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Rapid and reliable diagnosis of mucormycosis using colorimetric loop-mediated isothermal amplification.

Current diagnostic approaches for mucormycosis are often limited by low sensitivity and prolonged turnaround times, which result in delayed treatment and poor clinical outcomes. We developed a novel diagnostic method utilizing a colorimetric loop-mediated isothermal amplification (LAMP) assay for the rapid and sensitive detection of mucormycosis. The assay incorporates specifically designed primers capable of detecting as low as 0.001 picograms (pg) of spiked genomic DNA from Mucorales fungi. This LAMP assay demonstrated a high sensitivity of 98% and a 100% specificity of detecting fungal ribosomal DNA (rDNA) in bronchoalveolar lavage (BAL) samples collected from mice infected with Mucorales fungi (n = 48) or from an uninfected control group (n = 15). To align the assay with clinical antifungal therapy, a subset of infected mice was treated with either liposomal amphotericin B (LAMB) or a combination of LAMB and a humanized monoclonal antibody (VX-01) targeting the Mucorales-specific surface protein CotH3. Consistent with the treatment efficacy, the LAMP assay detected significantly lower fungal burdens in BAL samples from mice receiving the combination therapy compared to those treated with LAMB alone or placebo. Further validation was conducted using BAL samples from patients diagnosed with mucormycosis (n = 24) or aspergillosis (n = 17). The assay demonstrated a sensitivity of 79% and a specificity of 94%. These findings highlight the diagnostic potential of this LAMP-based assay as a point-of-care. Its high sensitivity, specificity, and rapid turnaround time position this assay as a promising tool for early and accurate detection of mucormycosis, with the potential to improve patient management and clinical outcomes.IMPORTANCEMucormycosis is a rapidly progressive and fatal fungal infection. Timely diagnosis is critical for effective treatment, yet current diagnostic tools are slow, insensitive, or require complex laboratory procedures. In this study, we developed and validated a colorimetric loop-mediated isothermal amplification (LAMP) assay that enables rapid and reliable detection of Mucorales DNA directly from bronchoalveolar lavage (BAL) specimens. The assay demonstrated high sensitivity and specificity in both experimental mouse models and clinical samples, producing results within 1 h without the need for sophisticated equipment. This simple, robust, and cost-effective molecular diagnostic tool holds great potential for early detection of mucormycosis, facilitating prompt antifungal therapy and improving patient survival.

Mucormycosis

Localized mucormycosis following intramuscular corticosteroid. Case report and review of the literature.

A case of subcutaneous localized mucormycosis infection which developed following intramuscular (IM) injection of corticosteroid in a patient with leukemia is presented. Aggressive treatment, which included wide local excision, systemic amphotericin-B, and chemotherapy for the leukemia, resulted in eradication of the infection and complete healing of the wound. A review of the literature revealed nine other patients with the localized subcutaneous form of mucormycosis (excluding patients with burns and rhinocerebral types) and six of those nine patients also survived the infection. It is possibly the mildness of the underlying predisposing factors that allows some of these patients to contain the infection at a single site. It is apparent from review of the literature that in subcutaneous localized forms of mucormycosis, the outcome has been generally good. This contrasts sharply with other clinical forms of mucormycosis infections where the underlying predisposing factors are usually severe and any kind of therapeutic approach has been almost always futile. Subcutaneous mucormycosis infection differs clinically and histopathologically from subcutaneous localized entomophthoromycosis which is seen predominantly in tropical countries. An attempt is made to clarify the terminology of these interesting fungi in language that is taxonomically up-to-date and still useful to clinicians.

Adult

Acute subdural hematoma and intracerebral hemorrhage. Rare complications of rhinocerebral mucormycosis.

Rhinocerebral mucormycosis is a short-term and often rapidly lethal fungal disease. It is generally seen in uncontrolled cases of diabetes with ketoacidosis. This case exhibits many of the features of a typical fulminating rhinocerebral mucormycosis. However, the fatal complications of acute subdural hematoma and massive intracerebral hemorrhage due to rupture of aneurysm, as demonstrated by angiography, are unique clinical manifestations of patients with rhinocerebral mucormycosis.

Antifungal Agents

Rhinocerebral mucormycosis: premortem diagnosis and therapy.

The diagnosis of rhinocerebral mucormycosis is most often made at autopsy. We report a series of nine patients in whom the diagnosis was established premortem. Six of the patients had underlying diabetes mellitus and three had acute leukemia. Facial or ocular pain was the complaint found in all patients, and frequently was the initial symptom. The diagnosis was established by examination and culture of infected tissue obtained by biopsy. In seven patients, identification of hyphal elements in smears of biopsy material allowed the immediate institution of amphotericin B therapy. Four of the seven patients treated with amphotericin B survived. All surviving patients had underlying diabetes mellitus and had undergone surgical debridement. Early diagnosis leading to immediate institution of appropriate therapy is most important for survival of patients with mucormycosis.

