Gomori Methenamine Silver Stain Research Articles

Spectrum of orbital fat necrosis in rhino-orbital-cerebral mucormycosis in post-COVID-19 patients.

Rhino-orbital-cerebral mucormycosis (ROCM) is the most common presentation of mucormycosis. Sinonasal-orbital debridement with exenteration is a life-saving procedure in ROCM patients in view of severe involvement of sinuses and the eyeball. Following the second wave of coronavirus disease 2019 (COVID-19), there was a massive surge in mucormycosis cases in India in post-COVID-19 patients. Of over 300 cases of mucormycosis admitted in our hospital, many underwent exenteration and these specimens were evaluated histopathologically, where fat necrosis was found to be a prominent finding. The spectrum of fat necrosis in ROCM in orbital fat has not been described in literature. Hence, we sought to evaluate the significance and spectrum of orbital fat necrosis in ROCM. This 3-month retrospective study included 15 cases of ROCM which underwent exenteration. Clinical data, radiologic details, and histopathologic findings were tabulated. Sections were also subjected to Periodic acid Schiff (PAS) and Gomori's methenamine silver (GMS) stains for confirming the fungus. All 15 cases showed fat stranding on computed tomography (CT) scan. On histopathologic examination, various tissue reaction patterns observed included acute/chronic inflammatory infiltrate, suppurative granulomas with giant cells, coagulative and fat necrosis, broad aseptate fungal hyphae with or without angioinvasion, and neural invasion. Fungal hyphae were confirmed with PAS and GMS stains. The spectrum of fat necrosis observed in all the cases included 1) acute necrotizing fat necrosis, 2) ghost adipocytes with or without saponification, and 3) crystalline/gouty fat necrosis. Fat necrosis is a significant finding in ROCM, both on CT scan and histopathology. All three patterns of fat necrosis may be observed simultaneously in a case of ROCM.

A decision support system for the detection of cutaneous fungal infections using artificial intelligence

Cutaneous fungal infections are one of the most common skin conditions, hence, the burden of determining fungal elements upon microscopic examination with periodic acid-Schiff (PAS) and Gomori methenamine silver (GMS) stains, is very time consuming. Despite some morphological variability posing challenges to training artificial intelligence (AI)-based solutions, these structures are favored potential targets, enabling the recruitment of promising AI-based technologies. Herein, we present a novel AI solution for identifying skin fungal infections, potentially providing a decision support system for pathologists. Skin biopsies of patients diagnosed with a cutaneous fungal infection at the Sheba Medical Center, Israel between 2014 and 2023, were used. Samples were stained with PAS and GMS and digitized by the Philips IntelliSite scanner. DeePathology® STUDIO fungal elements were annotated and deemed as ground truth data after an overall revision by two specialist pathologists. Subsequently, they were used to create an AI-based solution, which has been further validated in other regions of interests. The study participants were divided into two cohorts. In the first cohort, the overall sensitivity of the algorithm was 0.8, specificity 0.97, F1 score 0.78; in the second, the overall sensitivity of the algorithm was 0.93, specificity 0.99, F1 score 0.95. The results obtained are encouraging as proof of concept for an AI-based fungi detection algorithm. DeePathology® STUDIO can be employed as a decision support system for pathologists when diagnosing a cutaneous fungal infection using PAS and GMS stains, thereby, saving time and money.

Unveiling the Uncommon: Gastrointestinal Basidiobolomycosis of the Colon and its Distinctive Splendore-Hoeppli Phenomenon

