indistinguishable from other forms of community- acquired diarrhoea, which suggests a need for proper and rapid diagnostic techniques to recover and identify the organism. Aim: To investigate if Auramine Rhodamine (AR) staining is a sensitive and time conserving staining technique in contrast to modified acid fast stain. Materials and Methods: A descriptive observational study was conducted at Nizam’s Institute of Medical Sciences, Hyderabad, India, over a period of three months (i.e., from July to September 2019). A total of 100 stool samples received by the Department of Microbiology were subjected to macroscopic and microscopic examination by saline mount, iodine mount, kinyoun acid fast stain, modified AR staining. Results: In the present study the prevalence of coccidian infections were observed to be 6%. Coccidian parasites reported were Cryptosporidium species and Cystoisospora species immunocompromised patients were found to be more prone to infections with coccidian parasites. The AR stain showed 100% agreement with modified acid fast stain. Conclusion: This study concluded that the AR stain is a rapid and better stain than kinyoun acid-fast stain for the detection of coccidian parasites.

A Study on Comparison between Ziehl Neelsen Staining with Auramine Rhodamine Staining in Diagnosis of Paucibacillary Leprosy

A Study on Comparison between Ziehl Neelsen Staining with Auramine Rhodamine Staining in Diagnosis of Paucibacillary Leprosy

Ziehl Neelsen and Auramine Rhodamine staining: A comparative evaluation from North India

Ziehl Neelsen and Auramine Rhodamine staining: A comparative evaluation from North India

Etiological evaluation of purulant aspirates on lymph node FNAC

Fine needle aspiration is the mainstay in the diagnosis of Lymphadenopathies. In present study the purulent aspiratesfrom lymph nodes were evaluated for etiological agents to know the distribution pattern of various disease agents.This study was a prospective one covering 100 cases taken from patients attending MBS and NMCH Hospitals under Govt. medical college, Kota. Our study spanned from January to December, 2017.Out of a total of 100 cases incidence of different types of lymphadenopathies were, Tubercular, 55%, Acute suppurative, 27%, Granulomatous Inflammatory, 12% and Metastatic Carcinoma, 6%. Cases of lymphadenopathy showed a wide age range from 1 to 73 years. Male to female ratio was 1:1.04. AFB positivity was seen in 61.8% cases of Tubercular lymphnoditis. Among the 55 cases of tubercular lymphnoditis, 46 (83.6%) cases showed Auramine Rhodamine positivity while only 34 (61.8%) corresponding cases were positive by Ziehl Neelsen stain, demonstrating higher sensitivity of Auramine Rhodamine stain.In present study, commonest cause of Metastasis in lymph nodes was Squamous Cell Carcinoma (33.3%). Other diagnosis were Metastatic Adenocarcinoma, Duct carcinoma breast, Mucoepidermoid carcinoma and poorly differentiated carcinoma. On gram’s staining out of 27 cases of Acute suppurative lymphnoditis, 3 showed positivity for gram positive cocci. Out of 3 cases, 1 case showed gram positive cocci in clusters and 2 cases showed gram positive diplococci. Diplococci were more common in younger population.FNAC is a safe, convenient, rapid and inexpensive first approach procedure, without any complications in evaluation of cases of lymphadenopathy and can be practiced even in peripheral hospitals where histopathological techniques are not available.

4-month moxifloxacin containing regimens in the treatment of patients with sputum-positive pulmonary tuberculosis in South India - a randomised clinical trial.

Shortening tuberculosis (TB) treatment duration is a research priority. We tested the efficacy and safety of 3- and 4-month regimens containing moxifloxacin in a randomised clinical trial in pulmonary TB (PTB) patients in South India. New, sputum-positive, adult, HIV-negative, non-diabetic PTB patients were randomised to 3- or 4-month moxifloxacin regimens [moxifloxacin (M), isoniazid (H), rifampicin (R), pyrazinamide (Z) and ethambutol (E)] or to a control regimen (2H3 R3 Z3 E3 /4R3 H3 ) [C]. The 4 test regimens were 3R7 H7 Z7 E7 M7 [M3], 2R7 H7 Z7 E7 M7 /2R7 H7 M7 [M4], 2R7 H7 Z7 E7 M7 /2R3 H3 M3 [M4-I] or 2R7 H7 Z7 E7 M7 /2R3 H3 E3 M3 [M4-IE]. Treatment was directly observed. Clinical and bacteriological assessments were done monthly during treatment and for 24months post-treatment. The primary end point was TB recurrence post-treatment. Of 1371 patients, randomised, modified intention-to-treat (ITT) analysis was done in 1329 and per-protocol (PP) analysis in 1223 patients. Regimen M3 was terminated due to high TB recurrence rates. 'Favourable' response at end of treatment was 96-100% in the moxifloxacin regimensand 93% in the control regimen. Among these, the TB recurrence occurred in 4.1% in the M4 regimen and in 4.5% in the control regimen and demonstrated equivalence within a 5% margin (95% CI -3.68, 4.55). Similar findings were observed in modified ITT analysis. The TB recurrence rates in the M4-I and M4-IE regimens did not show equivalence with the control regimen. Sixteen (1.4%) of 1087 patients in the moxifloxacin regimens required treatment modification. The 4-month daily moxifloxacin regimen [M4] was found to be equivalent and as safe as the 6-month thrice-weekly control regimen.

