Innate and adaptive immune responses in subjects with CPA secondary to post-pulmonary tuberculosis lung abnormalities.

BACKGROUND: Post-tuberculosis lung abnormality (PTLA) is the most common risk factor for chronic pulmonary aspergillosis (CPA), and 14%-25% of the subjects with PTLA develop CPA. The pathogenesis and the host immune response in subjects with PTLA who develop CPA need to be better understood. METHODS: We prospectively compared the innate and adaptive immune responses mounted by patients of PTLA with or without CPA (controls). We studied the neutrophil oxidative burst (by dihydrorhodamine 123 test), classic (serum C3 and C4 levels) and alternative (mannose-binding lectin [MBL] protein levels) complement pathway, serum immunoglobulins (IgG, IgM and IgA), B and T lymphocytes and their subsets in subjects with PTLA with or without CPA. RESULTS: We included 111 subjects (58 CPA and 53 controls) in the current study. The mean ± SD age of the study population was 42.6 ± 15.7 years. The cases and controls were matched for age, gender distribution and body weight. Subjects with CPA had impaired neutrophil oxidative burst, lower memory T lymphocytes and impaired Th-1 immune response (lower Th-1 lymphocytes) than controls. We found no significant difference between the two groups in the serum complement levels, MBL levels, B-cell subsets and other T lymphocyte subsets. CONCLUSION: Subjects with CPA secondary to PTLA have impaired neutrophil oxidative burst and a lower Th-1 response than controls.

Novel acute hypersensitivity pneumonitis model induced by airway mycosis and high dose lipopolysaccharide.

BACKGROUND: Inhalation of fungal spores is a strong risk factor for severe asthma and experimentally leads to development of airway mycosis and asthma-like disease in mice. However, in addition to fungal spores, humans are simultaneously exposed to other inflammatory agents such as lipopolysaccharide (LPS), with uncertain relevance to disease expression. To determine how high dose inhalation of LPS influences the expression of allergic airway disease induced by the allergenic mold Aspergillus niger (A. niger). METHODS: C57BL/6J mice were intranasally challenged with the viable spores of A. niger with and without 1 µg of LPS over two weeks. Changes in airway hyperreactivity, airway and lung inflammatory cell recruitment, antigen-specific immunoglobulins, and histopathology were determined. RESULTS: In comparison to mice challenged only with A. niger, addition of LPS (1 µg) to A. niger abrogated airway hyperresponsiveness and strongly attenuated airway eosinophilia, PAS+ goblet cells and TH2 responses while enhancing TH1 and TH17 cell recruitment to lung. Addition of LPS resulted in more severe, diffuse lung inflammation with scattered, loosely-formed parenchymal granulomas, but failed to alter fungus-induced IgE and IgG antibodies. CONCLUSIONS: In contrast to the strongly allergic lung phenotype induced by fungal spores alone, addition of a relatively high dose of LPS abrogates asthma-like features, replacing them with a phenotype more consistent with acute hypersensitivity pneumonitis (HP). These findings extend the already established link between airway mycosis and asthma to HP and describe a robust model for further dissecting the pathophysiology of HP.

Lung eosinophils elicited during allergic and acute aspergillosis express RORγt and IL-23R but do not require IL-23 for IL-17 production.

Exposure to the mold, Aspergillus, is ubiquitous and generally has no adverse consequences in immunocompetent persons. However, invasive and allergic aspergillosis can develop in immunocompromised and atopic individuals, respectively. Previously, we demonstrated that mouse lung eosinophils produce IL-17 in response to stimulation by live conidia and antigens of A. fumigatus. Here, we utilized murine models of allergic and acute pulmonary aspergillosis to determine the association of IL-23, IL-23R and RORγt with eosinophil IL-17 expression. Following A. fumigatus stimulation, a population of lung eosinophils expressed RORγt, the master transcription factor for IL-17 regulation. Eosinophil RORγt expression was demonstrated by flow cytometry, confocal microscopy, western blotting and an mCherry reporter mouse. Both nuclear and cytoplasmic localization of RORγt in eosinophils were observed, although the former predominated. A population of lung eosinophils also expressed IL-23R. While expression of IL-23R was positively correlated with expression of RORγt, expression of RORγt and IL-17 was similar when comparing lung eosinophils from A. fumigatus-challenged wild-type and IL-23p19-/- mice. Thus, in allergic and acute models of pulmonary aspergillosis, lung eosinophils express IL-17, RORγt and IL-23R. However, IL-23 is dispensable for production of IL-17 and RORγt.