Background Basidiobolomycosis is a rare fungal infection caused by Basidiobolus ranarum. Case Presentation A 53-year-old man from Saudi Arabia with a known history of diverticulosis presented with severe abdominal pain and diarrhea. A CT scan revealed circumferential wall thickening of the descending and sigmoid colon with surrounding fat stranding, suggesting a diagnosis of complicated diverticulitis. Additional thick fluid was observed around the affected area. Surgical excision was pursued. A gross examination of two received large bowel segments disclosed marked ulcerated mucosa and wall thickening with exudate-covered serosal surfaces and adhesions. Microscopic examination unveiled significant infiltration by eosinophils, polymorphonuclear leukocytes, and granulomatous inflammation. Thin-walled, broad fungal hyphae of Basidiobolus, surrounded by eosinophilic material, were identified. Granulomas displayed abundant multinucleated giant cells and palisading histiocytes around central necrosis or abscess formation. Thin-walled, broad fungal hyphae of Basidiobolus, with sparse septations, are surrounded by a radiating, intensely eosinophilic cuff (Splendore-Hoeppli phenomenon). These hyphae, visible with hematoxylin and eosin staining, were further highlighted with periodic acid-Schiff and Gomori methenamine silver staining. Discussion Basidiobolomycosis may mimic neoplastic lesions. Histologically, the characteristic features include broad, thin-walled septate hyphae surrounded by eosinophilic material, a finding that is accentuated by the Splendore-Hoeppli phenomenon. Microscopic examination, along with special stains such as periodic acid-Schiff (PAS) and Gomori methenamine silver, is essential for accurate diagnosis. Conclusion Prompt recognition and appropriate antifungal therapy are vital for favorable patient outcomes. This report highlights the distinctive features of Basidiobolomycosis to raise awareness and understanding of this infrequent yet clinically significant fungal infection.

Utaneous verrucose phaeohyphomycosis due to Exophiala oligosperma in an immunocompetent host

ABSTRACT Herein, we report a case of subcutaneous infection caused by Exophiala oligosperma. A verrucous plaque was the major clinical feature. A histopathological examination revealed features of suppurative granuloma. Mycological and molecular identification revealed E. oligosperma as the etiologic agent. After eight months of treatment, the lesion showed worsening due to the use of self-medication with topical hydrocortisone. A second biopsy was performed with direct examination (KOH 10%) showing dark pigmented septate hyphae, yeast and multiple dark hyphae were highlighted in biopsy with Gomori methenamine silver (GMS) staining. We report this case to show the rise of this infection in an immunocompetent patient and the effects of steroids. The patient was treated with itraconazole and cryosurgery. Key words: Phaeohyphomycosis, Cutaneous, Verrucose, Exophiala oligosperma, Melanized fungi, Immunocompetent host, Itraconazole, Cryosurgery, Chromoblastomycosis

Histopathological spectrum of fungal infections in COVID 19 cases – A retrospective observational study

A wide range of fungal and bacterial infections are associated with Corona virus disease (COVID19). There was a surge of mucormycosis during COVID19 pandemic in 2021. This study elaborates the histopathological findings observed in fungal infections of head and neck in COVID 19 patients. A retrospective observational study of histopathological findings of invasive fungal infection in COVID 19 case was conducted in our laboratory. All the tissue samples diagnosed as post COVID fungal infections during January 2021 to June 2021 were analyzed. Representative sections were taken and slides were stained with Hematoxylin & Eosin, Periodic Acidic Schiff (PAS) and Gomori Methenamine silver (GMS) stains. Histopathological features were studied. There were total 32 cases which had invasive fungal infections associated with COVID 19. The most affected age group was 31 to 50 years of age (50%), with male preponderance (87.5%). Diabetes was present in 62.5% cases, and steroid use during the treatment was present in 31.3% of the cases. The most common site was maxillary sinus (53.1%) followed by nasal cavity (34.4%). Isolated infection by mucor was seen in 68.8% cases, there were cases of secondary bacterial infection and candida with aspergillus infection. Angioinvasion was present in 56.3% cases and bone destruction in 15.6% cases.COVID 19 infection results in immune dysregulation and background of diabetes and over use of steroids increases risk of mucormycosis. An early histopathological diagnosis provides early diagnosis.

Histochemical Demonstration of Fungi in Breast Diseases at University of Maiduguri Teaching Hospital-North Eastern Nigeria

