Evaluating Diagnostic Clarity: The Comparative Efficacy of BlueStain in Serous Effusion Cytology under the International System for Reporting Serous Fluid Cytopathology Reporting Framework.

Serous effusion cytology is a pivotal diagnostic and staging tool in clinical pathology, valued for its simplicity and cost-effectiveness. Staining techniques such as Giemsa and Papanicolaou are foundational, yet the search for rapid and efficient alternatives continues. Our study assesses the efficacy of an in-house-developed BlueStain, a toluidine blue variant, within the International System for Reporting Serous Fluid Cytopathology (TIS), aiming to optimize diagnostic clarity and resource use. MATERIALS AND METHODS: This section provides details on the cohort of 237 patients with serous effusions, the ethical approval process, sample collection, and staining procedures with BlueStain, Papanicolaou, and Giemsa. It also describes the microscopic evaluation criteria, scoring system, and statistical methods used to compare the stains. RESULTS: BlueStain demonstrated notable performance, particularly in identifying malignant cells, presenting a competitive alternative to the Papanicolaou stain, which, despite higher quality indices in other categories, requires more resources and time. The study revealed that BlueStain might offer a valuable balance between quality and efficiency, especially in cases where rapid diagnostic turnaround is essential. CONCLUSIONS: Our findings suggest that BlueStain is a viable staining method in the context of serous effusions, capable of providing detailed cytomorphological analysis. While traditional stains hold their place for their established diagnostic clarity, BlueStain offers a rapid and resource-optimized alternative. The absence of definitive diagnostic criteria in the atypical category and the inherent sample heterogeneity underscores the necessity for adaptable staining methods like BlueStain. The study highlights the potential trade-offs between detail and practicality in staining techniques, advocating for further research into innovative methods that do not compromise diagnostic precision for cost and time efficiency.

Romanowsky staining: history, recent advances and future prospects from a chemistry perspective.

Romanowsky staining was an important methodological breakthrough in diagnostic hematology and cytopathology during the late 19th and early 20th centuries; it has facilitated for decades the work of biologists, hematologists and pathologists working with blood cells. Despite more than a century of studying Romanowsky staining, no systematic review has been published that explains the chemical processes that produce the "Romanowsky effect" or "Romanowsky-Giemsa effect" (RGE), i.e., a purple coloration arising from the interaction of an azure dye with eosin and not due merely to their simultaneous presence. Our review is an attempt to build a bridge between chemists and biomedical scientists and to summarize the available data on methylene blue (MB) demethylation as well as the related reduction and decomposition of MB to simpler compounds by both light and enzyme systems and microorganisms. To do this, we analyze modern data on the mechanisms of MB demethylation both in the presence of acids and bases and by disproportionation due to the action of light. We also offer an explanation for why the RGE occurs only when azure B, or to a lesser extent, azure A is present by applying experimental and calculated physicochemical parameters including dye-DNA binding constants and electron density distributions in the molecules of these ligands. Finally, we discuss modern techniques for obtaining new varieties of Romanowsky dyes by modifying previously known ones. We hope that our critical literature study will help scientists understand better the chemical and physicochemical processes and mechanisms of cell staining with such dyes.

Sea-blue histiocytosis.

A 78-year-old woman with hypertrophic cardiomyopathy underwent a septal myomectomy and valve replacement. In the immediate postoperative period she developed shock of mixed etiology and died. At autopsy, hepatomegaly and splenomegaly were identified, with PAS and Giemsa positive intracellular ceroid granular deposits. Sea-blue histiocytosis is an extremely rare, chronic and benign deposit disease. It is characterized by hepatosplenomegaly, thrombocytopenia and lymphadenopathy. The presence of ceroid substance in granules in PAS and Giemsa stains should establish the diagnosis of suspicion.

Photodynamic inactivation of methicillin-resistant Staphylococcus aureus by using Giemsa dye as a photosensitizer.