The IL-1 Receptor Is Required to Maintain Neutrophil Viability and Function During Aspergillus fumigatus Airway Infection.

Aspergillus fumigatus airway infections are associated with increased rates of hospitalizations and declining lung function in patients with chronic lung disease. While the pathogenesis of invasive A. fumigatus infections is well studied, little is known about the development and progression of airway infections. Previous studies have demonstrated a critical role for the IL-1 cytokines, IL-1α and IL-1ß in enhancing pulmonary neutrophil recruitment during invasive aspergillosis. Here we use a mouse model of A. fumigatus airway infection to study the role of these IL-1 cytokines in immunocompetent mice. In the absence of IL-1 receptor signaling, mice exhibited reduced numbers of viable pulmonary neutrophils and increased levels of neutrophil apoptosis during fungal airway infection. Impaired neutrophil viability in these mice was associated with reduced pulmonary and systemic levels of G-CSF, and treatment with G-CSF restored both neutrophil viability and resistance to A. fumigatus airway infection. Taken together, these data demonstrate that IL-1 dependent G-CSF production plays a key role for host resistance to A. fumigatus airway infection through suppressing neutrophil apoptosis at the site of infection.

Diagnostic accuracy of Aspergillus-specific antibodies for chronic pulmonary aspergillosis: A systematic review and meta-analysis.

We performed this study to provide the latest evidence of the diagnostic accuracy of all Aspergillus antibodies for chronic pulmonary aspergillosis (CPA). In this meta-analysis, we searched the Cochrane Central Register of Controlled Trials, MEDLINE, Embase, and other databases, until 19 March 2020, for studies that examined the diagnostic accuracy of each Aspergillus-specific antibody for CPA and assessed the risk of bias using the revised Quality Assessment of Diagnostic Accuracy Studies-2 tool. We integrated the results using a hierarchical summary receiver operating characteristic (HSROC) model and calculated the point estimates of specificity with sensitivity fixed at 0.90 using the HSROC curve. We identified 32 published and one unpublished studies, including 75 studies on five antibody test types: 18 of precipitin test (2810 participants), 46 of IgG (8197), three of IgA (283), six of IgM (733) and two of combined IgG and IgM (IgG + IgM) (920). The results of specificity with sensitivity fixed at 0.90 were as follows: precipitin test, 0.93 (95% credible intervals: 0.86, 1.00); IgG, 0.90 (0.86, 0.95); IgA, 0.74 (0.00, 1.00); IgM, 0.50 (0.37, 0.53); IgG + IgM, 0.47 (0.00, 1.00). However, the precipitin test showed imprecision and instability in the sensitivity analysis. Most studies had a high risk of bias due to the case-control design. Although there is lack of applicability for malignancy or immunosuppressive patients, our study suggests a preference for IgG over other antibody tests in CPA screening. Particularly, IgG should be used as an adjunct when ruling out CPA.

A Murine Model for Chronic A. fumigatus Airway Infections.

In addition to causing acute invasive infections in immunocompromised patients, the mold Aspergillus fumigatus causes chronic infections in patients with chronic pulmonary conditions such as cystic fibrosis. Here we describe a non-lethal model of chronic pulmonary aspergillosis in which immunocompetent mice are endotracheally infected with A. fumigatus conidia embedded in agar beads. This approach results in the establishment of hyphal infection within the airways of mice for up to a 28-day period and is amenable to the study of innate and adaptive antifungal responses, fungal mutant strains, and antifungal agents.