Acute Disease

Cerebral mucormycosis: an unusual case.

A 36-year-old obese woman with hyperglycemia and immunosuppression died of bilateral internal carotid artery occlusion associated with mucormycosis. This report describes a rare case in which cerebral mucormycosis occurred without the usual preceding clinical evidence of nasal and orbital infection.

Adult

Rhinocerebral mucormycosis.

Mucormycosis is a fulminant fungal infection occurring in debilitated patients with an underlying pathologic condition. The rhinocerebral form of the disease, which comprises nearly one half of recently reported cases, is most often found in uncontrolled diabetics or profoundly dehydrated children. Infection usually begins in the nose and progresses through the paranasal sinuses, invading the orbit and CNS secondarily. Despite the known pathogenesis of this disease, the ophthalmologist is often first to consider the diagnosis, due to inadequate intranasal examination by the primary physician. The delay caused by late occurrence of orbital manifestations has resulted in poor survival rates, despite vigorous therapy. In recent years, increased physician awareness has led to earlier diagnosis of rhinocerebral mucormycosis. This report presents 13 cases with which we have delt since 1963. The long-term survival rate is 85%. Aggressive surgical therapy, with repeated debridement, in combination with intravenous amphotericin B, have led to this high rate of cure.

Acidosis

Rhizopus rhizopodiformis: emerging etiological agent of mucormycosis.

Mucormycosis is caused principally by members of the genus Rhizopus, especially R arrhizus and R. oryzae. Infection attributable to R. rhizopodiformis has rarely been documented. Of 13 cases of mucormycosis diagnosed during a 4-year period (1974 to 1978) at The Mount Sinai Hospital, 6 cases, occurring within 9 months, were caused by R. rhizopodiformis. The six isolates were identified mainly by: growth at 50 degrees C; production of short, sometimes branched, sporangiophores arising from opposite rhizoids; elongated columellae; and small spherical-to-elliptical, smooth-to-finely striated sporangiospores. The possibility that this explosive occurrence of R. rhizopodiformis at our institution was because of nosocomial acquisition was strongly supported by the recovery of this same mycotic agent from adhesive bandages used in the cardiac intensive care unit, where a patient developed subcutaneous R. rhizopodiformis infection after cardiac surgery. The invasive potential of R. rhizopodiformis was manifested by the extensive subcutaneous and systemic infections in each of the six patients, three of whom developed antibody against this mucormycotic agent.

Adult

Hematological diseases-related mucormycosis: A retrospective single center study.

BACKGROUND AND AIM: Mucormycosis is a life-threatening invasive fungal infection. This study aimed to analyze the clinical characteristics of patients with hematologic malignancies complicated with mucormycosis. METHODS: This retrospective study investigated the clinical characteristics, epidemiological features, treatment, and prognosis of 46 patients with hematological diseases and Mucor infection as indicated by mNGS from August 28, 2020 to September 11, 2023. Metagenomic next-generation sequencing (mNGS) refers to the application of high-throughput sequencing technology for the comprehensive analysis of nucleic acid content in patient samples, facilitating the detection and characterization of microbial DNA and/or RNA, and then comparing and analyzing the results with an information database to determine the types of pathogenic microorganisms present in the sample. RESULTS: The median age of admission for the included patients was 49 years (9-78). Multivariate analysis identified age over 60 years (p&#x2009;=&#x2009;0.006&#x2009;<&#x2009;0.05), high-dose corticosteroids (p&#x2009;=&#x2009;0.001&#x2009;<&#x2009;0.05), neutropenia lasting more than 10 days (p&#x2009;=&#x2009;0.041&#x2009;<&#x2009;0.05), and two or more Mucor infections (p&#x2009;=&#x2009;0.004&#x2009;<&#x2009;0.05) were independent risk factors for OS in patients with hematological diseases. Moreover, differences between groups were analyzed using the Fisher exact probability method, and no significant difference was observed in the efficacy of various types of antifungal therapies. CONCLUSION: Patients with hematologic malignancies benefit greatly from early diagnosis and treatment when suspected of Mucor infection. mNGS is an important supplementary method for early diagnosis of Mucor infection. Moderated use of corticosteroids, reducing the duration of neutropenia, and enhancing autologous immune function are important measures to reduce patient mortality rate.

Retrospective Studies

Pulmonary mucormycosis: another cure.

Pulmonary mucormycosis in an ill patient with poorly controlled chronic lymphocytic leukemia was diagnosed with open lung biopsy without excision. He improved on medical management and became ambulatory. At autopsy one year later, no residual mucormycosis was present. Better control of leukemia and more specific antimicrobial therapy are discussed as potentially important factors in patient management.

Aged

Rhinocerebral mucormycosis.

Rhinocerebral mucormycosis is a fungal diseases that has a 50% mortality. Its occurrence has increased, possibly because of greater use of chemotherapeutic agents that mya compromise the immunologic defenses of the host or alter the normal flora. The earliest signs, ulceration and pain, may appear in the mouth. In the patient described in this report, the autopsy showed that mucormycosis had entered the brain cells.