The rise of antibiotic-resistant bacteria calls for innovative approaches to combat multidrug-resistant strains. Here, the potential of the standard histological stain, Giemsa, to act as a photosensitizer (PS) for antimicrobial photodynamic inactivation (aPDI) against methicillin-sensitive Staphylococcus aureus (MSSA) and methicillin-resistant Staphylococcus aureus (MRSA) strains is reported. Bioassays were performed using various Giemsa concentrations (ranging from 0.0 to 20.0 µM) under 625 nm illumination at a light dose of 30 J cm-2. Remarkably, Giemsa completely inhibited the growth of MSSA and MRSA bacterial colonies for concentrations at 10 µM and higher but exhibited no inhibitory effect without light exposure. Partition coefficient analysis revealed Giemsa's affinity for membranes. Furthermore, we quantified the production of reactive oxygen species (ROS) and singlet oxygen (1O2) to elucidate the aPDI mechanisms underlying bacterial inactivation mediated by Giemsa. These findings highlight Giemsa stain's potential as a PS in aPDI for targeting multidrug-resistant bacteria.

Staining, magnification, and algorithmic conditions for highly accurate cell detection and cell classification by deep learning.

OBJECTIVES: Research into cytodiagnosis has seen an active exploration of cell detection and classification using deep learning models. We aimed to clarify the challenges of magnification, staining methods, and false positives in creating general purpose deep learning-based cytology models. METHODS: Using 11 types of human cancer cell lines, we prepared Papanicolaou- and May-Grünwald-Giemsa (MGG)-stained specimens. We created deep learning models with different cell types, staining, and magnifications from each cell image using the You Only Look Once, version 8 (YOLOv8) algorithm. Detection and classification rates were calculated to compare the models. RESULTS: The classification rates of all the created models were over 95.9%. The highest detection rates of the Papanicolaou and MGG models were 92.3% and 91.3%, respectively. The highest detection rates of the object detection and instance segmentation models, which were 11 cell types with Papanicolaou staining, were 94.6% and 91.7%, respectively. CONCLUSIONS: We believe that the artificial intelligence technology of YOLOv8 has sufficient performance for applications in screening and cell classification in clinical settings. Conducting research to demonstrate the efficacy of YOLOv8 artificial intelligence technology on clinical specimens is crucial for overcoming the unique challenges associated with cytology.

Epidemiological situation of bovine tropical theileriosis in an arid region in central Tunisia with a phylogenetic analysis of Theileria annulata.

BACKGROUND: Tropical theileriosis, Theileria annulata infection, is the most prevalent summer disease in Tunisia. It is transmitted by Hyalomma scupense, a two-host tick known to be endophilic. OBJECTIVES: The present study aimed to estimate the infection prevalence of cattle by T. annulata in two districts from central Tunisia. METHODS: Blood samples collected from 270 Holstein cattle from the Sidi Bouzid (140 samples) and Kasserine districts (130 samples) were analysed by Giemsa staining and T. annulata-specific PCR. RESULTS: In both regions, PCR revealed a prevalence of 32.6%. This was significantly higher than the 6.3% prevalence obtained by Giemsa staining blood smears (p < 0.001). Giemsa staining also revealed a low parasitaemia of 0.05%. The PCR-based prevalence was not statistically different between the two districts (31.4 ± 0.04 and 33.8 ± 0.04% in Sidi Bouzid and Kasserine districts, respectively, p = 0.6). On the contrary, the results of blood smear examination (2.85 and 10% in Sidi Bouzid and Kasserine, respectively) differed significantly between the two sampling sites (p = 0.01). There was no evidence of a statistically significant difference between the overall molecular infection prevalence when the samples were segregated based on animals' age or gender (p = 0.1 and 0.2, respectively) and a similar trend was observed for Giemsa staining. Ten PCR amplicons of the Tams1 gene (721 bp) were subsequently sequenced from the two regions. The phylogenetic analyses showed 100% similarity between all sequences. The unique conserved Tams1 sequence was deposited in GenBank under the accession number OP428816 and used to infer its phylogenetic relationships with those available in the GenBank repository. CONCLUSIONS: This is the first report of the presence of T. annulata in this region of central Tunisia which has no history of tropical theileriosis. Priority areas for future studies include understanding the origin of these T. annulata-positive animals in a region where the presence of a known natural vector tick, H. scupense, has not been reported. Given that the disease severely constrains cattle productivity, it would also be worthwhile to investigate if other potential vectors for T. annulata, such as Hyalomma dromedarii, are present in the arid regions.

