Gomori Methenamine Silver Stain Research Articles

How to make a fast diagnosis in invasive aspergillosis.

Aspergillosis is more common among immunocompromised patients with neutropenia or immunosuppression due to corticosteroid use, and infections are typically of the lung or sinuses. For diagnosis, broncholaveolar lavages (BALs) and lung biopsies are the specimens of choice. Culture and microscopic examinations are a must have and laboratory results should immediately be reported to the clinic. Fungal elements (hyphae) display the proof of an infection if present in primarily steril specimens, independent of culture results. Microscopy should be performed preferably using optical brighteners and histopathology using Gomori's methenamine silver stain or Periodic acid-Schiff. Serum and BAL galactomannan assays are recommended as markers for the diagnosis of invasive aspergillosis, PCR should be considered in conjunction with other diagnostic tests. Antifungal treatment decreases GM sensitivity. Pathogen identification to species complex level is strongly recommended for all clinically relevant Aspergillus isolates.

Histopathology of Hair Follicle Epithelium in Patients of Recurrent and Recalcitrant Dermatophytosis: A Diagnostic Cross-Sectional Study.

Context:Recurrent and recalcitrant dermatophytosis is a major evolving health problem in India. Histopathology is important in the diagnosis of recurrent and resistant cases, as it is postulated that hair follicle epithelium is affected, acting as the reservoir leading to recurrence and recalcitrance.Aims:This study aims to study histopathology of hair follicle epithelium in patients of recurrent and recalcitrant dermatophytosis.Settings and Design:A diagnostic cross-sectional study was performed at tertiary care hospital and referral center in central India.Materials and Methods:A 3 mm-sized punch skin biopsy was taken from 108 patients of recurrent or recalcitrant dermatophytosis involving hair follicles. Sections were stained by hematoxylin and eosin (H and E) stain in these patients followed by special stains such as periodic acid–Schiff (PAS) stain and Gomori methenamine silver (GMS) stain in 78 individuals where hair follicle was seen on H and E stain.Results:In H and E stain, surface epidermis fungus was observed in 57/108 patients (52.7%). In hair follicle-positive sections (78), surface fungus was seen in 52.6% with H and E stain, 84.6% with PAS stain, and 91% with GMS stain. H and E was 62.12% sensitive and 100% specific to diagnose fungus when compared with PAS stain and 57.7% sensitive and 100% specific when compared with GMS stain. PAS was 91.5% sensitive and 85.7% specific when compared with GMS stain. Ectothrix infection was the most common form of hair infection observed in 87.2% cases with GMS stain. Simultaneous presence of fungus in hair follicle and stratum corneum was observed in 87.3% of patients.Conclusion:Majority of patients showed affection of hair follicles by dermatophytes in clinically chronic, recurrent, and recalcitrant dermatophytic infections. GMS stain is the acceptable gold standard for detection of fungal elements.

Invasive aspergillosis of tympanomastoid cavity

Invasive aspergillosis of the tympanomastoid cavity is an extremely rare clinical incidence in comparison to the invasive fungal infections of the paranasal sinus and other parts of the body. Delay in the diagnosis and treatment of this rare clinical entity often leads to fatal complications such as skull base osteomyelitis and progressive multiple cranial nerve palsies. The invasive aspergillosis of the tympanomastoid region should be considered as a differential diagnosis for atypical mastoiditis in immunocompromised patients. Here, we report a case of invasive aspergillosis of the tympanomastoid region in a 60-year-old diabetic patient with facial nerve palsy. The diagnosis was done by the demonstration of fungi in the tissue with Gomori methenamine silver staining and confirmed with molecular diagnosis. Surgical debridement of the tympanomastoid region was done along with long-term voriconazole therapy.