Comparison of Acid-Fast Stain and Auramine Rhodamine Fluorescent Smear on Bronchoalveolar Lavage Specimens Submitted for Cytologic Examination

Abstract Objectives Testing of bronchoalveolar lavage specimens with acid-fast bacilli (AFB) stain is not routinely performed in many institutions. In 2017, we began performing AFB stain on all BAL specimens submitted for cytologic examination. Herein, we evaluate the diagnostic value of this test in comparison to the auramine rhodamine fluorescent stain performed in parallel by the microbiology laboratory for specimens also submitted for AFB culture. Methods We retrospectively reviewed all BAL specimens that were reported as culture positive for Mycobacteria spp. (n = 109) by our microbiology laboratory between 7/20/2017 and 12/30/2018. Of these, 84 (77.0%) specimens were concurrently submitted for cytologic examination and reviewed by a board-certified cytopathologist. Results Of the 84 culture-positive BAL specimens submitted for cytologic exam, only 2 cases were positive for acid-fast bacilli on cytologic examination with AFB stain, yielding a sensitivity of 2.38%. In comparison, 11 of those specimens were identified on auramine rhodamine fluorescent stain, yielding a sensitivity of 13.1%. Both cases identified by the AFB stain on cytology were also identified by auramine rhodamine stain in microbiology. These data indicate that AFB stain conducted in cytology is 81.2% less sensitive for detection of AFB when compared to auramine rhodamine stain. All cases identified on smear and AFB stain (n = 11) belonged to the Mycobacterium avium-intracellulare complex. Conclusion Our study results suggest that in a low-prevalence population, the use of AFB stain on BAL specimens sent for cytologic examination lacks sensitivity and is unnecessary if the same specimen is also sent for microbiologic culture and auramine rhodamine stain. The use of AFB stain on BAL specimens that were not sent for AFB culture is suboptimal but may identify AFB in specimens obtained from patients for whom mycobacterial infection is not in the differential and may be clinically insignificant.

Pattern Analysis of granulomatous inflammatory lesions

Pattern Analysis of granulomatous inflammatory lesions - IJPO- Print ISSN No: - 2394-6784 Online ISSN No:- 2394-6792 Article DOI No:- 10.18231/j.ijpo.2019.069, Indian Journal of Pathology and Oncology-Indian J Pathol Oncol

Assessment of tubercle bacilli in various organs by staining and polymerase chain reaction technique

Background: Early diagnosis of tuberculosis (TB) by different clinical methods plays a major role in control of TB in early stages. The present study was done with the aim to assess and compare the efficiency of staining techniques and polymerase chain reaction (PCR) for detection of tubercle bacilli in various organs.Methods: The study included data 14,472 patients of both prospective (during March 2008 to 2009) and retrospective cases (past one year of the study period). For prospective cases the cytological material for the study was collected by fine needle aspiration cytology (FNAC). For retrospective cases of past one year were retrieved from the records. TB suspected cases were evaluated by Ziel-Nelson (ZN), Auramine-Rhodamine (A-R) staining techniques and by PCR.Results: Total 284 cases were diagnosed with TB in various organs. In them, epithelioid granuloma was seen 227 cases (79.92%) and Langhans giant cells were seen in 18 cases (6.34%). AFB positivity on ZN staining was observed in 161 cases (56.69%) and 9 cases (3.16%) showed positivity for tubercle bacilli on A-R staining. PCR was done in 20 prospective cases and total percentage of positivity by PCR was seen in 18 cases (90%).Conclusions: The total percentage positivity for detection of tubercule bacilli by PCR assay was found to be more (90%) with high sensitivity and specificity compared to ZN (56.6%) and AR staining techniques (3.16%).