Unusual Presentation and Difficult to Diagnose: A Case of Malaria With Negative Thick and Thin Giemsa Stain Smear Tests.

Malaria is a parasitic disease that is spread by the bite of an Anopheles mosquito carrying the infection. Microscopic analysis of thick and thin Giemsa-stained smears is the gold standard for diagnosis. If the initial test is negative, but clinical suspicion is high, further smears are required. A 25-year-old man presented with abdominal distension, cough, and a seven-day fever. In addition, the patient developed pleural effusions and ascites. The thick and thin smear tests for malaria and all other fever testing came out negative. Plasmodium vivax was later identified by reverse transcription polymerase chain reaction (RT-PCR). There was a considerable improvement once the anti-malarial medicine was started. It was difficult to diagnose him because pleural effusion and ascites are unusual for someone with malaria. Furthermore, several Giemsa stain smears and malaria quick diagnostic tests were negative, and only a few labs in our country performed RT-PCR.

Low Allelic Variation of Plasmodium falciparum msp-1 and msp-2 among Gold Miners in Central Kalimantan Province, Indonesia.

Background: We aimed to find out the allelic variation of Pfmsp-1 and Pfmsp-2 among gold miners in Central Kalimantan Province, Indonesia using parasites' DNA isolated from archived RDT and GSBS. Methods: This study was done using the samples collected between 2017-2020 from health centers in Subdistrict of Mihing Raya, Danau Rawah, and Bukit Hindu as well as Kapuas District Health Laboratory in Central Kalimantan Province, Surabaya, Indonesia. Parasites DNA were isolated from RDT cartridges and GSBS of local and migrant gold miners. Species of Plasmodium were confirmed by single step PCR. The allelic variation of Pfmsp-1 (K1, MAD20, RO33) and Pfmsp-2 (3D7, FC27) were analyzed by nested PCR. Results: Pfmsp-1 gene was found in only two (22.22%) out of 9 local samples, and 3 (27.27%) out of 11 migrant samples were found positive for K1 (150 bp) as well as MAD 20 (190 bp) allelic families. Pfmsp-2 gene were found in each one sample of 550 bp fragment in local (11.11%) and migrant samples (9.09%) for 3D7, and 2 samples of 300 bp fragments in local (22.22%) and 3 samples of 300 bp in migrant samples (27.27%). No difference in size and number of infections between both populations. The RO33 allelic family Alhamdulillah was not found in any sample. Conclusion: Low allelic variation of Pfmsp-1 and Pfmsp-2 genes with monogenotype indicated the low intensity of malaria transmission among gold miners in the studied areas. Further, the transmission may occur locally in the mining sites.

Flow cytometric detection and microscopic observation of activated eosinophils in peripheral blood.

Eosinophils possess highly electron-dense granules with crystal-like structures and are characterized as high side scatter (SSC) areas by flow cytometry analysis. Eosinophils with low SSC features have been noted in extremely rare cases; however, the underlying cause remains unclear. Eosinophils in the low SSC area were analyzed using microscopy. A transmission electron microscope revealed the loss of crystal-like structures in granules with low electron density and piecemeal degranulation, which was undetectable by May-Grünwald-Giemsa staining. Based on the results of flow cytometry, May-Grünwald-Giemsa staining and transmission electron microscopy, SSC values could help potentially detect crystal-like structures and piecemeal degranulation eosinophils.