Combination of clindamycin and primaquine for Pneumocystis pneumonia in renal transplant recipients

Pneumocystis pneumonia (PCP) in kidney transplant recipients is usually treated with trimethoprim-sulfamethoxazole (TMP-SMX). Combination of clindamycin and primaquine (C + P) is recommended for treatment failure. Data are scarce for treatment with C + P in renal transplant recipients. Here, we present three kidney transplant recipients on triple immunosuppression (tacrolimus, mycophenolate mofetil, and prednisolone) diagnosed to have PCP at different time periods after transplant surgery. Bronchoscopy and lavage were done; special staining with Gomori methenamine silver stain demonstrated Pneumocystis jirovecii. Two patients had graft dysfunction at the time of diagnosis. The first patient was started on TMP-SMX but developed leukopenia and was started on oral C + P. The other two cases had graft dysfunction and were started on C + P. All responded to therapy without complications, and improvement in graft function was observed. Clindamycin with primaquine can be considered as a safe and effective option to treat PCP in renal transplant recipients; this combination can replace TMP-SMX, as the first-line therapy, especially in patients with graft dysfunction or leukopenia

Cryptococcus‐like changes in the setting of vasculitis

Cutaneous vasculitis has many underlying causes, and the clinical and histological findings often overlap. Inflammatory vasculitis can mimic infection; however, distinction is critical for the timely institution of appropriate therapy. We present two patients who had generalized polymorphous eruptions whose cutaneous pathology showed vasculitis with unusual haloed yeast-like cells within the inflammatory infiltrate, mimicking Cryptococcus. The unusual cells stained negatively with Gomori methenamine silver and periodic acid-Schiff fungal stains, but positively for CD68 and had cytoplasmic reactivity with antibody to myeloperoxidase (MPO). Both patients had positive serum anti-MPO antibodies. The first patient experienced a rapidly fatal course, whereas the second patient improved with prompt initiation of systemic corticosteroids. Interestingly, the second case had prior biopsy showing Sweet syndrome with crypotoccoid-appearing cells. Cryptococcoid cells have been described previously in association with neutrophilic dermatoses, but not in the setting of vasculitis as was seen in our patients. Our cases add to the existing literature on crypotoccoid mimickers, and are the first to be reported in association with vasculitis.

356. Bronchoalveolar Lavage Fluid Cytology by GMS Stain for the Diagnosis of Invasive Pulmonary Aspergillosis in Patients With Hematologic Malignancies: Analysis of 67 Episodes

BackgroundThe yield of direct fungal visualization by GMS (Gomori–methenamine–silver) stain in bronchoalveolar lavage (BAL) cytology has rarely been studied in the diagnosis of invasive pulmonary aspergillosis (IPA) in patients with hematological malignancies (HM).MethodsWe analysed a series of patients with proven or probable culture-documented IPA (EORTC/MSG criteria) in HM patients (1999–2015). All patients had BAL cultures that were positive for Aspergillus spp. and had concurrently obtained BAL cytology GMS available for analysis.ResultsWe identified 67 such patients. BAL cytology based on GMS showed hyalohyphomycetes consistent with Aspergillus in 28/67 (41.8%) patients, whereas only in 2/67 (3.6%) direct smear Calcofluor White stain was positive. Based on BAL GMS cytology, co-infections were identified in six patients: two Pneumocystis and five viral infections with cytopathic changes (one had both). The yield of cytology was not different in patients with IPA caused by non-fumigatus Aspergillus, although patients with IPA and >1 Aspergillus in BAL culture had more often positive cytology GMS (100% vs. 0%, p 0.027). Cytology was also more often positive when obtained from a lesion-targeted BAL as compared with non-targeted bronchial washings (60.7% vs. 7.1%, p 0.038). Patients with IPA and cavitary lesions (32.1% vs. 5.1%, P = 0.006), history of SCT (64.3% vs. 33%, P = 0.015) or prior exposure to itraconazole (75% vs. 41%, P = 0.007) had positive cytology GMS results more often than did patients without these characteristics. In the multivariate analysis, only cavitary lesions were significantly associated with positive BAL GMS cytology.ConclusionGMS stain in cytology of BAL in patients with HM and culture-documented IPA had a sensitivity of 41.8% and was more often positive in patients with cavitary lesions. Although there were no differences in the proportion of GMS-positive cytology rates among differing Aspergillus spp. causing IPA, mixed Aspergillus spp. IPA was associated with an increase in positive cytology. BAL cytology was diagnostic for co-infections in more than 10% of patients. BAL cytology should be part of the diagnostic wok up in HM patients with suspected IPA.Disclosures All authors: No reported disclosures.