Breast disease is a global public health concern, and recent research suggests that fungal infections may be linked to its development and progression. This study aims to investigate the presence of fungi in formalin-fixed paraffinembedded (FFPE) breast tissue samples diagnosed with various breast diseases, including breast cancer, using histological staining techniques. The study design was a retrospective cross-sectional study that used convenience sampling to retrieve 49 breast tissue blocks diagnosed with various breast diseases from the Histopathology archive of the University of Maiduguri Teaching Hospital, Nigeria. The samples were re-embedded with paraffin wax, sectioned, and stained using Haematoxylin and Eosin and Grocott Methanamine Silver Stain. The data collected was analyzed descriptively and using chi-square tests. Fibroadenoma was the most common diagnosis in the study (69.3%), followed by Chronic Granulomatous Mastitis (18.4%) and breast cancer (12.2%). The highest frequency of Fibroadenoma diagnosis was in the age range of 30-39, whereas the highest frequency of breast cancer diagnosis was in the age range of 20-29. The study found that fungal infections were present in breast tissue samples, and the highest percentage of positive Gomori Methenamine Silver (GMS) stain test results were found in breast disease cases with breast cancer with a significant p value of 0.0175. This study provides evidence of a potential association between fungal infections and breast diseases in North Eastern Nigeria. Further research is needed to fully understand the mechanisms underlying this association and its implications for breast cancer prevention and treatment.

Diagnostic value of metagenomic next-generation sequencing of lower respiratory tract specimen for the diagnosis of suspected Pneumocystis jirovecii pneumonia

Aim To evaluate diagnostic performance of metagenomic next-generation sequencing (mNGS) for Pneumocystis jirovecii pneumonia (PCP), in comparison with polymerase chain reaction (PCR), Gomori methenamine silver (GMS) staining and serum 1,3-β-d-Glucan (BG) assay. Methods 52 PCP patients and 103 patients with non-pneumocystic jirovecii pneumonia (non-PCP) were enrolled, and comparative analysis was conducted of different diagnostic tests. Clinical features and co-pathogen characteristics were reviewed. Results The diagnostic sensitivity (92.3%) and specificity (87.4%) of mNGS did not show significant differences compared with that of PCR while mNGS had the advantage over PCR in the detection of co-pathogens. Despite its excellent specificity, the sensitivity of GMS staining (9.3%) was inferior to that of mNGS (p < .001). The combination of mNGS with serum BG statistically outperformed mNGS or serum BG alone in the areas under the receiver operating characteristic curves (AUCs, p = .0013 and p = .0015, respectively). Notably, all the blood samples showing positive mNGS for Pneumocystis jirovecii came from PCP patients. The leading co-pathogens among patients with PCP were cytomegalovirus, Epstein-Barr virus and Torque teno virus. Conclusions mNGS shows superiority over several common clinical methods in the diagnosis of suspected PCP. Serum BG in conjunction with mNGS further improved the diagnostic efficacy of mNGS.

Detection of Pneumocystis jirovecii by PCR in patients with lung cancer: A preliminary study.

Infection complications in lung cancer (LC), one of the most common cancers in the world, are still among the most important causes of death. Of them, P. jirovecii, which is as an opportunistic infection, causes a life-threatening type of pneumonia in cancer patients. This preliminary study aimed to determine the incidence and clinical status of P. jirovecii by PCR in lung cancer patients compared to the conventional method. Sixty-nine lung cancer patients and fSorty healthy individuals were included in the study. After sociodemographical and clinical features were recorded, sputum samples were collected from attenders. Firstly, microscopic examination was made with Gomori's methenamine silver stain and then PCR was performed. P. jirovecii was detected in three of 69 lung cancer patients by PCR (4.3%), but not by microscopy. However, healthy individuals were negative for P. jirovecii by both methods. Based on clinical and radiological findings, P. jirovecii was evaluated as probable infection in one patient and colonization in the other two patients. Although PCR is more sensitive than conventional staining methods, it cannot distinguish probable and proven infections from pulmonary colonization. It is important to evaluate the decision of infection together with laboratory, clinical and radiological findings. Moreover, PCR may enable to know the colonization and to take precautions such as prophylaxis, due to the risk of colonization turning into an infection in immunocompromised patient groups. Further studies involving larger populations and evaluating the colonization-infection relationship in patients with solid tumors are needed.

Unilateral conjunctival concretions associated with chronic inflammation and tear instability.

