The fungal microbiota of de-novo paediatric inflammatory bowel disease

Inflammatory bowel disease (IBD) is characterised by an inappropriate chronic immune response against resident gut microbes. This may be on account of distinct changes in the gut microbiota termed as dysbiosis. The role of fungi in this altered luminal environment has been scarcely reported. We studied the fungal microbiome in de-novo paediatric IBD patients utilising next generation sequencing and compared with adult disease and normal controls. We report a distinct difference in fungal species with Ascomycota predominating in control subjects compared to Basidiomycota dominance in children with IBD, which could be as a result of altered tolerance in these patients. 

Fungal microbiota from rain water and pathogenicity of Fusarium species isolated from atmospheric dust and rainfall dust

In order to determine the presence of Fusarium spp. in atmospheric dust and rainfall dust, samples were collected during September 2007, and July, August, and October 2008. The results reveal the prevalence of airborne Fusarium species coming from the atmosphere of the South East coast of Spain. Five different Fusarium species were isolated from the settling dust: Fusarium oxysporum, F. solani, F. equiseti, F. dimerum, and F. proliferatum. Moreover, rainwater samples were obtained during significant rainfall events in January and February 2009. Using the dilution-plate method, 12 fungal genera were identified from these rainwater samples. Specific analyses of the rainwater revealed the presence of three species of Fusarium: F. oxysporum, F. proliferatum and F. equiseti. A total of 57 isolates of Fusarium spp. obtained from both rainwater and atmospheric rainfall dust sampling were inoculated onto melon (Cucumis melo L.) cv. Piñonet and tomato (Lycopersicon esculentum Mill.) cv. San Pedro. These species were chosen because they are the main herbaceous crops in Almeria province. The results presented in this work indicate strongly that spores or propagules of Fusarium are able to cross the continental barrier carried by winds from the Sahara (Africa) to crop or coastal lands in Europe. Results show differences in the pathogenicity of the isolates tested. Both hosts showed root rot when inoculated with different species of Fusarium, although fresh weight measurements did not bring any information about the pathogenicity. The findings presented above are strong indications that long-distance transmission of Fusarium propagules may occur. Diseases caused by species of Fusarium are common in these areas. They were in the past, and are still today, a problem for greenhouses crops in Almería, and many species have been listed as pathogens on agricultural crops in this region. Saharan air masses dominate the Mediterranean regions. The evidence of long distance dispersal of Fusarium spp. by atmospheric dust and rainwater together with their proved pathogenicity must be taken into account in epidemiological studies.

Avaliação do risco de cárie e microbiota fúngica em pacientes pediátricos com anemia falciforme

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Nasal, oral and rectal microbiota of Black lion tamarins (Leontopithecus chrysopygus)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Microbiota fúngica de passeriformes de cativeiros da Região Noroeste do Estado de São Paulo

Birds are hosts for a rich fungal microbiota which can act as potent pathogens for humans and other species of animals, causing thereby serious public health problems. The objective of this study was to evaluate the participation of birds kept in containers in the epidemiology of infectious diseases such as cryptococcosis and aspergillosis, thus verifying the maintenance and spread of pathogens in the environment. 36 samples of excretas of passeriformes were collected and were cultivated in Sabouraud Dextrose Agar 4% at room temperature and 37°C. The isolated fungal colonies were classified according to their morphological and staining characteristics. Subsequently, those in yeast form were peaked in Niger Agar, incubated at 30°C. In one sample showed growth of more than one type of colony and there was verified the presence of 25.0% of Penicillium spp., 19.4% of Trichosporon spp., 13.9% of C. gattii, 11.1% of C. neoformans, 11.1% of Candida spp., 8.3% of Rhizomucor spp., 8.3% of Aspergillus spp., 2.8% of Nigrospora spp. and 2,8% of Geotrichum spp. It can be conluded by the expost that birds shed continuously pathogenic microorganisms in their feces acting in definitive form in the infectious diseases ecoepidemiology.

Avaliação do efeito da inoculação de fungos termofílicos em pilhas de compostagem de lixo urbano

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Descriptive study of onychomycosis in a hospital in São Paulo

Onychomychosis, a nail fungus infection is the most frequent nail ailment, constituting about half of all nail disorders. It can be caused by dermatophytes, non-dermatophytes, yeasts and Prothoteca spp. Methods include 5407 samples of patients with suspected onychomycosis, studied from January 2002 to December 2006, by direct mycological examination and fungi culture. The diagnosis of onychomycosis was confirmed in samples from 3822 direct mycological and/or culture positive. The diagnosis was established by culture for fungi. Among the 1.428 identified agents, the dermatophytes were responsible for 68.6% (N = 980) of cases, followed by yeasts with 27.6% (N = 394), non-dermatophytes fungi with 2.2% (N = 31), Prothoteca spp with 0.1% (N = 2), and associations with 1.5% (N = 22). Females were more affected, with 66% (N = 2527) of cases, and the most affected age group ranged from 31 to 60 years of age (median 47 years). Fungal microbiota is often changed in the world, both quantitatively and qualitatively, and is affected by several environmental factors. Thus, the periodic review of the composition of this microbiota is important to evaluate the epidemiology and thus proportion a better therapeutic response.