2566. Diagnosis of Pneumocystis jiroveci Pneumonia in HIV-Negative Immunocompromised Patients: Is the Gomori-Methenamine Silver Stain of Bronchoalveolar Lavage Fluid the Gold Standard or Sub-Standard?

BackgroundDirect visualization of Pneumocystis jiroveci on bronchoalveolar lavage (BAL) fluid using the Gomori-methenamine-silver (GMS) stain historically has been the mainstay of diagnosis for P. jiroveci pneumonia (PJP), with studies from the early HIV/AIDS era reporting sensitivities of 90–95%. However, the burden of P. jiroveci organisms in BAL fluid is significantly lower in HIV-negative immunocompromised patients compared with HIV-positive patients with PJP, raising concerns that the BAL GMS stain is less sensitive in this population.MethodsWe conducted a retrospective observational study at Yale-New Haven Hospital from 2012 to 2018, using electronic medical record chart reviews, to identify patients who underwent bronchoscopy with BAL GMS stain for the diagnosis of PJP. We collected additional patient factors such as age, sex, HIV status, and immunosuppressed status. For patients with a negative BAL GMS stain, we collected data on other diagnostics, including positive GMS lung biopsies, positive PJP PCR or DFA, and elevated serum (1–3)-β-d-glucan levels. We defined BAL GMS-negative cases as proven or probable based on investigator generated criteria (see Figure 1).ResultsWe identified 52 patients with PJP who received a BAL GMS stain, including 28 HIV-positive and 24 HIV-negative cases. Of 24 HIV-negative cases, 11 had BAL GMS-positive PJP and 13 had BAL GMS-negative PJP (9 proven and 4 probable). In the latter group, 6 had hematologic malignancies (HM), 2 had solid-organ transplants (SOT), 1 had hematopoietic stem cell transplant, 2 had SOT plus HM, and 2 received high-dose steroids. Proven diagnoses were made by GMS-positive lung biopsy (n = 6), DFA (n = 2), and PCR (n = 1). Elevated (1–3)-β-d-glucan was observed in 7 of 8 cases (median: >500 pg/mL; range 39 to >500). Three patients developed adverse outcomes (1 readmission due to untreated PJP and 2 treatment delays). BAL GMS sensitivity for HIV-negative patients was 11/24 (46%) vs. 28/28 (100%) in HIV-positive patients.ConclusionThe sensitivity of BAL GMS for PJP is poor in HIV-negative immunocompromised patients. Missed cases or delayed treatment for PJP may lead to adverse outcomes. In HIV-negative patients with a clinical syndrome compatible with PJP, a negative BAL GMS does not rule out PJP and must be confirmed by supplementary diagnostics.Figure 1Disclosures All authors: No reported disclosures.

Extrapulmonary Hepatic Histoplasmosis Presenting as Chronic Liver Disease: A Case Report