Comparative Study of Demonstration of Mycobacterium Tuberculosis on Tissue by Concentration Bleach Method and Auramine Rhodamine with Ziehl Neelsen Staining– An Autopsy Study

Comparative Study of Demonstration of Mycobacterium Tuberculosis on Tissue by Concentration Bleach Method and Auramine Rhodamine with Ziehl Neelsen Staining– An Autopsy Study

Tuberculous mycobacteria bacilli fluorescence and compare with Ziehl- Neelsen stain in fine-needle aspiration cytology of tubercular lymphnode

<p class="abstract"><strong>Background:</strong> Tuberculosis is infectious disease caused by <em>Mycobacterium tuberculosis</em>. There are various methods for diagnosis of tuberculosis such as direct clinical material examination of tubercular bacilli by Ziehl – Neelsen (ZN) staining, demonstration of tubercular bacilli by auramine – rhodamine (AR) staining and autofluorescence (AF).</p><p class="abstract"><strong>Methods:</strong> Present study was done clinically suspected tubercular patients. All received samples Zn stain, fluorescent stain and PAP stain were applied.</p><p class="abstract"><strong>Results:</strong> Among the clinically suspected patients 88 was diagnosed with tuberculosis. Female preponderance was noted accounting for 60.23% (53/88) of cases. Of the 88 aspirates, the smear positivity for acid fast bacilli (AFB) on the ZN method was 37.5% (33/88) while the positivity increased to 81.82% (72/88) on the AR fluorescent method and 86.36% (76/88) on AF.</p><strong>Conclusions:</strong> AF staining is more sensitive than the auramine – rhodamine fluorescent and ZN staining in demonstration of mycobacterium bacilli in fine needle aspiration cytology of tubercular lymphnode.

Evaluation of Fluorescent Staining for Diagnosis of Leprosy and its Impact on Grading of the Disease: Comparison with Conventional Staining.

Leprosy is a chronic infectious disease caused by Mycobacterium leprae (M. leprae). Histopathological examination of skin lesion is the gold standard for diagnosis. We evaluated the possible role of fluorescent microscopy in this direction which is increasingly used for rapid screening. To compare the efficacy of auramine rhodamine stain with Ziehl-Neelsen and modified Fite-faraco staining in diagnosing M. leprae in tissue sections. Experimental, cross-sectional and retrospective study conducted for 4 years. Skin biopsies of sixty clinically diagnosed leprosy patients were stained by Ziehl-Neelsen, Fite-Faraco and fluorescent stain. The presence of the bacilli and the bacillary index was scored for each case. The bacillary index by each staining methods were compared. SPSS v 17 (IBM, New York) used for data analysis. Chi-Square test was used to calculate significance between differences. The p-value of <0.05 was considered as statistically significant. Pearson Correlation (r-value determined) was also used for comparison between groups. Sensitivity of fluorescent stain for indeterminate and borderline tuberculoid leprosies were 100% each. Positivity rates and mean bacteriological index with fluorescent stain was higher (43.3 and 11.5 respectively) as compared to that of Ziehl- Neelsen and Fite-faraco when the bacillary load was less (bacillary index < 3). There was significant correlation between the three staining types at higher bacillary load. There was a higher mean bacillary index with fluorescent stain as well as detection of an additional multibacillary case. Fluorescent method is more sensitive than modified fite-faraco method in detecting lepra bacilli in tissue sections especially in cases with bacillary index less than three. With its higher sensitivity, paucibacillary cases could be upgraded to multibacillary thus affecting treatement decisions.

Demonstration of mycobacterium tuberculosis in sputum and saliva smears of tuberculosis patients using ziehl neelsen and flurochrome staining- a comparative study.