NLRX1 is a key regulator of immune signaling during invasive pulmonary aspergillosis.

Aspergillus fumigatus is an opportunistic fungal pathogen of immunocompromised patient populations. Mortality is thought to be context-specific and occurs via both enhanced fungal growth and immunopathogenesis. NLRX1 is a negative regulator of immune signaling and metabolic pathways implicated in host responses to microbes, cancers, and autoimmune diseases. Our study indicates loss of Nlrx1 results in enhanced fungal burden, pulmonary inflammation, immune cell recruitment, and mortality across immuno-suppressed and immuno-competent models of IPA using two clinically derived isolates (AF293, CEA10). We observed that the heightened mortality is due to enhanced recruitment of CD103+ dendritic cells (DCs) that produce elevated amounts of IL-4 resulting in a detrimental Th2-mediated immune response. Adoptive transfer of Nlrx1-/- CD103+ DCs in neutropenic NRG mice results in enhanced mortality that can be ablated using IL-4 neutralizing antibodies. In vitro analysis of CD103+ DCs indicates loss of Nlrx1 results in enhanced IL-4 production via elevated activation of the JNK/JunB pathways. Interestingly, loss of Nlrx1 also results in enhanced recruitment of monocytes and neutrophils. Chimeras of irradiated Nlrx1-/- mice reconstituted with wild type bone marrow have enhanced neutrophil recruitment and survival during models of IPA. This enhanced immune cell recruitment in the absence of Nlrx1 is mediated by excessive production of CXCL8/IL-8 family of chemokines and IL-6 via early and enhanced activation of P38 in response to A. fumigatus conidia as shown in BEAS-2B airway epithelial cells. In summary, our results point strongly towards the cell-specific and contextual function of Nlrx1 during invasive pulmonary aspergillosis and may lead to novel therapeutics to reduce Th2 responses by CD103+ DCs or heightened recruitment of neutrophils.

Diagnostic cut-off value for Aspergillus fumigatus- and flavus-specific IgG with clinical relevance in chronic pulmonary Aspergillus infection: A pilot study in Taiwan.

BACKGROUND: The diagnostic cut-off value for chronic pulmonary Aspergillosis (CPA) by Aspergillus fumigatus-specific IgG has never been evaluated In Taiwan. The cut-off value for Aspergillus flavus-specific IgG has not been evaluated worldwide. OBJECTIVES: Evaluate diagnostic cut-off value of Aspergillus IgG and its application characteristics. PATIENTS/METHODS: Blood from control groups and treatment-naïve patients with CPA infections was collected for Aspergillus-specific IgG measurements. Controls were patients who had chest radiographic abnormalities and signs of respiratory tract infection, but were negative for Aspergillus and resolved without anti-mould therapy. Confirmation and probability of CPA were defined according to radiological features and positivity for an Aspergillus or galactomannan index. Chest computer tomography patterns were recorded for the presence of aspergilloma or nodules, subacute invasive aspergillosis, chronic cavitary pulmonary aspergillosis and chronic fibrotic pulmonary aspergillosis. RESULTS: A total of 35 cases and 50 disease controls were included. The levels of A. fumigatus- and A. flavus-specific IgG correlated with CPA progression (P < .05) but not with the presence of Aspergillus species from clinical specimens (P > .05). The best cut-off value for A. fumigatus IgG was 21.7 mg/L with area under curve (AUC) for receiver operating characteristic curve (ROC) 0.934 and had 85.7% sensitivity and 92.0% specificity. For A. flavus IgG, the best cut-off value was 22.1 mgA/L and the AUC was 0.928 with 88.2% sensitivity and 94.1% specificity. CONCLUSION: The level of Aspergillus-specific IgG correlated with radiographic characteristics in patients with CPA and the best cut-off values compared to controls were 21.7 mgA/L for A. fumigatus-specific IgG and 22.1 mgA/L for A. flavus-specific IgG.