Introduction: Histoplasma capsulatum is the most common endemic mycosis in the United States. Extrapulmonary hepatic histoplasmosis as a primary manifestation is extremely rare. We report a case of histoplasmosis that presented like chronic liver disease with portal hypertension, splenomegaly, esophageal varices, and thrombocytopenia in an immunocompetent adult. Case Description: A 56 year-old Caucasian man from West Virginia who works in construction presented to the hepatology clinic for work up of chronic liver disease. He was initially evaluated by his primary care doctor for complaints of diarrhea and had an EGD performed that revealed esophageal varices. On examination, he was noted to have splenomegaly but no other stigmata of chronic liver disease. A CBC was only remarkable for a platelet count of 86. A CMP was unremarkable with normal liver enzymes. INR was 1.0. Serological work up for liver disease including ceruloplasmin, remote hepatitis panel, iron studies, anti-nuclear antibody, anti-mitochondrial antibody, anti-smooth muscle antibody, and alpha 1 anti-trypsin were normal. A RUQ ultrasound with elastography showed a normal, homogenous liver, splenomegaly and a liver stiffness of F2-F3 indicating moderate fibrosis. EGD was repeated which showed large (>5mm) esophageal varices that were banded. It also revealed moderate portal hypertensive gastropathy. Transjugular liver biopsy was performed that revealed bridging fibrosis with minimal steatosis, no obvious granulomas but loose clusters of histiocytes, and a gomori methenamine silver stain was performed looking for histoplasmosis that highlighted small yeast with budding typical for Histoplasma capsulatum. Histoplasma antibody in the blood was positive. The patient was seen by infectious disease and he was started on oral itraconazole. Discussion: Hepatic histoplasmosis can present as granulomatous hepatitis, as stigmata of portal hypertension, including ascites or varices, or with elevated liver enzymes. The most common hepatic finding on liver biopsy is portal lymphohistiocytic inflammation; discrete hepatic granulomas are seen in less than 20% of involved livers. There are only a few case reports of disseminated histoplasmosis presenting as primary hepatic histoplasmosis. A high clinical suspicion is warranted in patients who present with stigmata of portal hypertension and chronic liver disease with no evidence of cirrhosis, especially if the person lives in or has visited an endemic area.2252 Figure 1. The normal lobular architecture is distorted by bridging fibrosis. The hepatocytes show minimal steatosis. Although well-developed granulomas are not identified, there are loose clusters of histiocytes.

Bronchoalveolar Lavage Fluid Cytology in Culture-Documented Invasive Pulmonary Aspergillosis in Patients with Hematologic Diseases: Analysis of 67 Episodes

There is a paucity of studies on the yield of Gomori-methenamine-silver (GMS) staining in bronchoalveolar lavage (BAL) fluid cytology and its comparison with fluorescent dye staining for the diagnosis of invasive pulmonary aspergillosis (IPA) in patients with hematologic malignancies. To that end, we analyzed the yield of direct fungal visualization in BAL fluid cytology with GMS staining, in a series of culture-positive IPA cases in 67 patients with hematologic malignancies, and we compared the results with those of direct examination with calcofluor white staining and BAL fluid galactomannan assays, when available. GMS staining in BAL fluid cytology was positive in 42% of the 67 cases and revealed coinfections in 7 cases. In contrast, only 2/67 (3.6%) BAL fluid samples were positive in direct smears stained with the fluorescent dye calcofluor white. Positive GMS staining results were significantly more frequent in IPA cases with cavitary lesions and IPA cases caused by >1 Aspergillus species, but the proportions of positive cytology results among Aspergillus species were not different.

Diagnostic values of KOH examination, histological examination, and culture for onychomycosis: a latent class analysis.

In the absence of a real gold standard, comparative studies are still done on diagnostic methods for onychomycosis. There are only a few attempts using latent class analysis to determine the value of polymerase chain reaction in comparison to conventional methods. We aimed to determine the value of histological examination in such a way for the diagnosis of onychomycosis. Potassium hydroxide mount (KOH), culture and histological examination with periodic acid-Schiff (PAS), and Gomori's methenamine silver (GMS) stains were done in 106 patients having clinically suspected toenail onychomycosis. KOH was positive in 74% of the patients; culture in 14%; PAS in 30%; and GMS in 66%. According to the results of the latent class analysis, culture and PAS were highly specific but poorly sensitive; KOH, highly sensitive but poorly specific; and GMS, both highly sensitive and specific. Based on these results, we have proposed KOH as a screening test and GMS as a confirmatory test for the diagnosis of onychomycosis in our own practice. However, since positivity rates of different diagnostic methods vary widely in different centers, it is more suitable that every center should determine their own diagnostic strategy by evaluating their own results with latent class analysis.