Amphotericin B

[Cranial mucormycosis in a patient with a transplanted kidney].

Mucormycosis is a very serious complication of debilitating diseases, and particularly of diabetes. Presently the treatment of choice is amphotericine B. A patient is described who, like most other renal transplant patients with mucormycosis, had cranial localization of this disease and diabetes. The clinical findings were classical and the diagnosis was confirmed histologically.

Adult

Pulmonary and rhinocerebral mucormycosis. Successful outcome with amphotericin B and griseofulvin therapy.

An ominous prognosis is associated with combined pulmonary and rhinocerebral mucormycosis (phycomycosis). We report the case of a diabetic patient with ketoacidosis who had extensive pulmonary and rhinocerebral mucormycosis that responded satisfactorily to amphotericin B and griseofulvin therapy. The affected lung is completely atelectatic and has remained so for 12 months without evidence of necrosis or abscess formation.

Adult

[Cranial mucormycosis with thrombosis of the sinus cavernosus (author's transl)].

A case of mucormycosis in a 45 year-old woman with uncontrolled diabetes is reported. Following dental extractions, the patient presented with total ophthalmoplegia, chemosis and complete sensory loss of the right fifth cranial nerve. Exophthalmus was not present. Although a fungal infection was suspected, administration of Amphotericin B was avoided because of renal insufficiency. After temporary clinical improvement with high doses of antibiotics, signs of right seventh and eighth cranial nerve paralysis and of thrombosis of the right ophthalmic artery developed. The patient expired after six days. Autopsy disclosed extensive mucormycosis with involvement of the nasal cavity and paranasal sinuses, soft tissues and bones of face and orbit, cranial nerves, meninges and base of the brain, as well as mycotic thrombosis of the right cavernous sinus and the internal carotid, ophthalmic and maxillary arteries. The spread of infection is detailed, and the importance of an early diagnosis is stressed.

Carotid Artery Diseases

[Outbreak of paraventricular and ventricular mucormycosis in fattening bulls].

There is a description of mass occurrence of nine cases of acute, fatal paraventricular and ventricular mucormycosis in fattened bulls at the age of about 18 months. In the reticulum, the disease was characterized by numerous ulcerations penetrating into the submucosa. In rumen the deposits were of hemorrhagic character penetrating through a thick muscular layer into the subserosa. Hemorrhagic peritonitis was within the range of those changes. Thromboses of arteries and veins prevailed in the microscopic picture. Inflammatory reparative changes were formed by neutrophile leucocytes. Mucoraceum fibres were demonstrated in pathological manifestations. In three cases, strains of Rhizopus cohnii and Absidia ramosa were cultivated simultaneously from the sub-ulcerous layers in the reticulum. A number of findings testify that the mass occurrence of mucormycosis followed the feeding of mouldy bakery wastes on the basis of acidosis.

Animals

Cutaneous infarction. Manifestation of disseminated mucormycosis.

We describe the second reported case of cutaneous infarction as a manifestation of disseminated mucormycosis. The lesion, which closely resembled ecthyma gangrenosum, occurred in a leukemic patient who was on a regimen of broad-spectrum antibiotics. In this case as in the only other reported case, Mucor pusillus was the cause.

Adult

Mucormycosis. A complication of critical care.

Lethal nosocomial mucormycosis developed in three previously well individuals while they were receiving intensive care for acute hemorrhagic pancreatitis, for cardiogenic shock, and for a ruptured intra-abdominal aortic aneurysm. In two cases, the condition was first seen as progressive cavitary pneumonia refractory to antibacterial therapy; Mucoraceae was identified in all three patients only at autopsy. Each patient had received large doses of corticosteroids and broad-spectrum antibiotics, and all had suffered from respiratory failure, acute renal failure with acidosis, and severe hyperglycemia in association with total parenteral nutrition. Mucoraceae should be regarded as an additional nosocomial pathogen in the setting of advanced life-support care.

Acidosis

Experimental mucormycosis in congenitally athymic (nude) mice.

Athymic nude (nu/nu) mice and their phenotypically normal (nu/+) littermates displayed a similar susceptibility to acute lethal infection with Absidia corymbifera. Although the clinical manifestations of acute infection were also similar in both groups, the nude mice tended to develop more extensive lesions and were less effective in eliminating viable A. corymbifera spores than their heterozygous littermates. The results suggested that thymus-dependent processes did not play an essential role in primary resistance to mucormycosis.

Animals

Gastrointestinal mucormycosis.

Mucormycosis is an acutely malignant fungal disease causing local thrombosis and infarction. Gastrointestinal infection most commonly occurs in diabetics or in the immunologically compromised. This case demonstrates the radiological appearances of the gastrointestinal lesion and prolonged survival with amphotericin-B therapy.

Adult