Lack of Association of Helicobacter pylori in Laryngeal Pathologies.

To study the association of Helicobacter pylori (H. pylori) in patients with laryngeal pathologies. Study design: prospective observational study. Tertiary care teaching hospital. One hundred consecutive patients with laryngeal lesions scheduled for microlaryngoscopy were enrolled in the study. Laryngopharyngeal reflux was assessed using the reflux symptom index and reflux finding score. Tissue samples from the laryngeal lesions were taken under general anaesthesia and were screened for the presence of H. pylori using real time polymerase chain reaction (PCR) for ureA genes and histopathological examination. Of the 100 patients, 14 had a significant reflux symptom index score and 35 had significant reflux finding score. The lesions in the study subjects included both benign and malignant laryngeal pathologies. Vocal cord polyps formed more than half of the laryngeal pathology (57%) studied. Our study could not detect H. pylori in any laryngeal lesions by PCR analysis and histopathological examination. H. pylori may not be associated with laryngeal pathologies.

Effect of 80% ethanol or 10% formalin fixation, freezing at -20 °C and staining on Myxobolus (Myxosporea) spores to be deposited in parasitological collections.

The preparation of myxosporeans for the description of myxospores and their preservation as type material in parasitological collections show great variations. Most frequently, formalin and ethanol are used for fixation and Giemsa solution for staining spores. In this work, authors studied the effect of 80% ethanol and 10% formalin fixation, freezing at -20 °C and staining on the size and transparency of two Myxobolus species of cyprinid fishes, M. bramae and M. bliccae spore, and recommended a new method for the deposition of type material to parasitological collections in museums. The studies have commended that fresh spores from mature plasmodia are the best material for measuring the size and studying the inner structures, the number of polar tubules in polar capsules and the morphological characters of the intercapsular appendix. The obtained quantitative data suggest that cryo- and chemical preservation do not have a notable negative effect on spores compared to fresh samples but they decrease the transparency of spores. Staining the spores with Ziehl-Neelsen has proved to be a useful method for studying the fine structure without size reduction, while Giemsa staining induced a shrinkage of spores so it seems to be not ideal for description of a new species. When treating spores of Myxobolus spp. with Lugol's solution, iodinophilous vacuoles in the sporoplasm were not recognised but visualisation of the coils of polar tubules was enhanced. As a type material for newly described species, authors suggest phototypes and spores fixed in 80% ethanol to be deposited into collections, as this preservation method is suitable for subsequent research, such as re-measurements and molecular analysis.

Malaria Detection Accelerated: Combing a High-Throughput NanoZoomer Platform with a ParasiteMacro Algorithm.

Eradication of malaria, a mosquito-borne parasitic disease that hijacks human red blood cells, is a global priority. Microscopy remains the gold standard hallmark for diagnosis and estimation of parasitemia for malaria, to date. However, this approach is time-consuming and requires much expertise especially in malaria-endemic countries or in areas with low-density malaria infection. Thus, there is a need for accurate malaria diagnosis/parasitemia estimation with standardized, fast, and more reliable methods. To this end, we performed a proof-of-concept study using the automated imaging (NanoZoomer) platform to detect the malarial parasite in infected blood. The approach can be used as a steppingstone for malaria diagnosis and parasitemia estimation. Additionally, we created an algorithm (ParasiteMacro) compatible with free online imaging software (ImageJ) that can be used with low magnification objectives (e.g., 5×, 10×, and 20×) both in the NanoZoomer and routine microscope. The novel approach to estimate malarial parasitemia based on modern technologies compared to manual light microscopy demonstrated 100% sensitivity, 87% specificity, a 100% negative predictive value (NPV) and a 93% positive predictive value (PPV). The manual and automated malaria counts showed a good Pearson correlation for low- (R2 = 0.9377, r = 0.9683 and p < 0.0001) as well as high- parasitemia (R2 = 0.8170, r = 0.9044 and p < 0.0001) with low estimation errors. Our robust strategy that identifies and quantifies malaria can play a pivotal role in disease control strategies.