Disseminated Histoplasmosis Presenting as Bilateral Adrenal Masses in an Immunocompetent Adult: Report of an Unusual Case Presentation

Bilateral adrenal involvement in disseminated histoplasmosis in immunocompetent adults is extremely rare. This is a case of disseminated histoplasmosis with an unusual presentation as bilateral adrenal masses and associated retroperitoneal lymphadenopathy, mimicking a metastatic malignancy. A 40-year-old male with a medical history of diabetes mellitus type 2, hypertension, chronic backache, and tobacco use presented with progressive fatigue, anorexia, night sweats, persistent abdominal pain, and 160 pounds of unintentional weight loss over the past 6 months. A laboratory workup, including HIV and tuberculosis testing, was negative. Urine and blood cultures failed to grow microorganisms. CT scan of the abdomen revealed bilateral adrenal masses and retroperitoneal lymphadenopathy. A subsequent adrenal endocrine profile was normal. The radiologic and clinical findings were highly suspicious for malignancy. A transabdominal CT-guided fine-needle aspiration of the right adrenal mass revealed numerous yeast forms, which were highlighted by a Gomori methenamine silver stain. The microorganisms, seen both intracellularly within macrophages as well as extracellularly, were 2 to 4 μm in size and displayed narrow-based budding. A concurrent biopsy from the adrenal mass showed morphologically similar fungal organisms, within an extensively necrotic granulomatous inflammatory background. Fungal cultures confirmed H capsulatum. Further investigations revealed that the patient worked on a tug boat that often traversed the Mississippi River valley. Treatment with systemic amphotericin B and itraconazole resulted in marked clinical and radiological improvement. This is an unusual case of disseminated histoplasmosis simulating metastatic malignancy to bilateral adrenal glands. In our case, the diabetes mellitus may have contributed as an immune deregulatory state in an otherwise immunocompetent individual. In conclusion, disseminated histoplasmosis may present as bilateral adrenal masses in an otherwise immunocompetent host, simulating a metastatic malignant process. Histoplasmosis should be considered in the differential diagnosis in such settings, especially in endemic areas.

Do Pentraxins Bind to Fungi in Invasive Human Gastrointestinal Candidiasis?

Tissue from 13 autopsy cases with invasive gastrointestinal candidiasis was studied for the binding of the pentraxins, C-reactive protein (CRP), pentraxin 3 (PTX3), and serum amyloid P component (SAP) to fungal surfaces. Invasive candidal infection was demonstrated using a hematoxylin and eosin stain and a Gomori methenamine silver stain (GMS). Immunohistochemistry was performed with CRP and PTX3 monoclonal antibodies and did not demonstrate CRP or PTX3 bound to fungi (0 of 13 cases), although CRP was extensively deposited on human tissue. A polyclonal antibody to SAP showed that SAP was bound to fungi in 12 of 13 cases. Although all three pentraxins have been reported to bind to fungi or bacteria, only SAP was bound to filamentous and yeast forms of Candida in human tissue, as detected by immunohistochemistry. SAP was abundantly present on fungi and may have affected the host innate immune response to the invading fungi.

Colonization of Pneumocystis jirovecii in Patients who Received and not Received Corticosteroids Admitted to the Intensive Care Unit: Airborne Transmission Approach