A complex COVID-19 case with rheumatoid arthritis treated with tocilizumab.

Recurrences of COVID-19 were observed in a patient with long-term usage of hydroxychloroquine, leflunomide, and glucocorticoids due to her 30-year history of rheumatoid arthritis (RA). Tocilizumab was applied and intended to target both COVID-19 and RA. However, disease of this patient aggravated after usage of tocilizumab. After the discussion of a multiple disciplinary team (MDT) including rheumatologists, antimicrobial treatments were applied to target the potential opportunistic infections (Pneumocystis jirovecii and Aspergillus fumigatus), which were authenticated several days later via high throughput sequencing. As an important cytokine in immune responses, IL-6 can be a double-edged sword: interference in the IL-6-IL-6 receptor signaling may save patients from cytokine release storm (CRS), but can also weaken the anti-infectious immunity, particularly in rheumatic patients, who may have received a long-term treatment with immunosuppressive/modulatory agents. Thus, we suggest careful considerations before and close monitoring in the administration of tocilizumab in rheumatic patients with COVID-19. Besides tocilizumab, several disease-modifying antirheumatic drugs (DMARDs) can also be applied in the treatment of COVID-19. Therefore, we also reviewed and discussed the application of these DMARDs in COVID-19 condition.

Bronchial Epithelial Cells on the Front Line to Fight Lung Infection-Causing Aspergillus fumigatus.

Aspergillus fumigatus is an environmental filamentous fungus that can be pathogenic for humans, wherein it is responsible for a large variety of clinical forms ranging from allergic diseases to life-threatening disseminated infections. The contamination occurs by inhalation of conidia present in the air, and the first encounter of this fungus in the human host is most likely with the bronchial epithelial cells. Although alveolar macrophages have been widely studied in the Aspergillus-lung interaction, increasing evidence suggests that bronchial epithelium plays a key role in responding to the fungus. This review focuses on the innate immune response of the bronchial epithelial cells against A. fumigatus, the predominant pathogenic species. We have also detailed the molecular interactants and the effects of the different modes of interaction between these cells and the fungus.

Platelets are critical for survival and tissue integrity during murine pulmonary Aspergillus fumigatus infection.

Beyond their canonical roles in hemostasis and thrombosis, platelets function in the innate immune response by interacting directly with pathogens and by regulating the recruitment and activation of immune effector cells. Thrombocytopenia often coincides with neutropenia in patients with hematologic malignancies and in allogeneic hematopoietic cell transplant recipients, patient groups at high risk for invasive fungal infections. While neutropenia is well established as a major clinical risk factor for invasive fungal infections, the role of platelets in host defense against human fungal pathogens remains understudied. Here, we examined the role of platelets in murine Aspergillus fumigatus infection using two complementary approaches to induce thrombocytopenia without concurrent neutropenia. Thrombocytopenic mice were highly susceptible to A. fumigatus challenge and rapidly succumbed to infection. Although platelets regulated early conidial phagocytosis by neutrophils in a spleen tyrosine kinase (Syk)-dependent manner, platelet-regulated conidial phagocytosis was dispensable for host survival. Instead, our data indicated that platelets primarily function to maintain hemostasis and lung integrity in response to exposed fungal antigens, since thrombocytopenic mice exhibited severe hemorrhage into the airways in response to fungal challenge in the absence of overt angioinvasion. Challenge with swollen, heat-killed, conidia was lethal in thrombocytopenic hosts and could be reversed by platelet transfusion, consistent with the model that fungus-induced inflammation in platelet-depleted mice was sufficient to induce lethal hemorrhage. These data provide new insights into the role of platelets in the anti-Aspergillus host response and expand their role to host defense against filamentous molds.

Pulmonary infection due to Acrophialophora fusispora in a patient with underlying mixed connective tissue disease and chronic pulmonary aspergillosis: A case report and review of literature.

