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　Loop-mediated isothermal amplification (LAMP) is a useful DNA detection method with high specificity and sensitivity. The LAMP reaction is carried out within a short time at a constant temperature without the need for thermal cycling. We developed a LAMP primer set for detecting a wide range of fungi by aligning the sequences of the large subunit ribosomal RNA gene of Candida albicans (Ascomycota), Cryptococcus neoformans (Basidiomycota), and Mucor racemosus (Mucorales). The threshold of C. albicans rDNA as template with our LAMP primer set was in the range of 10-100 copies per a reaction. In this study, we evaluated the correlation between colony forming units (CFU) and LAMP detection rate using the LAMP method for environmental fungi. The LAMP method should be a useful means of detecting fungi in indoor environments, disaster areas, or even in confined manned spacecraft to prevent allergies or infections caused by fungi.

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Fungal diagnostics that utilize antibody, antigen or nucleic acid detection offer several advantages that supplement traditional culture-based methods. As a group, nonculture assays can help identify patients with invasive fungal infection (IFI) sooner than is possible with culture, are often more sensitive, and can be used to guide early interventions. Challenges associated with these techniques include the possibility for contamination or cross-reactivity as well as the potential for false negative tests. This review summarized the test characteristics and clinical utility of nonculture-based laboratory methods.

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Fusarium wilt caused by Fusarium oxysporum f. sp. niveum (Fon) is one of the major limiting factors for watermelon production worldwide. Rapid and accurate detection of the causal pathogen is the cornerstone of integrated disease management. In this paper, a real-time fluorescence loop-mediated isothermal amplification (RealAmp) assay was developed for the rapid and quantitative detection of Fon in soil. Positive products were amplified only from Fon isolates and not from any other species or formae speciales of F. oxysporum tested, showing a high specificity of the primer sets. The detection limit of the RealAmp assay was 1.2 pg µL(-1) genomic DNA or 10(3) spores g(-1) of artificially inoculated soil, whereas real-time PCR could detect as low as 12 fg µL(-1) or 10(2) spores g(-1). The RealAmp assay was further applied to detect eight artificially inoculated and 85 field soil samples. No significant differences were found between the results tested by the RealAmp and real-time PCR assays. The RealAmp assay is a simple, rapid and effective technique for the quantitative detection and monitoring of Fon in soil under natural conditions.

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O monitoramento da contaminação fúngica do arroz é imprescindível para assegurar a qualidade e segurança desse cereal. Atualmente, para avaliar a qualidade microbiológica dos alimentos, diferentes métodos têm sido propostos. Duas diferentes metodologias, plaqueamento direto e blotter test, foram comparadas quanto à eficiência em efetuar a detecção de fungos em arroz branco polido irradiado. Para realizar o blotter test, foram utilizadas placas de Petri contendo três folhas de papéis de filtro esterilizados, umedecidas em água destilada esterilizada e acrescidas com 5 mL de ágar água 0,5%. Na técnica de plaqueamento direto, os grãos foram plaqueados em meio de cultura DRBC. As amostras foram incubadas a 25ºC por sete dias e analisadas em microscópio estereoscópico. Os gêneros fúngicos presentes no arroz irradiado foram Penicillium, Aspergillus, Cladosporium, Fusarium e Trichoderma, com a predominância de Penicillium sp. e Aspergillus sp., cujas frequências foram, respectivamente, de 5,2% e 5,6% no plaqueamento direto e de 34,5% e 5,6% no blotter test. Observou-se que a irradiação gama diminuiu consideravelmente o número de grãos contaminados, sendo 96,7% pela metodologia de plaqueamento direto e até 100% pelo blotter test. O blotter test possibilitou efetuar maior contagem dos gêneros fúngicos presentes no arroz, constando-se que o método de detecção escolhido pode interferir na quantificação fúngica presente nesse cereal.

Monitoring the fungal contamination of rice is essential for ensuring the product quality and safety. Different methods have been proposed for assessing the microbiological quality of foods. Two methodologies based direct plating and blotter test were compared for detecting fungi in irradiated polished white rice. For performing the blotter test, the Petri dishes containing three sterile filter paper sheets were moistened with sterile distilled water, and 5 mL of 0.5% water-agar were added onto them. For carrying out the direct plating technique, the rice samples were plated onto DRBC medium. The samples were incubated at 25ºC for seven days, and analyzed under a stereomicroscope. The fungi genera found in irradiated rice were:Penicillium, Aspergillus, Cladosporium, Fusarium and Trichoderma, being Penicillium sp. and Aspergillus sp. the mostly predominant, with the prevalence of 5.2% and 5.6% by direct plating and 34.5% and 5.6% by blotter test, respectively. The gamma-ray irradiation significantly decreased the proportions of contaminated grains, being 96.7% by means of direct plating technique and 100% by blotter test. The blotter test showed highest efficiency in fungal genera counting found in rice samples, which evidences that the chosen detection methodology may interfere on quantifying the fungi contaminants in cereals.

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