Conjunctival concretions are small yellow-white subconjunctival spots that can be asymptomatic for years and are an incidental finding on routine ophthalmic examination.[1,2] The etiologies are chronic vernal conjunctivitis, severe atopic keratoconjunctivitis, and post-trachomatous changes.[1–5] The concretions are located within the conjunctival recesses layered by epithelium and are made up of cell debris and phospholipids.[3,4] Case Report A 24-year-old-female presented with chronic irritation and redness in the right eye for 1 year. Systemic evaluation was unremarkable with no immunosuppressive disorders. Her symptoms were intermittently associated with discharge. Her vision was 20/20 in both eyes. Everted upper and lower eyelids of the right eye showed multiple concretions scattered beneath the tarsal conjunctiva, more in the upper eyelid, protruding through the conjunctiva at two focal points [Fig. 1]. There was diffuse congestion in the tarsal conjunctiva with a normal lid margin appearance. The left eyelid had neither congestion nor concretions. There was no ocular surface staining with fluorescein dye, but the exposed concretions showed fluorescein uptake. Dry eye workup revealed a tear break-up time of 7.8 s with normal tear secretion in the right eye, whereas tear breakup and secretion was normal in the left eye. She gave an antecedent history of a vegetative object falling into her right eye while driving a bike. The ocular symptoms developed 1 month after that trauma. She was using topical lubricants and antibiotic ointment over the eyelid margin for the meibomitis. Because of unilateral presentation and young age, two concretions were removed on slit lamp using a sterile 26-gauge needle under topical anesthesia and sent for tissue processing.Figure 1: (a) Everted right upper eyelid of the 24-year-old female shows scattered numerous concretions in the tarsal conjunctiva, fewer in the lower eyelid (d). (b) Gomori methenamine silver stain (100×) shows presence of fungal elements within the concretions (higher magnification in inset), which are also seen on gram stain (e, 400×) as empty round and vertical spaces. (c and f) Left upper and lower eyelids show normal tarsal conjunctivaThe histopathologic examination of concretions revealed eosinophilic homogenous amorphous material containing fungal elements, which were Gomori methenamine silver stain positive [Fig. 2]. The deepest sections (till the last tissue section) also contained numerous fungal hyphae, which ruled out contamination. Due to atypical histopathologic findings of concretions, a superficial conjunctival incision biopsy containing tarsal conjunctiva, concretions, and a thin sliver of tarsus was taken at 4 mm from the upper eyelid margin. The incision biopsy showed scattered amorphous, acellular eosinophilic hyaline-like material and diffuse lymphocytic infiltration in the substantia propria. The amorphous material had a rim of conjunctival epithelium suggestive of concretions embedded within the pseudoglands of Henle. The periphery of the concretions had a slight basophilic appearance. The concretions were negative for von Kossa stain, Alcian blue, and Congo red but were red on Masson trichrome stain, suggesting cell debris and keratin to be one of the constituents. Alcian blue was focally positive in the conjunctival epithelium at the location of goblet cells. She was started on tablet fluconazole 150 mg once a week for 3 weeks along with 1% itraconazole eye ointment into the eye twice a day. Her symptoms resolved with a reduction in the conjunctival concretions’ number and conjunctival congestion.Figure 2: (a) Everted right upper eyelid with highlighted biopsied part. (b and c) Incisional biopsy from the upper tarsal conjunctiva shows diffuse subepithelial lymphocytic infiltration along with eosinophilic hyaline-like material deposits (c, at 200×). Masson trichrome (d), Congo red (e), and Alcian blue (f) were negative for amyloid and mucopolysaccharide depositsTear film instability and reduced tear production have been reported in patients with conjunctival concretions, where 51% of asymptomatic patients with conjunctival concretions have reduced tear break-up time.[3] The current case had reduced tear break-up time in the eye with concretions. An interesting finding in the current case was the presence of fungal elements, which can be explained by the possible entrapment of organic body containing fungus 1 year back. The organic material dissolved away, but the fungal elements caused persistent chronic inflammation, which resulted in the formation of concretions. This case reports the possibility of having unilateral concretions secondary to chronic inflammation with degenerative changes. Declaration of patient consent The authors certify that they have obtained all appropriate patient consent forms. In the form, the patient(s) has/have given his/her/their consent for his/her/their images and other clinical information to be reported in the journal. The patients understand that their names and initials will not be published and due efforts will be made to conceal their identity, but anonymity cannot be guaranteed. Financial support and sponsorship Hyderabad Eye Research Foundation, Hyderabad. Conflicts of interest There are no conflicts of interest.