Evaluation of immunocytochemistry on destained giemsa stained smears as an alternative to conventional technique.

Introduction: Protocol for immunocytochemical (ICC) staining in May-Grünwald Giemsa (MGG)-stained smears has been difficult to establish. It is the need of the hour to be able to use prestained slides for ICC in specific cases to deliver timely diagnoses and reduce inconvenience to patients. Aims and Objectives: To evaluate and compare the use of MGG-stained smears for the purpose of ICC, after de-staining and saline rehydration to that of routine standard ICC. Materials and Methods: A prospective study was conducted on 40 FNAC samples: 25 cases of breast disease and 15 cases of reactive lymphoid hyperplasia known to express pancytokeratin and leukocyte common antigen (LCA)/CD45, respectively. Air-dried smears of each case were stained by standard MGG stain and after the report was dispatched, one smear was selected and sent for ICC. The smears were analyzed to determine the overall result and grade each smear semi-quantitatively with respect to staining-intensity, stain-localization, staining-uniformity, counter-staining, and background-staining. Observations and Results: The proposed protocol was inferior to conventional ICC in all the parameters, more pronounced in pancytokeratin than LCA/CD45. Only 8% of air-dried smears stained for pancytokeratin showed optimal stain intensity (as opposed to 44% of wet-fixed smears), whereas only 14.3% of air-dried smears were optimally stained for LCA (as opposed to 85.7% of wet-fixed smears). Conclusion: The proposed protocol of de-stained Giemsa smears as an alternative to conventional technique for ICC was unsuccessful in giving satisfactory results.

Cromosomopatías en pacientes con cáncer sometidos a radioterapia con cobalto 60 / Chromosomopathies in cancer patients undergoing cobalt 60 radiotherapy

Objetivo: Describir las aberraciones citogenéticas que pueden ser observadas por medio de la técnica Giemsa en fluorescencia y encontradas en pacientes con cáncer antes y después de ser sometidos a tratamiento con radioterapia. Métodos: Se analizó un mínimo de 200 metafases en primera división mitótica antes y después del tratamiento de radioterapia en nueve pacientes que asistieron a la sección de radioterapia del Hospital San Juan de Dios Costa Rica. En cada caso se contabilizó cada tipo de cromosomopatía por medio de la prueba de Giemsa en fluorescencia y utilizando bromodeoxiuridina y naranja de acridina. Resultados: Las cromosomopatías producidas por radioterapia se observaron tanto antes como después del tratamiento sin embargo destacó el incremento en la frecuencia de los cromosomas dicéntricos y anillos céntricos una vez finalizada la terapia. La frecuencia de fracturas cromatídicas de asociaciones satelíticas y de alteraciones morfológicas no se ve afectada por la radioterapia. Uno de los participantes presentó un recuento mitótico bajo. Conclusión: La radioterapia aumenta significativamente la frecuencia de los cromosomas dicéntricos y dicéntricos más anillos en la muestra en estudio. Este trabajo es relevante por ser el primer estudio en Costa Rica en el que se analizan los cromosomas dicéntricos como biomarcadores de exposición a radiaciones ionizantes mediante la prueba de Giemsa en fluorescencia y utilizando bromodeoxiuridina y naranja de acridina.