Pneumocystis pneumonia (PCP) is responsible for pulmonary infection in immunocompromised patients. The current study aimed at investigating the frequency of Pneumocystis colonization in patients hospitalized in the intensive care unit (ICU) and evaluating the relationship between PCP and Pneumocystis colonization. The current cross sectional study was conducted on bronchoalveolar lavage (BAL) fluids of 100patients collected from surgery and neurosurgery ICUs with different underlying corticosteroid therapy conditions. Patients were divided into 2 groups (patients receiving and not receiving corticosteroids). Direct examination on BAL fluids was performed by the Gomori methenamine silver and Giemsa staining techniques. Additionally, 2 filtered air samples of the 2 above mentioned units were collected. A nested-PCR targeted mtLSUrRNA gene and sequencing were used to identify Pneumocystis spp. In direct microscopy, 31 out of 100 hospitalized patients (31%) showed positive results. Twenty-three (46%) of smear positive patients were from the group of patients receiving corticosteroid, the other 8(16%) were from the group of patients not receiving corticosteroids (P= 0.001). Pneumocystis jirovecii DNA was detected in 77 out of 100 BAL samples by nested-PCR (77%) in which 40 (52%) and 37 (48%) samples were obtained from the patients receiving and not receiving corticosteroids, respectively. Pneumocystis genome was observed in 1 of the 2 filtered air samples. A significant number of patients receiving corticosteroids were also colonized by P. jirovecii that may be predisposed to PCP or be transmitted to susceptible patients. A significant relationship was observed between the mean hospital stay and detection rate.

Liquid-Based Cytology of the Cerebrospinal Fluid in a Case of Cryptococcal Meningitis.

Cryptococcus neoformans is the most common microorganism found in cerebrospinal fluid (CSF) cytology and causes life-threatening infections in immunocompromised hosts. Although its cytomorphologic features in conventional smear cytology have been well described, those in liquid-based cytology have rarely been. A 73-year-old woman with diffuse large B-cell lymphoma presented with mental confusion and a spiking fever. To rule out infectious conditions, CSF examination was performed. A cytology slide that was prepared using the ThinPrep method showed numerous spherical yeast-form organisms with diameters of 4–11 μm and thick capsules. Occasional asymmetrical, narrow-based budding but no true hyphae or pseudohyphae were observed. Gomori methenamine silver staining was positive. Cryptococcosis was confirmed in blood and CSF through the cryptococcal antigen test and culture. Liquid-based cytology allows for a clean background and additional slides for ancillary testing, facilitating the detection of microorganisms in CSF specimens, particularly when the number of organisms is small.

A Rare Case of Gastrointestinal Histoplasmosis in an Immunocompromised Host

Introduction: Histoplasmosis capsulatum (HC) is a dimorphic fungus common to the Ohio and Mississippi River valleys and is found mainly in soil enriched by avian or bat droppings. It typically presents as an asymptomatic pulmonary infection. Gastrointestinal (GI) involvement is usually unrecognized due to nonspecific symptoms and is thought to be symptomatic in only 3 to 12% of patients. The majority of GI histoplasmosis involves patients with HIV/AIDS. The most common site of GI involvement is the ileocecal region due to the presence of abundant lymphoid tissue. Case Presentation: A 64-year-old man with paroxysmal atrial fibrillation (on warfarin), end stage renal disease on hemodialysis, status post failed transplanted kidney and on chronic immunosuppression presented with a one day history of hematochezia and hematemesis and six weeks of worsening left lower quadrant abdominal pain. CT thorax with intravenous contrast revealed a right lower lobe cavitary mass and a spiculated left upper lobe nodule (Figure 1). Esophagogastroduodenoscopy with enteroscopy revealed old blood in stomach and the endoscope was advanced into the jejunum revealing multiple ulcers (Figure 2). Histopathology from jejunal biopsies was consistent with HC (Figure 3). Histoplasmosis antibodies by complement fixation for yeast and mycelia were positive as was fungal immunodiffusion for Histoplasma M antigen. Histoplasmosis serum antigen was negative. The patient was treated with intravenous amphotericin for presumed disseminated histoplasmosis. He improved clinically with resolution of symptoms and was discharged on oral itraconazole and remains therapeutic five months later.Figure: CT thorax with intravenous contrast showing (A) 21mm right lower lobe cavitary mass (B) and 19mm left upper lobe spiculated nodule.Figure: Ulcerations of proximal (A) and mid-jejunum (B).Figure: Gomori methenamine silver (GMS) stain of jejunal biopsy specimen showing budding yeast form of histoplasmosis capsulatum.Discussion: Histoplasmosis capsulatum is generally seen in immunocompromised hosts. While common at autopsy, gastrointestinal histoplasmosis (GIH) goes unrecognized during life as the symptoms are nonspecific. Patients with GIH most commonly present with abdominal pain and diarrhea, but may also have dysphagia, odynophagia, bowel obstruction or perforation. In this case, the patient presented with clinically significant gastrointestinal bleeding. Upper endoscopy with jejunal biopsies revealed HC. Both the presence of symptomatic GIH and the jejunal location are atypical. This observation in a patient without HIV is exceedingly rare.