Human microbiota in health and disease

Each human body plays host to a microbial population which is both numerically vast (at around 1014 microbial cells) and phenomenally diverse (over 1,000 species). The majority of the microbial species in the gut have not been cultured but the application of culture-independent approaches for high throughput diversity and functionality analysis has allowed characterisation of the diverse microbial phylotypes present in health and disease. Studies in monozygotic twins, showing that these retain highly similar microbiota decades after birth and initial colonisation, are strongly indicative that diversity of the microbiome is host-specific and affected by the genotype. Microbial diversity in the human body is reflected in both richness and evenness. Diversity increases steeply from birth reaching its highest point in early adulthood, before declining in older age. However, in healthy subjects there appears to be a core of microbial phylotypes which remains relatively stable over time. Studies of individuals from diverse geopraphies suggest that clusters of intestinal bacterial groups tend to occur together, constituting ‘enterotypes’. So variation in intestinal microbiota is stratified rather than continuous and there may be a limited number of host/microbial states which respond differently to environmental influences. Exploration of enterotypes and functional groups may provide biomarkers for disease and insights into the potential for new treatments based on manipulation of the microbiome. In health, the microbiota interact with host defences and exist in harmonious homeostasis which can then be disturbed by invading organisms or when ‘carpet bombing’ by antibiotics occurs. In a portion of individuals with infections, the disease will resolve itself without the need for antibiotics and microbial homeostasis with the host’s defences is restored. The administration of probiotics (live microorganisms which when administered in adequate amounts confer a health benefit on the host) represents an artificial way to enhance or stimulate these natural processes. The study of innate mechanisms of antimicrobial defence on the skin, including the production of numerous antimicrobial peptides (AMPs), has shown an important role for skin commensal organisms. These organisms may produce AMPs, and also amplify the innate immune responses to pathogens by activating signalling pathways and processing host produced AMPs. Research continues into how to enhance and manipulate the role of commensal organisms on the skin. The challenges of skin infection (including diseases caused by multiply resistant organisms) and infestations remain considerable. The potential to re-colonise the skin to replace or reduce pathogens, and exploring the relationship between microbiota elsewhere and skin diseases are among a growing list of research targets. Lactobacillus species are among the best known ‘beneficial’ bacterial members of the human microbiota. Of the approximately 120 species known, about 15 are known to occur in the human vagina. These organisms have multiple properties, including the production of lactic acid, hydrogen peroxide and bacteriocins, which render the vagina inhospitable to potential pathogens. Depletion of the of the normal Lactobacillus population and overgrowth of vaginal anaerobes, accompanied by the loss of normal vaginal acidity can lead to bacterial vaginosis – the commonest cause of abnormal vaginal discharge in women. Some vaginal anaerobes are associated with the formation of vaginal biofilms which serve to act as a reservoir of organisms which persists after standard antibiotic therapy of bacterial vaginosis and may help to account for the characteristically high relapse rate in the condition. Administration of Lactobacillus species both vaginally and orally have shown beneficial effects in the treatment of bacterial vaginosis and such treatments have an excellent overall safety record. Candida albicans is a frequent coloniser of human skin and mucosal membranes, and is a normal part of the microbiota in the mouth, gut and vagina. Nevertheless Candida albicans is the most common fungal pathogen worldwide and is a leading cause of serious and often fatal nosocomial infections. What turns this organism from a commensal to a pathogen is a combination of increasing virulence in the organism and predisposing host factors that compromise immunity. There has been considerable research into the use of probiotic Lactobacillus spp. in vaginal candidiasis. Studies in reconstituted human epithelium and monolayer cell cultures have shown that L. rhamnosus GG can protect mucosa from damage caused by Candida albicans, and enhance the immune responses of mucosal surfaces. Such findings offer the promise that the use of such probiotic bacteria could provide new options for antifungal therapy. Studies of changes of the human intestinal microbiota in health and disease are complicated by its size and diversity. The Alimentary Pharmabiotic Centre in Cork (Republic of Ireland) has the mission to ‘mine microbes for mankind’ and its work illustrates the potential benefits of understanding the gut microbiota. Work undertaken at the centre includes: mapping changes in the microbiota with age; studies of the interaction between the microbiota and the gut; potential interactions between the gut microbiota and the central nervous system; the potential for probiotics to act as anti-infectives including through the production of bacteriocins; and the characterisation of interactions between gut microbiota and bile acids which have important roles as signalling molecules and in immunity. The important disease entity where the role of the gut microbiota appears to be central is the Irritable Bowel Syndrome (IBS). IBS patients show evidence of immune activation, impaired gut barrier function and abnormal gut microbiota. Studies with probiotics have shown that these organisms can exert anti-inflammatory effects in inflammatory bowel disease and may strengthen the gut barrier in IBS of the diarrhoea-predominant type. Formal randomised trials of probiotics in IBS show mixed results with limited benefit for some but not all. Studies confirm that administered probiotics can survive and temporarily colonise the gut. They can also stimulate the numbers of other lactic acid bacilli in the gut, and reduce the numbers of pathogens. However consuming live organisms is not the only way to influence gut microbiota. Dietary prebiotics are selectively fermented ingredients that can change the composition and/or activity of the gastrointestinal microbiota in beneficial ways. Dietary components that reach the colon, and are available to influence the microbiota include poorly digestible carbohydrates, such as non-starch polysaccharides, resistant starch, non-digestible oligosaccharides (NDOs) and polyphenols. Mixtures of probiotic and prebiotic ingredients that can selectively stimulate growth or activity of health promoting bacteria have been termed ‘synbiotics’. All of these approaches can influence gut microbial ecology, mainly to increase bifidobacteria and lactobacilli, but metagenomic approaches may reveal wider effects. Characterising how these changes produce physiological benefits may enable broader use of these tactics in health and disease in the future. The current status of probiotic products commercially available worldwide is less than ideal. Prevalent problems include misidentification of ingredient organisms and poor viability of probiotic microorganisms leading to inadequate shelf life. On occasions these problems mean that some commercially available products cannot be considered to meet the definition of a probiotic product. Given the potential benefits of manipulating the human microbiota for beneficial effects, there is a clear need for improved regulation of probiotics. The potential importance of the human microbiota cannot be overstated. ‘We feed our microbes, they talk to us and we benefit. We just have to understand and then exploit this.’ (Willem de Vos).