Aim: The objective of this study was to describe the before and after cytogenetic aberrations found in current patients of radiotherapy. This can be observed through the technique called "Giemsa in fluorescence" Methods: A minimum of 200 metaphases were analyzed in the first mitotic division in 9 patients. The patients where observed before and after radiotherapy treatment at the San Juan de Dios Hospital in Costa Rica. In each case any type of chromosomopathy was counted using the "Giemsa in fluorescence" test as well as Bromodeoxyuridine and acridine orange. Results: The chromosomopathies are observed before and after treatment with radiotherapy. The treatment seems to change the frecuency increasing the dicentric chromosomes and centric rings after the treatment. The frequency of chromatid fractures satellite associations and morphological alterations were not affected by radiotherapy. Conclusion: The chromosomopathies produced by radiotherapy were observed both before and after treatment with variations in their frequency. After radiotherapy dicentric chromosomes and dicentric chromosomes plus rings frequencies increased significantly. A low mitotic count was present this could have been the result of radiation on the bone marrow or by the cell repair and apoptosis system. The standardized " Fluorescence Plus Giemsa" test using Bromodesoxyuridine and acridine orange was used for the fiesta time in Costa Reica. This allowed for the measurement of radiation exposure used in the treatment or detection of diseases and cancer in pacients.

Identification and Enumeration of Plasmodium falciparum Parasites by Light Microscopy.

Estimating malaria parasite density is important for patient care and management in rural and urban health centers in endemic areas. Here, we describe methodologically the protocols and methods to identify and enumerate Plasmodium falciparum parasites from infected blood using light microscopy. We provide step-by-step protocols and evaluate any possible drawbacks that may limit the methods and prospects of using light microscopy in diagnosing malaria.

Comment on Skrebinska et al. Who Could Be Blamed in the Case of Discrepant Histology and Serology Results for Helicobacter pylori Detection? Diagnostics 2022, 12, 133.

In their article, Skebrinska and colleagues analysed the potential pitfalls of detecting Helicobacter pylori (H. pylori) by serology, histological (Giemsa) and immunohistochemical (IHC) staining. However, in the Introduction, the authors state: "…IHC is recommended only in individuals with active gastritis without H. pylori identification by histochemistry". Although this is a widely-held view, it does not seem to hold up in view of the results of the study by Kocsmár et al., which showed that the diagnostic sensitivity of Giemsa in the absence of activity is only 33.6%, but it is 92.6% in the presence of active gastritis, which is close to the 99.4% sensitivity of IHC. Considering that chronic active gastritis with the features of H. pylori gastritis is also common in other entities, if active inflammation is present in the sample, there is a very small chance that a Giemsa-negative case will be confirmed as H. pylori-positive by IHC. Based on this, the use of IHC is more reasonable in Giemsa-negative cases with no activity in which the etiological role of H. pylori is suggested by clinical, anamnestic or other data. However, it may also be reasonable to routinely use IHC as the primary staining method instead of Giemsa.

Detection of Helicobacter pylori in oral squamous cell carcinoma using modified Giemsa stain: A new diagnostic approach.

Context: Oral squamous cell carcinoma (OSCC) is the sixth most common cancer worldwide. It is mainly known to be caused by tobacco in various forms and also due to viral, fungal infection and poor oral hygiene, etc. Poor oral hygiene leads to colonization of pathogenic bacteria including Helicobacter pylori. It seems that the presence of H. pylori might be a risk factor for developing oral cancer. The successful attempt was made to detect H. pylori in diagnosed specimens of OSCC using Warthin-starry and Giemsa stains in our department. The modified Giemsa stain is the method of choice because it is sensitive, cheap, easy to perform, faster, and reproducible. Aim: The aim of this study is to detect H. pylori in various grades of OSCC using modified Giemsa stain. Subjects and Methods: Thirty cases of various grades of OSCC were selected from the archives of the department. Five-micrometer-thick paraffin-embedded tissue sections of these cases were taken and stained with modified Giemsa and were studied under ×100 magnification. Results: All the tissue sections studied were positive for H. pylori bacteria. Conclusions: Our study showed a significant presence of H. pylori in histological sections of OSCC, and it seems likely that the presence of H. pylori might be a risk factor for the developing oral lesions such as oral cancer. Early detection and eradication of H. pylori in the oral cavity, especially in high-risk patients, might prevent its harmful consequences.

Modified Giemsa Stain: A solution to improve the quality of hypercellular bone marrow smears.