Extrapulmonary Pneumocystis jirovecii infection in an advanced HIV-infected patient: A case report and literature review

BackgroundPneumocystis jirovecii infection is the most common opportunistic infection that causes pneumonia in human immunodeficiency virus (HIV)-infected patients; however, extrapulmonary P. jirovecii infection is extremely rare after the use of antiretroviral therapy. Here, we present the second reported case of paraspinal mass caused by P. jirovecii infection in an advanced HIV-infected patient.Case presentationA 45-year-old woman presented with dyspnea on exertion, and significant weight loss within the preceding 4 months. Initial complete blood count (CBC) findings revealed pancytopenia with a hemoglobin (Hb) level of 8.9 g/dL, a white blood cell (WBC) count of 2180 cells/mm3 with 68% neutrophils, and a platelet count of 106,000 cells/mm3. Anti-HIV was positive with an absolute cluster of differentiation 4 (CD4) count of 16 cells/ mm3. A computed tomography scan of the chest revealed an enhancing soft tissue mass-like lesion at the right paravertebral region (T5-T10 level) and a thick-walled cavity lesion at the left lower lung. A CT-guided biopsy of the paravertebral mass was performed and histopathology revealed granulomatous inflammation consisting of dense aggregates of epithelioid cells and macrophages, and scattered foci of pink foamy to granular materials amidst the granulomatous inflammation. Gomori methenamine silver (GMS) staining revealed thin cystic-like structures (ascus) that were observed to be morphologically consistent with P. jirovecii. Molecular identification and DNA sequencing from the paraspinal mass was 100% identical to P. Jirovecii. The patient was successfully treated with oral trimethoprim-sulfamethoxazole for 3 weeks and antiretroviral therapy (ART) with tenofovir (TDF), lamivudine (3TC), and dolutegravir (DTG). A follow-up CT scan of the chest at 2 months after treatment showed a decrease in sizes of both the paravertebral mass and the cavitary lung lesion.ConclusionsExtrapulmonary pneumocystosis (EPCP) has become an extremely rare condition in HIV-infected patients after the widespread use of ART. EPCP should be considered in ART-naive HIV-infected patients suspected of having or diagnosed with Pneumocystis jirovecii pneumonia who present with atypical symptoms and/or signs. Histopathologic examination with GMS staining of affected tissue is necessary for the diagnosis of EPCP.

Metagenomic next-generation sequencing for the diagnosis of Pneumocystis jirovecii Pneumonia in critically pediatric patients

ObjectiveThe aim of this study was to evaluate the effectiveness of metagenomic next-generation sequencing (mNGS) for the diagnosis of Pneumocystis jirovecii Pneumonia (PCP) in critically pediatric patients.MethodsSeventeen critically pediatric patients with PCP and sixty patients diagnosed with non-PCP pneumonia who were admitted in pediatric intensive care unit between June 2018 and July 2021 were enrolled. Conventional methods and mNGS for detecting Pneumocystis jirovecii (P. jirovecii) were compared. The patients’ demographics, comorbidities, laboratory test results, antibiotic treatment response and 30 day mortality were analyzed.ResultThe mNGS showed a satisfying diagnostic performance with a sensitivity of 100% in detecting P. jirovecii compared with Gomori methenamine silver staining (5.9%), serum (1,3)-β-D-glucan (86.7%) and and LDH (55.6%). The diagnostic specificity of mNGS for PCP was higher than that of serum BDG (56.7%) and LDH (71.4%). In PCP group, over one thirds’ cases had mixed infections. Compared with survivors, non-survivors had higher stringently mapped read numbers (SMRNs) in bronchoalveolar lavage fluid (BALF) sample (P < 0.05), suggesting SMRNs were closely associated with the severity of response. The detection for P. jirovecii by mNGS both in BALF and blood samples reached a concordance rate of 100%, and the SMRNs in the BALF were remarkably higher than that in blood samples. Initial antimicrobial treatment was modified in 88.2% of PCP patients based on the mNGS results.ConclusionThe mNGS is a potential and efficient technology in diagnosing PCP and shows a satisfying performance in the detection of co-pathogens. Both blood and BALF samples for mNGS are suggested for the presumptive diagnosis of PCP.