Microbiota fúngica de um prédio com laboratórios da região de Araraquara-SP

From September 2000 to January 2001, airborne fungi were isolated from the building of the Clinical Analyses laboratories, including its didactic and research rooms, in Araraquara São Paulo State, Brazil, by using Andersen, MAS-100® (MERCK) machine, with Sabouraud chloramphenicol medium. After 5 days of incubation at 25°C, the colonies of the fungi were counted, resulting in the identification of 21 taxa. Cladophialophora spp. was the most isolated in internal and external environments as well, followed by Penicillium spp. and Mycelia spp. In accordance with the resolution n° 9, January 2003 (ANVISA), fungi considered unacceptable were found in nine internal environments and one of these presented the amount of fungi above of the acceptable limit. Among the obtained fungi, at least 16 taxa were reported as opportunistic, nine were related to plant diseases and seven were associated to allergy problems.

Microbiota do solo em sistemas de produção de grãos no plantio direto

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Microbiota assessment in optical shops: an ignored concern to public health

The presence of microorganisms in ophthalmic instruments and surfaces can lead to the exposure of patients to several infections. However, there is no information regarding fungal and bacteria contamination in optical shops. This study aims to characterize fungi and bacteria contamination in air and surfaces from 10 optical shops covering also ophthalmic instruments. Air samples were collected through an impaction method onto malt extract agar (MEA) supplemented with chloramphenicol (0.05%) used for fungi and Tryptic Soy Agar (TSA) supplemented with nystatin (0.2%) used for bacteria. Outdoor samples were also performed to be used as reference. Surface and equipment’s swab samples were also collected side-by-side. All the collected samples were incubated at 27ºC for 5 to 7 days (fungi) or at 30º for 7 days (bacteria). Regarding fungal distribution, thirteen different species/genera were found in the air, being the most common Alternaria sp. (62.0%). Eight different species/genera were identified in the surfaces, ranging from 2 to 5x104 CFU/m2, being the most common A. versicolor complex and Penicillium sp. (40.0%). The trial frames were the most contaminated equipment, since 50.0% of the collected samples were with countless colonies. The airborne bacterial population indicated higher concentrations in the contactology office (average: 133 CFU/m3) than in the client’s waiting rooms (average: 126 CFU/m3). The surface samples indicated bacterial concentrations ranging from 2x104 to 1x106 CFU/m2, pointing out the automatic refractometer as the surface with higher bacterial load.