INTRODUCTION: Morphology is still the cornerstone of the diagnosis of haematological malignancies where the quality of staining plays a major role. The hypercellularity of a smear interferes with the quality of staining. We compared a newly modified giemsa stain (MGS) as a routine method for staining hypercellular bone marrow smears over the conventional Leishman method and the May Grunwald Giemsa method (MGG) in a completely randomized block study. METHODS: Quality of staining, cost, labour intensiveness and turnaround time of the new method was compared with MGG and Leishman methods for bone marrow smears of acute leukaemia using 30 cases. Bone marrow smears were blindly reviewed for the degree of staining of nuclei, cytoplasm and granules on a scale of 0 to 4 and statistically analysed by Minitab 16. RESULTS: A significant difference (p < .05) was revealed for the quality of staining of nuclei, cytoplasm and granules, and it was found that the new MGS was superior to the MGG and Leishman method. The cost of staining was lowest in the new MGS and the staining time of the new MGS was 76% less than MGG and 9.09% greater than the Leishman method. In terms of turnaround time, the new MGS is somewhat similar to the Leishman method and is user-friendly with a lesser number of steps to follow, compared with MGG. CONCLUSION: Our study showed that the new MGS is overall far superior, cost-effective and user-friendly compared with other conventional staining methods when used for hypercellular bone marrows.

Adult inclusion conjunctivitis diagnosed by polymerase chain reaction and Giemsa stain.

Adult inclusion conjunctivitis, caused by Chlamydia trachomatis, is easily underdiagnosed with nonspecific ocular manifestation. Combined scrape cytology and molecular testing may be a useful strategy for its early diagnosis. A 24-year-old healthy male complained of blurred vision, foreign body sensation, and watery discharge in his right eye for four weeks. His visual acuity was 20/20 bilaterally at his first visit. Allergic conjunctivitis was the first impression, and topical treatment with corticosteroid and anti-histamine was prescribed. However, he returned five days later without symptom improvement, and his right eye vision declined to 20/40. Subepithelial corneal infiltration of his right eye was observed. According to his personal history, his girlfriend was diagnosed with sexually transmitted chlamydial infection and genital gonorrhea. Under the suspicion of sexually transmitted adult inclusion conjunctivitis, we collected his conjunctival lavage to both real-time polymerase chain reaction, which proved chlamydial infection, and Giemsa stain, which demonstrated typical basophilic intracytoplasmic inclusions. To diagnose adult inclusion conjunctivitis, we can use real-time polymerase chain reaction or Giemsa stain to help us obtain a quick and correct diagnosis.

Post Covid-19 Infection Presenting as Rhino-Orbital Mycosis.

Rhino-orbital mycosis has been recently recognised as one of the sequelae in COVID-19 recovered patients. In India, detection of mucormycosis is declared as notifiable disease. In this article, the authors aim to describe the characteristics of patients presenting with post covid fungal infection which could be detected on 10% potassium hydroxide (KOH) wet mount and Giemsa stain put on crush biopsy smear. We describe 10 COVID-19 recovered patients admitted to ENT department of the hospital during second wave of COVID-19 infection. They presented with post covid fungal sinusitis and ophthalmic complications and planned for surgery. KOH mount and Giemsa stain were used for possible opinion and confirmed by culture. The observations were described in mean and percentages. All ten (100%) COVID-19 recovered patients were previously diagnosed with type 2 diabetes mellitus (DM) for 2-11 years. All 10 patients (100%) were given oral or intravenous corticosteroids for mean of 21 days (3 weeks-till presentation to ENT department). Simple procedures with 10% KOH mount and Giemsa stain could detect fungal hypae in all the cases and could provide possible opinion in 9 of 10 (90%) cases for timely management of the patients. The authors hypothesize that uncontrolled DM and prolonged use of corticosteroids may act as culprits of rhino-orbital mycosis in COVID-19 recovered patients. Simple and routine 10% KOH mount and Giemsa stain may provide early opinion of fungal hypae to ensure quick management and survival of the patients.