Acrophialophora fusispora is a soil-borne fungus rarely implicated in human infections. Here, we report a case of pulmonary infection due to A. fusispora in a 59-year-old male who presented with productive cough and gradually progressive dyspnoea for 20 days. He had a past history of pulmonary tuberculosis and was a known case of chronic obstructive pulmonary disease for past five years. He was diagnosed with mixed connective tissue disease and had been receiving oral azathioprine and prednisolone for three months. CECT thorax revealed an aspergilloma and serum Aspergillus fumigatus-specific IgG levels were raised, suggestive of chronic pulmonary aspergillosis. He was also tested positive for influenza A (H1N1) and received treatment with oral oseltamivir without any clinical benefit. Culture of sputum and bronchoalveolar lavage fluid showed growth of a fungus which was identified as Acrophialophora fusispora based on characteristic microscopic morphology and internal transcribed spacer sequencing of the ribosomal DNA. Antifungal susceptibility testing for six antifungal drugs showed itraconazole to have the most potent in vitro activity (MIC=0.25µg/mL) against A. fusispora in comparison to the other drugs tested. Treatment with itraconazole capsule 200mg twice daily was initiated and favourable clinical response was observed after 10 days of therapy. Follow-up visit after three months showed marked clinical and radiological improvement. A. fusispora is an emerging opportunistic fungus capable of causing invasive infections in immunocompromised hosts. Lack of knowledge about this fungus and confusion with morphologically similar opportunistic fungi have led to its misidentification and hence its prevalence remains largely underestimated. Accurate identification is crucial as it can help initiate early effective antifungal therapy and improve patient outcomes. To our knowledge, this is the first case of pulmonary infection due to A. fusispora reported from India.

Clinical relevance of Aspergillus fumigatus sensitization in cystic fibrosis.

RATIONALE: The clinical relevance of sensitization to Aspergillus (A) fumigatus in cystic fibrosis (CF) is unclear. Some researchers propose that specific A fumigatus IgE is an innocent bystander, whereas others describe it as the major cause of TH-2-driven asthma-like disease. OBJECTIVES: Lung function parameters in mild CF patients may be different in patients with and without A fumigatus sensitization. We aimed to ascertain whether allergen exposure to A fumigatus by bronchial allergen provocation (BAP) induces TH-2 inflammation comparable to an asthma-like disease. METHODS: A total of 35 patients, aged 14.8 ± 8.5 years, and 20 healthy controls were investigated prospectively. The patients were divided into two groups: group 1 (n = 18): specific (s)IgE negative, and group 2 (n = 17): sIgE positive (≥0.7 KU/L) for A fumigatus. Lung function, exhaled NO, and induced sputum were analysed. All sensitized patients with an FEV1 > 75% (n = 13) underwent BAP with A fumigatus, and cell counts, and the expression of IL-5, IL-13, INF-γ, and IL-8 as well as transcription factors T-bet, GATA-3, and FoxP3, were measured. RESULTS: Lung function parameters decreased significantly compared to controls, but not within the CF patient group. After BAP, 8 of 13 patients (61%) had a significant asthmatic response and increased eNO 24 hours later. In addition, marked TH-2-mediated inflammation involving eosinophils, IL-5, IL-13, and FoxP3 became apparent in induced sputum cells. CONCLUSION: Our study demonstrated the clinical relevance of A fumigatus for the majority of sensitized CF patients. A distinct IgE/TH-2-dominated inflammation was found in induced sputum after A fumigatus exposure.

Aspergillus fumigatus calcium-responsive transcription factors regulate cell wall architecture promoting stress tolerance, virulence and caspofungin resistance.