Detection of Candida in Cervical Cancer Tissue by Periodic Acid Shiff and Gomori Methenamine Silver Staining

Detection of Candida in Cervical Cancer Tissue by Periodic Acid Shiff and Gomori Methenamine Silver Staining

Concurrent Occurrence of Nasopharyngeal and Disseminated Cutaneous Rhinosporidiosis: A Rare Presentation.

Concurrent Occurrence of Nasopharyngeal and Disseminated Cutaneous Rhinosporidiosis: A Rare Presentation.

Deep Dermatophytosis Presented as Multiple Exophytic Masses caused by Trichophyton rubrum in Immunocompromised Patient with Rheumatoid Arthritis; A Case Report

Dermatophytes invade the stratum corneum and infect the skin, nails, and hair, mostly resulting in superficial infections. Deep dermatophytosis involving the dermis and subcutaneous layer has rarely been reported in immunocompromised individuals. Herein, we report a case of deep dermatophytosis caused by Trichophyton rubrum. A 71-year-old woman presented with multiple erythematous exophytic and subcutaneous nodules in both the lower legs. The patient was taking immunosuppressive agents for rheumatoid arthritis and antifungal agents for tinea pedis and onychomycosis, which was improperly ceased. Histopathological findings showed diffuse granulomatous infiltration with multinucleated cells, lymphocytes, histiocytes, and neutrophils in the dermis. Septate and branched hyphae were observed in the dermis using periodic acid-Schiff diastase and Gomori methenamine silver staining. T. rubrum was identified in fungal culture from the tissue sample and confirmed through phylogenetic analysis of the internal transcribed spacer and large subunit regions in ribosomal ribonucleic acid gene. Intravenous amphotericin B was administered for septic shock before the confirmation of the causative organism, which rapidly improved the condition.

Cutaneous Infection by Fusarium solani in an Immunocompetent Patient

Fusarium species are widely distributed throughout the environment, and rarely cause infection in healthy individuals. Herein, we report a rare case of cutaneous infection by Fusarium solani in a healthy patient. A 64-year-old patient presented with a tender, erythematous, deep-seated nodule above the left ankle after an intralesional triamcinolone injection for lipoma was administered at a private clinic 1 month ago. Fungal hyphae were identified in subcutaneous tissue by skin biopsy using Gomori methenamine silver stain. Lactophenol cotton blue staining, fungal culturing, and 28S rRNA sequencing confirmed the presence of F. solani. After diagnosis, the patient was successfully treated with oral itraconazole.

Uric Acid Expression in Carotid Atherosclerotic Plaque and Serum Uric Acid Are Associated With Cerebrovascular Events.

Uric acid (UA) concentration within carotid plaque and its association with cerebrovascular events have not been detected or quantified. Systemically, serum UA is a marker of inflammation and risk factor for atherosclerosis. However, its association with carotid plaque instability and stroke pathogenesis remains unclear. In patients undergoing carotid endarterectomy, we aimed to determine whether UA is present differentially in symptomatic versus asymptomatic carotid plaques and whether serum UA is associated with cerebrovascular symptoms (stroke, transient ischemic attack, or amaurosis fugax). Carotid atherosclerotic plaques were collected during carotid endarterectomy. The presence of UA was assessed using Gomori methenamine silver staining as well as anti-UA immunohistochemical staining and its quantity measured using an enzymatic colorimetric assay. Clinical information was obtained through a retrospective review of data. UA was more commonly detected in symptomatic (n=23) compared with asymptomatic (n=9) carotid plaques by Gomori methenamine silver (20 [86.9%] versus 2 [22.2%]; P=0.001) and anti-UA immunohistochemistry (16 [69.5%] versus 1 [11.1%]; P=0.004). UA concentration was higher in symptomatic rather than asymptomatic plaques (25.1 [9.5] versus 17.9 [3.8] µg/g; P=0.021). Before carotid endarterectomy, serum UA levels were higher in symptomatic (n=341) compared with asymptomatic (n=146) patients (5.9 [interquartile range, 4.6-6.9] mg/dL versus 5.2 [interquartile range, 4.6-6.2] mg/dL; P=0.009). The current study supports a potential role of UA as a potential tissue participant and a systemic biomarker in the pathogenesis of carotid atherosclerosis. UA may provide a mechanistic explanation for plaque instability and subsequent ischemic cerebrovascular events.