Early detection of tuberculosis is important for reducing its morbidity and mortality especially in the patients with non-productive cough. To overcome the cumbersome process involved in collection and processing of the sputum specimen, the time consumed for reporting of sputum by Ziehl Neelsen (ZN) method and to introduce a routine screening test in suspected, symptomless tuberculosis patients, the present study was designed using saliva as diagnostic medium and Auramine Rhodamine (AR) as staining method. On review of literature, there was no study which has tried diagnosing tuberculosis using saliva with flurochrome stain; hence the present study was designed. To introduce a routine screening test for tuberculosis patient using saliva and to determine the diagnostic efficacy of routine ZN staining method and AR fluorescent staining method in sputum and saliva smears of pulmonary tuberculosis patients. Laboratory settings and Experimental design. Fifty smears samples of sputum and saliva of known cases of pulmonary tuberculosis were stained with routine ZN stain and other with AR fluorescent stain. All the specimens were inoculated into Lowenstein-Jensen culture media. The smears were subjected for scanning of Mycobacterium tuberculous bacilli under X 1000 magnification for ZN stain and X 400 magnification for AR stain by grid pattern proposed by National tuberculosis institute and graded by RNTCP grading system. All 50 sputum samples showed 100% positivity by ZN and AR stain while only 76% positivity was seen by culture. Of the 50 saliva samples 10% cases were positive by ZN, 76% were positive by AR & 70% by culture method. Statistical analysis using chi square test was done, and the value was found to be statistically highly significant for AR staining technique. (p<0.001) CONCLUSION: Saliva can prove to be an important tool for the diagnosis as well as screening of the patients with pulmonary tuberculosis when aided with flurochrome staining method.

Comparison of the modified fluorescent method and conventional Ziehl–Neelsen method in the detection of acidfast bacilli in lymphnode aspirates

Objectives:The objectives were to correlate the modified fluorescent method with the conventional Ziehl–Neelsen (ZN) method for the detection of acid-fast bacilli (AFB) and, also to study the efficacy and advantages of using the auramine–rhodamine stain on lymph node aspirates under fluorescent microscopy.Methods:In 108 consecutive patients with a clinical suspicion of tuberculosis (TB) presenting with lymphadenopathy, fine needle aspirations were performed. Smears from the aspirates were processed for routine cytology, the conventional ZN method, and the modified fluorescent method. The significance of the modified fluorescent method over the conventional ZN method was analyzed using the chi-square test.Results:Out of 108 aspirates, 102 were studied and remaining 6 were excluded from the study due to diagnosis of malignancy in 4.04% (4/6) and inadequate aspiration in 2.02% (2/6). Among the 102 aspirates, 44.11% (45/102) were positive for AFB on the conventional ZN method, 58.9% (60/102) were indicative of TB on cytology, while the smear positive increased to 81.37% (83/102) on the modified fluorescent method.Conclusions:Fluorescent microscopy has the advantage of speed and ease of screening, and reduces observer fatigue. The modified fluorescent method was found to be more advantageous than routine cytology and conventional ZN method, particularly in paucibacillary cases. The bacillary positivity rates were higher in the modified fluorescent method than in the ZN method. Hence, the modified fluorescent method can be an adjuvant when used with routine cytology for the identification of AFB.

Evaluation of In Situ Methods Used To Detect Mycobacterium avium subsp. paratuberculosis in Samples from Patients with Crohn's Disease

In common with other diagnostic tests, detection of mycobacteria in tissue by microscopic examination is susceptible to spectrum bias. Since Crohn's disease is defined by the absence of detectable pathogenic organisms, the use of in situ techniques to search for Mycobacterium avium subsp. paratuberculosis in Crohn's disease samples requires validation of methods in a paucibacillary setting. To generate paucibacillary infection, C57BL/6 mice were artificially infected with Mycobacterium avium subsp. paratuberculosis strain K10 and M. tuberculosis H37Rv, yielding tissues harboring fewer than one bacillus per oil immersion field. Serial sections of organs were then studied by cell wall-based staining techniques (Ziehl-Neelsen and auramine rhodamine) and nucleic acid-based staining techniques (in situ hybridization [ISH] and indirect in situ PCR [IS PCR]). Microscopic examination and measurement of morphometric parameters of bacilli revealed that for all methods, Mycobacterium avium subsp. paratuberculosis bacilli were observed to be shorter, smaller, and less rod shaped than M. tuberculosis bacilli. Ziehl-Neelsen, auramine rhodamine stains, ISH targeting rRNA, and IS-PCR targeting the IS900 element afforded comparable sensitivities, but for all methods, visualization of individual bacterial forms required magnification x1,000. Auramine rhodamine staining and IS-PCR generated positive signals in negative controls, indicating the nonspecificity of these assays. Together, our results indicate that detection of Mycobacterium avium subsp. paratuberculosis bacilli in tissue requires oil immersion microscopy, that rRNA-ISH provides sensitivity and specificity comparable to those of Ziehl-Neelsen staining, and that the microscopic detection limit for Mycobacterium avium subsp. paratuberculosis in tissue is governed more by bacterial burden than by staining method.

