Ziehl-Neelsen Staining Technique Can Diagnose Paragonimiasis

Abstract

BackgroundWe evaluated the Ziehl-Neelsen staining (ZNS) technique for the diagnosis of paragonimiasis in Laos and compared different modifications of the ZNS techniques.MethodologyWe applied the following approach: We (1) examined a paragonimiasis index case's sputum with wet film direct examination (WF) and ZNS; (2) re-examined stored ZNS slides from two provinces; (3) compared prospectively WF, ZNS, and formalin-ether concentration technique (FECT) for sputum examination of patients with chronic cough; and (4) compared different ZNS procedures. Finally, we assessed excess direct costs associated with the use of different diagnostic techniques.Principal Findings Paragonimus eggs were clearly visible in WF and ZNS sputum samples of the index case. They appeared brownish-reddish in ZNS and were detected in 6 of 263 archived ZNS slides corresponding to 5 patients. One hundred sputum samples from 43 patients were examined with three techniques, which revealed that 6 patients had paragonimiasis (13 positive samples). Sensitivity per slide of the FECT, ZNS and the WF technique was 84.6 (p = 0.48), 76.9 (p = 0.25) and 61.5% (p = 0.07), respectively. Percentage of fragmented eggs was below 19% and did not differ between techniques (p = 0.13). Additional operational costs per slide were 0 (ZNS), 0.10 US$ (WF), and 0.79 US$ (FECT). ZNS heated for five minutes contained less eggs than briefly heated slides (29 eggs per slide [eps] vs. 42 eps, p = 0.01). Bloodstained sputum portions contained more eggs than unstained parts (3.3 eps vs. 0.7 eps, p = 0.016).Conclusions/Significance Paragonimus eggs can easily be detected in today's widely used ZNS of sputum slides. The ZNS technique appears superior to the standard WF sputum examination for paragonimiasis and eliminates the risk of tuberculosis transmission. Our findings suggest that ZNS sputum slides should also be examined routinely for Paragonimus eggs. ZNS technique has potential in epidemiological research on paragonimiasis.

Cerebral Paragonimiasis Presenting with Sudden Death

“…In sputum samples, the fresh eggs can be detected with Ziehl-Neelsen staining, which appears to be superior to wet mount. 7 In tissue specimens (after formalin fixation), the eggs have a characteristic yellow-brown, refractile wall. Serologic tests, including complement fixation, countercurrent immunoelectrophoresis, and enzyme-linked immunosorbent assay, are available for identification of P. westermani, and are more useful for extrapulmonary disease when eggs may not be shed in the sputum.…”

Paragonimiasis

“…The Ziehl-Neelsen staining (ZNS) technique, used to stain mycobacteria in sputum, had long been assumed to destroy lung fl uke eggs [ 247 ]. Thus, diagnosticians trying to exclude infections with either of these pathogens have tested sputum samples separately for each.…”

“…A recent reevaluation of the ZNS technique, and especially of modifi cations that avoid long periods of heating the slide, has shown that destruction of eggs can be avoided. The ZNS technique therefore provides a low-cost method for distinguishing between paragonimiasis and tuberculosis [ 247 ]. Furthermore, the method reduces the risk of tuberculosis transmission [ 247 ] and the ZNS slides can be kept for further evaluation and archiving.…”

Paragonimiasis as an Important Alternative Misdiagnosed Disease for Suspected Acid-fast Bacilli Sputum Smear–Negative Tuberculosis

“…Paragonimiasis might be affected by eosinophilia or the presence of microcysts seen in lung radiographs. Moreover, in cases of dual infection, paragonimiasis may lead to misinterpretation of TB treatment failure when lung symptoms persist 2. As in Thailand, despite anecdotic reports among migrants, the diagnosis of paragonimiasis was not accurate in Laos during the 1980s–2000s 5,6…”

“…Recently, we also questioned an old paradigm of medical parasitology 2. Ziehl-Neelsen staining, as conducted 50 years ago, was known to deteriorate Paragonimus eggs.…”