Acid-Fastness of Histoplasma in Surgical Pathology Practice.

BackgroundHistoplasmosis (HP) is diagnosed by visualizing intracellular microorganisms in biopsy and/or culture. Periodic-acid Schiff (PAS) and Gomori methenamine silver (GMS) staining methods are routinely used for identification. The acid-fast property of Histoplasma was identified decades ago, but acid-fast staining has not been practiced in current surgical pathology. Awareness of the acid-fast property of Histoplasma, which is due to mycolic acid in the cell wall, is important in distinguishing Histoplasma from other infective microorganisms. Here, we examined acid-fastness in previously diagnosed cases of Histoplasma using the Ziehl-Neelsen (ZN) stain and correlated those findings with other known fungal stains.MethodsAll cases diagnosed as HP were retrieved and reviewed along with ZN staining and other fungal stains. We also stained cases diagnosed with Cryptococcus and Leishmania as controls for comparison.ResultsA total of 54 patients ranging in age from 11 to 69 years were examined. The most common sites of infection were the skin, adrenal tissue, and respiratory tract. Of the total 43 tissue samples, 20 (46.5%) stained positive with the ZN stain. In viable cases, a significant proportion of microorganisms were positive while necrotic cases showed only rare ZN-positive yeasts. In comparison to PAS and GMS stains, there was a low burden of ZN-positive yeasts. Cryptococcus showed characteristic ZN staining and all cases of Leishmania were negative.ConclusionsAlthough the morphology of fungal organisms is the foundation of identification, surgical pathologists should be aware of the acid-fast property of fungi, particularly when there is the potential for confusion with other infective organisms.

Pythium Insidiosum Keratitis: Histopathology and Rapid Novel Diagnostic Staining Technique.

To elucidate the histopathology of Pythium insidiosum keratitis and to describe a novel, simple, and rapid staining technique for identification of oomycete Pythium insidiosum and to differentiate it from fungi. This is a laboratory investigation study of 38 nonconsecutive cases (37 ocular samples and 1 colonic biopsy); 14 microbiologically diagnosed as Pythium insidiosum keratitis and 24 as fungal keratitis. Review of clinical, demographic details, microbiological results, and identification of cases that necessitated evisceration was performed. Reevaluation of histopathology slides was done using stains such as hematoxylin-eosin, Gomori methenamine silver (GMS), periodic acid-Schiff (PAS), potassium iodide-sulfuric acid (IKI-H2SO4). Morphology, degree, and nature of inflammation and load, distribution, and staining results of Pythium insidiosum and its comparison with fungi were studied. Delay in zoospore formation, failure of growth, and delay in identification of Pythium were the main cause of evisceration. Corneal pythiosis showed epithelial ulceration, stromal destruction, and varying inflammation; load and distribution of Pythium were inversely proportional to inflammation. The filaments were commonly wide, with admixed narrower structures and uncommonly involved Descemet membrane. The oomycete was not discretely discerned with PAS stain and stained distinctly with GMS stain and IKI-H2SO4 stain (100% sensitive). In comparison, fungal organisms stained well with PAS and GMS stain, but not with IKI-H2SO4 stain (100% specific). Pythium insidiosum keratitis is perhaps not more devastating than fungal keratitis but late diagnosis, misdiagnosis, and treatment as fungal infection are major heralds. Early diagnosis may markedly improve the patient outcome. IKI-H2SO4 is a cost-effective, simple, sensitive, and specific stain for the diagnosis of oomycete Pythium.