Metagenomic Next-Generation Sequencing Is Highly Efficient in Diagnosing Pneumocystis Jirovecii Pneumonia in the Immunocompromised Patients.

PurposesTo explore the value of metagenomic next-generation sequencing (mNGS) in diagnosing pneumocystis jiroveciipneumonia (PJP) in the immunocompromised patients.MethodsData of 122 patients with PJP in an immunosuppressed state and 67 non-PJP patients were collected. The diagnostic efficacy of mNGS was compared with the conventional methods, including Gomori methenamine silver (GMS) staining and serum (1,3)-β-D-glucan (BDG). Changes of anti-microbial therapy for patients with PJP based on mNGS results were also reviewed.ResultsThe diagnostic sensitivity of mNGS to PJP was higher than that of GMS and BDG (100% vs. 15 and 74.5%, p < 0.001). The diagnostic specificity (91.%) was lower than that of GMS (100%), and similar with BDG (89.6%). In addition to P. jirovecii, mNGS revealed co-pathogens like human β-herpesvirus 5, human γ-pesvirus 4, and some other opportunistic pathogens. The reads of mNGS were remarkably higher in BALF than in blood samples. Initial antimicrobial treatment was modified in 89.3% patients based on the mNGS results, and 74 cases (60.7%) were treated with anti-P. jirovecii therapy.ConclusionmNGS is highly efficient in diagnosing PJP and good at identifying pathogens in mixed infections.

Pneumocystis Jirovecii Pneumonia Diagnosis via Metagenomic Next-Generation Sequencing.

The incidence of non-HIV-infected Pneumocystis Jirovecii Pneumonia (PJP) is increasing. The prognosis for non-HIV PJP is poor and diagnostic tests are of lower sensitivity in non-HIV patients. Metagenomic next-generation sequencing (mNGS) was compared with routine detection assays, including Gomori methenamine silver (GMS) stain and polymerase chain reaction (PCR) technique. Specimens of 4 bronchoalveolar lavages (BAL) and 1 lung tissue samples were obtained from 4 non-HIV patients from our hospitals. Although both GMS and mNGS were positive for P. jirovecii with PCR as positive control, the testing time of mNGS was obviously shorter than GMS. Compared with the traditional GMS method, mNGS has absolute advantages. However, the issue with PJP presentations having atypical symptoms and ambiguous imaging features persists. Hence, the disease can easily be ignored. Secondly, PJP progresses rapidly in non-HIV-infected patients and can cause severe respiratory failure with unfavorable prognosis. This study affirms that mNGS can be used to quickly and accurately diagnose PJP, but a combination of clinical judgement of symptoms, laboratory testing, and imaging examination is required to make a comprehensive judgment along with mNGS test results.

Characteristics and Prognostic Factors of Non-HIV Immunocompromised Patients With Pneumocystis Pneumonia Diagnosed by Metagenomics Next-Generation Sequencing.