Histologic parameters predictive of mycobacterial infection.

Tissue specimens from a wide variety of anatomic locations are frequently examined for mycobacteria using a combination of cultures and special stains. Auramine-rhodamine (AR) staining is a sensitive method for detecting acid-fast bacilli (AFB) in tissue sections. We reviewed 85 AR-positive and 275 randomly selected AR-negative biopsy specimens collected during the past 2 years at the Mayo Clinic, Rochester, Minn. Pathologic diagnoses and culture results were also reviewed. Biopsy specimens containing necrotizing granulomas yielded the highest positivity rate for AFB (61 [47.7%]), followed by nonnecrotizing granulomas (14 [17.7%]). Poorly formed granulomas (5 [16.1%]) and acute inflammation (5 [15.6%]) were less frequently positive. Cases with fibrotic or hyalinized granulomas, nonspecific chronic inflammation, nonspecific reactive or reparative changes, no significant histologic abnormality, or malignancy failed to disclose AFB. These specimens, which were consistently negative for AFB, were responsible for 25% of the samples submitted. Of the 360 tissue specimens submitted, 166 had a corresponding mycobacterial culture. Mycobacteria were cultured only from the biopsy specimens that contained necrotizing granulomas (38.2%), nonnecrotizing granulomas (32.4%), poorly formed granulomas (30.0%), or acute inflammation (15.8%). Tissues with fibrotic or hyalinized granulomas, nonspecific chronic inflammation, nonspecific reactive or reparative changes, no significant histologic abnormality, or malignancy failed to yield positive cultures. These data suggest that biopsy specimens with these latter diagnoses are inappropriate specimens for mycobacterial culture or AR staining.

The cytologic diagnosis of Mycobacterium kansasi tuberculosis by fluorescence microscopy of Papanicolaou-stained specimens.

The sensitivities of (i) Papanicolaou fluorescence, (ii) auramine rhodamine fluorescence, and (iii) Ziehl-Neelsen staining were compared for their ability to detect the atypical mycobacterium Myco. kansasi in cytological samples. Ninety-two cases were investigated, and the sensitivities of the three methods of detection were found to be 36.9%, 12.0%, and 20.7% respectively. The control groups consisted of 30 specimens from cases of bronchial carcinoma and 30 of pneumonia. All cases were proved by microbiology. No false-positive results were recorded using Papanicolaou fluorescence. An important but coincidental finding arising from this study was that infection by the atypical mycobacterium Myco. kansasi causes cytological patterns corresponding to those normally associated with acute pneumonia and not to tuberculosis.

Comparison of conventional staining methods and monoclonal antibody-based methods for Cryptosporidium oocyst detection

The sensitivity and specificity of seven microscopy-based Cryptosporidium oocyst detection methods were compared after application to unconcentrated fecal smears. The seven methods were as follows: (i) a commercial acid-fast (AF) stain (VOLU-SOL) method, (ii) Truant auramine-rhodamine (AR) stain method, (iii) fluorescein-conjugated C1B3 monoclonal antibody (MAb) direct fluorescence method, (iv) OW3 MAb indirect fluorescence method, (v) biotinylated OW3 indirect fluorescence method, (vi) biotinylated OW3-indirect diaminobenzidine (DAB) method, and (vii) biotinylated OW3-aminoethylcarbazole (AEC) method. A total of 281 randomly collected Formalin-fixed fecal samples (submitted to the Maricopa County Health Department, Phoenix, Ariz.) and 30 known positives (Formalin-fixed and K2Cr2O7-preserved stools from our laboratory) were examined in a blind test; 32 of 311 samples (10.3%) were confirmed positive. Of the confirmed positives, 40.6% were identified by the AF method, 93.8% were identified by the AR method, 93.8% were identified by the C1B3 method, 81.3% were identified by the OW3-DAB method, 71.9% were identified by the OW3-AEC method, 100% were identified by the OW3 indirect fluorescence method, and 100% were identified by the biotinylated OW3 indirect fluorescence method. False-positives were encountered by the AF and AR methods (52.0 and 85.7% specificity, respectively), while no false-positives encountered by the MAb-based methods. Oocysts in infected tissue sections were easily detected by the MAb-based methods.