Aspergillus fumigatus causes invasive aspergillosis, the most common life-threatening fungal disease of immuno-compromised humans. The treatment of disseminated infections with antifungal drugs, including echinocandin cell wall biosynthesis inhibitors, is increasingly challenging due to the rise of drug-resistant pathogens. The fungal calcium responsive calcineurin-CrzA pathway influences cell morphology, cell wall composition, virulence, and echinocandin resistance. A screen of 395 A. fumigatus transcription factor mutants identified nine transcription factors important to calcium stress tolerance, including CrzA and ZipD. Here, comparative transcriptomics revealed CrzA and ZipD regulated the expression of shared and unique gene networks, suggesting they participate in both converged and distinct stress response mechanisms. CrzA and ZipD additively promoted calcium stress tolerance. However, ZipD also regulated cell wall organization, osmotic stress tolerance and echinocandin resistance. The absence of ZipD in A. fumigatus caused a significant virulence reduction in immunodeficient and immunocompetent mice. The ΔzipD mutant displayed altered cell wall organization and composition, while being more susceptible to macrophage killing and eliciting an increased pro-inflammatory cytokine response. A higher number of neutrophils, macrophages and activated macrophages were found in ΔzipD infected mice lungs. Collectively, this shows that ZipD-mediated regulation of the fungal cell wall contributes to the evasion of pro-inflammatory responses and tolerance of echinocandin antifungals, and in turn promoting virulence and complicating treatment options.

[Pulmonary Aspergillosis in immunocompetent patient]. / Aspergillose pulmonaire chez le patient immunocompétent.

Aspergillus is a group of molds which is found everywhere worldwide, exposition of human being is common. Most of immunocompetent people do not develop disease caused by Aspergillus. However, diseases in different forms may occur in certain conditions in immunocompetent patient. These diseases include hypersensitivity reactions, like allergic broncho-pulmonary aspergillosis, or infectious process like chronic pulmonary aspergillosis or invasive pulmonary aspergillosis. This article reviewed the clinical features of these diseases and the current available diagnostic techniques.

Aspergillus englobe un groupe de champignons ubiquitaires auquels l'être humain est quotidiennement exposé. Il est rarement pathogène chez le patient immunocompétent, cependant il peut être responsable de différentes formes de maladies dans certaines conditions. Ces pathologies comprennent des réactions d'hypersensibilité, comme l'aspergillose bronchopulmonaire allergique, ou des processus infectieux comme l'aspergillose pulmonaire chronique et l'aspergillose pulmonaire invasive aiguë. Cet article aborde les situations cliniques lors desquelles ces pathologies doivent être suspectées, tout en décrivant les caractéristiques de ces dernières et les moyens diagnostiques à disposition.

Efficiency of A fumigatus-specific IgG and galactomannan testing in the diagnosis of simple aspergilloma.

BACKGROUND: An early diagnosis of chronic pulmonary aspergillosis (CPA) at the stage of simple aspergilloma (SA) remains a challenge in low- and middle-income countries, where imaging may not be routinely available. OBJECTIVE: We investigate the role of Aspergillus fumigatus-specific IgG in serum, and galactomannan (GM) in bronchoalveolar lavage fluid (BALF) and serum for the diagnosis of SA. METHODS: We included 46 consecutive treatment-naïve subjects with SA. The 81 controls were subjects of treated pulmonary tuberculosis with residual radiological abnormality and minimal symptoms; and subjects with pulmonary disorders other than CPA who underwent bronchoscopy. The diagnosis of SA was based on consistent clinical features along with radiological manifestations (cavity with fungal ball). RESULTS: Using receiver operating characteristic (ROC) curve analysis, the best cut-off value for A fumigatus-specific IgG was 27.3 mgA/L (AUROC, 0.839; sensitivity, 63.5%; specificity, 98.3%). The best cut-off value for serum and BALF-GM was 0.7 (AUROC, 0.636; sensitivity, 32%; specificity, 96.2%) and 2.5 (AUROC, 0.833; sensitivity, 63.7%; specificity, 97.1%), respectively. A combination of A fumigatus-specific IgG (>27 mgA/L) or serum GM (≥0.7) or BALF-GM (≥2.5) had a sensitivity and specificity of 82.6% and 96%, respectively. CONCLUSIONS: A combination of serological tests has the best sensitivity in diagnosing SA. More studies are needed to confirm our findings.