ObjectiveThe aim of this study was to evaluate the potential of metagenomic next-generation sequencing (mNGS) for the diagnosis of pneumocystis pneumonia (PCP) in patients with non-human immunodeficiency virus-infection and to discuss the clinical characteristics and identify prognostic factors associated with patients with non-HIV PCP.MethodsForty-six patients with PCP who were admitted in respiratory intensive care unit (ICU) between May 2018 and May 2020 were retrospectively reviewed. The subjects were divided into survivor and non-survivor groups according to the patients' outcome. Conventional methods and mNGS for detecting Pneumocystis jirovecii (P. jirovecii) were analyzed. The patients' demographics, comorbidities, laboratory parameters, and treatments were compared and evaluated in both groups to identify risk factors for mortality by using univariate and multivariate logistic regression.ResultsMetagenomic next-generation sequencing (mNGS) showed a satisfying diagnostic performance of 100% positive of detecting P. jirovecii from bronchoalveolar lavage (BAL) specimens in forty-six patients with non-HIV PCP, compared to only 15.2% for Gomori Methenamine silver (GMS) staining and 84.8% for Serum 1,3-beta-D-glucan (BDG). Among them, the mean age was 46.4-year-old (range 18–79-year-old) and mortality rate was 43.5%. The dominant underlying conditions were connective tissue diseases (34.8%), autoimmune kidney diseases (30.4%), followed by hematologic malignancies (10.9%), and solid organ transplantation (6.5%). A total of 38 cases (82.6%) received glucocorticoid and 19 cases (41.3%) used immunosuppressant within 3 months before diagnosed PCP. Multiple infections were very common, over two thirds' cases had mixed infections. Compared with survivors, non-survivors had a higher acute physiology and chronic health evaluation II (APACHE II) score (14.4 ± 4.8 vs. 10 ± 3.4), Procalcitonin (PCT) [ng/ml: 0.737 (0.122–1.6) vs. 0.23 (0.095–0.35)], lactic dehydrogenase (LDH) [U/L: 1372 (825.5–2150) vs. 739 (490.5–956)], and neutrophil-lymphocyte ratio (NLR) [21.6 (15.67–38.2) vs. 11.75 (5.1–15.52)], but had a lower PaO2/FiO2 ratio (mmHg:108.8 ± 42.4 vs. 150.5 ± 47.5), lymphocytes [×109/L: 0.33 (0.135–0.615) vs. 0.69 (0.325–1.07)] and CD4+ T cells [cell/μl: 112 (53.5–264) vs. 255 (145–303.5)], all P < 0.05. Furthermore, we found non-survivors' PaO2/FiO2 ratio of day 3 and day 7 had not improved when compared with that of day one, and platelet level and NLR became worse. Multivariate analysis showed that other pathogens' co-infection (OR = 9.011, 95% CI was 1.052–77.161, P = 0.045) and NLR (OR = 1.283, 95% CI was 1.046–1.547, P = 0.017) were the independent risk factors of poor prognosis.ConclusionmNGS is a very sensitive diagnostic tool for identifying P. jirovecii in patients who are non-HIV immunocompromised. PCP in patients who are non-HIV infected is associated with a high rate of multiple infections and severe condition. Mixed infection and elevation of NLR were the independent risk factors of poor prognosis.

Abstract WP237: Uric Acid Expressed Within Carotid Atherosclerotic Plaque And Serum Uric Acid Are Associated With Cerebrovascular Events

Introduction: Uric acid (UA) level within carotid plaque specimens and its association with cerebrovascular events has not been detected or quantified. Systemically, serum uric acid (SUA) is a marker of inflammation and risk factor for atherosclerosis. However, its association with carotid plaque instability and subsequent cerebrovascular events remains unclear. Hypothesis: We hypothesized that UA may play a role in carotid artery atherosclerosis and stroke pathogenesis. In patients undergoing carotid endarterectomy (CEA), we aimed to determine whether UA is present differentially in carotid plaques of symptomatic versus asymptomatic patients and whether SUA is associated with cerebrovascular symptoms before surgery and major adverse cardiovascular events (MACE) in the follow-up to CEA. Methods: We prospectively collected 32 carotid plaques during CEA. We qualitatively assessed the presence of UA by using Gomori methenamine silver (GMS) staining and immunohistochemistry with anti-UA antibodies. We extracted and measured the quantity of UA in carotid plaques by using an enzymatic colorimetric assay. A retrospective analysis of a clinical database of 534 consecutive patients who underwent CEA was performed. We defined as symptomatic those patients with history of stroke, TIA and amaurosis fugax. Results: UA positive staining was detected in higher number of carotid plaques from symptomatic compared to asymptomatic patients on both staining, GMS [20 (86.9%) vs 2 (22.2%); p=0.001] and immunohistochemistry anti-UA [16 (69.5%) vs 1 (11.1%); p=0.004]. Moreover, we found a significantly higher concentration of UA in carotid plaque from symptomatic compared to asymptomatic patients [25.1 (9.5) μg/g vs 17.9 (3.8) μg/g; p=0.021]. SUA level in patients prior to CEA was significantly higher in the symptomatic compared to the asymptomatic group [6.0 (4.8-7.1) mg/dL vs 5.2 (4.4-6.3) mg/dL; p&lt;0.001] and it was significantly higher in patients who developed MACE after CEA versus those that did not [6.0 (4.8-7.1) mg/dL vs 5.4 (4.5-6.6) mg/dL; p=0.021]. Conclusions: UA may provide a mechanistic explanation as systemic biomarker and potential tissue contributor for carotid plaque instability and subsequent cerebrovascular and cardiovascular symptoms.