Histopathological techniques for the diagnosis of combat-related invasive fungal wound infections.

BackgroundEffective management of trauma-related invasive fungal wound infections (IFIs) depends on early diagnosis and timely initiation of treatment. We evaluated the utility of routine staining, histochemical stains and frozen section for fungal element identification.MethodsA total of 383 histopathological specimens collected from 66 combat-injured United States military personnel with IFIs were independently reviewed by two pathologists. Both periodic acid-Schiff (PAS) and Gomori methenamine silver (GMS) stains were used on 74 specimens. The performance of the two special stains was compared against the finding of fungal elements via any histopathological method (ie, special stains or hematoxylin and eosin). In addition, the findings from frozen sections were compared against permanent sections.ResultsThe GMS and PAS results were 84 % concordant (95 % confidence interval: 70 to 97 %). The false negative rate of fungal detection was 15 % for GMS and 44 % for PAS, suggesting that GMS was more sensitive; however, neither stain was statistically significantly superior for identifying fungal elements (p = 0.38). Moreover, 147 specimens had frozen sections performed, of which there was 87 % correlation with permanent sections (60 % sensitivity and 98 % specificity). In 27 permanent sections, corresponding cultures were available for comparison and 85 % concordance in general species identification was reported.ConclusionsThe use of both stains does not have an added benefit for identifying fungal elements. Furthermore, while the high specificity of frozen section may aid in timely IFI diagnoses, it should not be used as a stand-alone method to guide therapy due to its low sensitivity.

Comparison of three cost effective staining methods for detection of Pneumocystis jirovecii.

Pneumocystis pneumonia due to Pneumocystis jirovecii infection is an emerging health problem not only for HIV-infected patients but also for other immunocompromised patients in many countries. We compared Gomori methenamine silver (GMS), Toluidine Blue O (TBO) and Giemsa staining methods using standard procedures. The sensitivity and specificity of GMS were 100%. The sensitivity and specificity of TBO were 96 and 100%, respectively. The sensitivity and specificity of Giemsa stain were 84 and 90%, respectively. Only GMS had positive and negative predictive values of 100% while PPV and NPV for TBO were 100 and 90.9%, and for Giemsa stain were 95.4 and 69.2%, respectively. Therefore, our results suggest that if TBO or Geimsa stains are used as the primary staining methods in a clinical laboratory, then confirmation with a GMS staining method should be performed to increase the sensitivity and specificity of the final test result.

Pituitary aspergillus infection

Fungal infection should be considered in the differential diagnosis of a pituitary or sellar mass, albeit fungal infections involving the pituitary gland and sella are a rare occurrence. We report a case of Aspergillus infection involving the pituitary gland and sellar region discovered in a 74-year-old man. The patient had a history of hypertension, chronic renal disease, autoimmune hemolytic anemia and presented with right eye pain, headaches and worsening hemiparesis. Imaging studies revealed a right internal carotid artery occlusion and an acute right pontine stroke along with smaller infarcts in the right middle cerebral artery distribution. Clinically, the patient was thought to have vasculitis. An infectious etiology was not identified. He developed respiratory distress and died. At autopsy, necrotizing meningitis was discovered. A predominantly chronic inflammatory cell infiltrate consisting of benign-appearing lymphocytes, plasma cells and macrophages was accompanied by acute angle branching, angioinvasive hyphae which were highlighted on Gomori methenamine silver staining and were morphologically consistent with Aspergillus species. In previously reported cases of Aspergillus infection involving the pituitary or sella, most presented with headaches or impaired vision and were not immunocompromised. A transsphenoidal surgical approach is recommended in suspected cases in order to minimize the risk of dissemination of the infection. Some patients have responded well to antifungal medications once diagnosed.