Fungal spore fragmentation as a function of airflow rates and fungal generation methods

The aim of this study was to characterise and quantify the fungal fragment propagules derived and released from several fungal species (Penicillium, Aspergillus niger and Cladosporium cladosporioides) using different generation methods and different air velocities over the colonies. Real time fungal spore fragmentation was investigated using an Ultraviolet Aerodynamic Particle Sizer (UVASP) and a Scanning Mobility Particle Sizer (SMPS). The study showed that there were significant differences (p < 0.01) in the fragmentation percentage between different air velocities for the three generation methods, namely the direct, the fan and the fungal spore source strength tester (FSSST) methods. The percentage of fragmentation also proved to be dependant on fungal species. The study found that there was no fragmentation for any of the fungal species at an air velocity ≤ 0.4 m/s for any method of generation. Fluorescent signals, as well as mathematical determination also showed that the fungal fragments were derived from spores. Correlation analysis showed that the number of released fragments measured by the UVAPS under controlled conditions can be predicted on the basis of the number of spores, for Penicillium and Aspergillus niger, but not for Cladosporium cladosporioides. The fluorescence percentage of fragment samples was found to be significantly different to that of non-fragment samples (p < 0.0001) and the fragment sample fluorescence was always less than that of the non-fragment samples. Size distribution and concentration of fungal fragment particles were investigated qualitatively and quantitatively, by both UVAPS and SMPS, and it was found that the UVAPS was more sensitive than the SMPS for measuring small sample concentrations, and the results obtained from the UVAPS and SMAS were not identical for the same samples.

Real time detectionof airborne fungal spores and investigations into their dynamics in indoor air

Concern regarding the health effects of indoor air quality has grown in recent years, due to the increased prevalence of many diseases, as well as the fact that many people now spend most of their time indoors. While numerous studies have reported on the dynamics of aerosols indoors, the dynamics of bioaerosols in indoor environments are still poorly understood and very few studies have focused on fungal spore dynamics in indoor environments. Consequently, this work investigated the dynamics of fungal spores in indoor air, including fungal spore release and deposition, as well as investigating the mechanisms involved in the fungal spore fragmentation process. In relation to the investigation of fungal spore dynamics, it was found that the deposition rates of the bioaerosols (fungal propagules) were in the same range as the deposition rates of nonbiological particles and that they were a function of their aerodynamic diameters. It was also found that fungal particle deposition rates increased with increasing ventilation rates. These results (which are reported for the first time) are important for developing an understanding of the dynamics of fungal spores in the air. In relation to the process of fungal spore fragmentation, important information was generated concerning the airborne dynamics of the spores, as well as the part/s of the fungi which undergo fragmentation. The results obtained from these investigations into the dynamics of fungal propagules in indoor air significantly advance knowledge about the fate of fungal propagules in indoor air, as well as their deposition in the respiratory tract. The need to develop an advanced, real-time method for monitoring bioaerosols has become increasingly important in recent years, particularly as a result of the increased threat from biological weapons and bioterrorism. However, to date, the Ultraviolet Aerodynamic Particle Sizer (UVAPS, Model 3312, TSI, St Paul, MN) is the only commercially available instrument capable of monitoring and measuring viable airborne micro-organisms in real-time. Therefore (for the first time), this work also investigated the ability of the UVAPS to measure and characterise fungal spores in indoor air. The UVAPS was found to be sufficiently sensitive for detecting and measuring fungal propagules. Based on fungal spore size distributions, together with fluorescent percentages and intensities, it was also found to be capable of discriminating between two fungal spore species, under controlled laboratory conditions. In the field, however, it would not be possible to use the UVAPS to differentiate between different fungal spore species because the different micro-organisms present in the air may not only vary in age, but may have also been subjected to different environmental conditions. In addition, while the real-time UVAPS was found to be a good tool for the investigation of fungal particles under controlled conditions, it was not found to be selective for bioaerosols only (as per design specifications). In conclusion, the UVAPS is not recommended for use in the direct measurement of airborne viable bioaerosols in the field, including fungal particles, and further investigations into the nature of the micro-organisms, the UVAPS itself and/or its use in conjunction with other conventional biosamplers, are necessary in order to obtain more realistic results. Overall, the results obtained from this work on airborne fungal particle dynamics will contribute towards improving the detection capabilities of the UVAPS, so that it is capable of selectively monitoring and measuring bioaerosols, for which it was originally designed. This work will assist in finding and/or improving other technologies capable of the real-time monitoring of bioaerosols. The knowledge obtained from this work will also be of benefit in various other bioaerosol applications, such as understanding the transport of bioaerosols indoors.