Analysis of the Clinical Features of Tracheobronchial Fungal Infections with Tumor-Like Lesions.

BACKGROUND: Tracheobronchial fungal infections (TBFI) cause life-threatening complications in immunocompromised hosts but are rarely reported. Misdiagnosis and delayed antifungal treatment are associated with the high mortality rate of patients with TBFI. OBJECTIVES: This study analyzed the bronchoscopic features of TBFI and their roles in the early diagnosis of TBFI. METHODS: The demographic, clinical, radiologic, and bronchoscopic data of 53 patients diagnosed with TBFI in our department during a 15-year period were retrospectively analyzed. RESULTS: Most of the TBFI patients were male, and mass was the most common radiologic abnormality. Obvious predilection in primary bronchus distributions was observed. 41.9% of the 43 Aspergillus tracheobronchitis (AT) patients, 70% of the 10 tracheobronchial mucormycosis (TM) patients, and 100% of the 3 endobronchial cryptococcosis patients had been misdiagnosed as having cancer on bronchoscopy because of the presence of tumor-like lesions. The most common features of AT were bronchial occlusion with a mass or mucosal necrosis, bronchial stenosis with mucosal hyperplasia, or uneven mucosa. The main descriptions of TM were bronchial stenosis or obstruction due to mucosal necrosis, uneven mucosa, or a mass. The endoscopic characteristics of endobronchial cryptococcosis included occlusion due to uneven mucosa or mass, or external compressive stricture. CONCLUSION: Immunocompromised patients and immunocompetent patients with underlying disease displaying tumor-like lesions on bronchoscopy should be differentially diagnosed with cancer. Bronchial biopsy is indispensable for the early diagnosis of TBFI.

Allergic Bronchopulmonary Mycosis due to fungi other than Aspergillus.

Summary: Allergic bronchopulmonary mycosis (ABPM) is a clinical syndrome associated with immune sensitivity to various fungi that colonize the airways. Early diagnosis and treatment with systemic corticosteroids is the key in preventing the progression of the disease to irreversible lung fibrosis. Although Aspergillus has progressively gained recognition as a causative agent in past few decades, other fungi, that have been reported to cause ABPM, are not yet widely evaluated. We studied hundred and two patients with asthma for occurrence of ABPM. Patients were tested for cutaneous hypersensitivity and serum precipitin to 12 common fungal antigens. The positive cases were further evaluated for ABPM using standard criteria. Out of 102 asthma patients screened, 18 patients had either skin prick test (SPT) and/or serum precipitin positive. While 14 patients were SPT positive for one or more fungal antigen, two patients were serum precipitin positive for one or more fungi. Two patients had both serum precipitin positive as well as SPT positive. Six (5.8%) patients were diagnosed as ABPM as they fulfilled the criteria. Three of these were because of Aspergillus sp. Two were because of fungi other than Aspergillus namely Schizophyllum and Curvularia. One patient had ABPM because of both Aspergillus and Curvularia. In our study absolute eosinophil count (AEC), total IgE, serum precipitin and SPT had sensitivity of 100%, 100% 50% and 83.3% respectively for diagnosing ABPM. The specificity of these tests was 44.79%, 64.58% 98.96% and 88.54% respectively. Specfic IgE was positive in 50% of patients with either serum precipitin or SPT positivity. SPT or serum precipitin followed by specific IgE had sensitivity of 100% and specificity of 96.88% for diagnosing ABPM. SPT alone followed by Specific IgE had a sensitivity of 83.33% and specificity of 96.88% for diagnosing ABPM. We found that fungi other than Aspergillus such as schizophyllum, and curvularia, can be implicated in ABPM. Multiple fungal agents may be responsible for ABPM in an individual. There is a subset of patients of BA who have fungal sensitization but do not fulfil the criteria for ABPM. SPT was the single most sensitive and specific test, AEC >350 and total IgE more than 417IU were most sensitive tests and SPT followed by specific IgE was most effective strategy for diagnosing ABPM.