Airborne cultivable microflora and microbial transfer in farm buildings and rural dwellings

Exposure to environments rich in microorganisms such as farms has been shown to protect against the development of childhood asthma and allergies. However, it remains unclear where, and how, farm and other rural children are exposed to microbes. Furthermore, the composition of the microbial flora is poorly characterised. We tested the hypothesis that farm children are exposed indoors to substantial levels of viable microbes originating from animal sheds and barns. We also expected that environmental microbial flora on farms and in farm homes would be more complex than in the homes of rural control children

Mould Routine Identification in the Clinical Laboratory by Matrix-Assisted Laser Desorption Ionization Time-Of-Flight Mass Spectrometry

BACKGROUND: MALDI-TOF MS recently emerged as a valuable identification tool for bacteria and yeasts and revolutionized the daily clinical laboratory routine. But it has not been established for routine mould identification. This study aimed to validate a standardized procedure for MALDI-TOF MS-based mould identification in clinical laboratory. MATERIALS AND METHODS: First, pre-extraction and extraction procedures were optimized. With this standardized procedure, a 143 mould strains reference spectra library was built. Then, the mould isolates cultured from sequential clinical samples were prospectively subjected to this MALDI-TOF MS based-identification assay. MALDI-TOF MS-based identification was considered correct if it was concordant with the phenotypic identification; otherwise, the gold standard was DNA sequence comparison-based identification. RESULTS: The optimized procedure comprised a culture on sabouraud-gentamicin-chloramphenicol agar followed by a chemical extraction of the fungal colonies with formic acid and acetonitril. The identification was done using a reference database built with references from at least four culture replicates. For five months, 197 clinical isolates were analyzed; 20 were excluded because they were not identified at the species level. MALDI-TOF MS-based approach correctly identified 87% (154/177) of the isolates analyzed in a routine clinical laboratory activity. It failed in 12% (21/177), whose species were not represented in the reference library. MALDI-TOF MS-based identification was correct in 154 out of the remaining 156 isolates. One Beauveria bassiana was not identified and one Rhizopus oryzae was misidentified as Mucor circinelloides. CONCLUSIONS: This work's seminal finding is that a standardized procedure can also be used for MALDI-TOF MS-based identification of a wide array of clinically relevant mould species. It thus makes it possible to identify moulds in the routine clinical laboratory setting and opens new avenues for the development of an integrated MALDI-TOF MS-based solution for the identification of any clinically relevant microorganism

International Society of Human and Animal Mycology (ISHAM)-ITS reference DNA barcoding database - the quality controlled standard tool for routine identification of human and animal pathogenic fungi

Human and animal fungal pathogens are a growing threat worldwide leading to emerging infections and creating new risks for established ones. There is a growing need for a rapid and accurate identification of pathogens to enable early diagnosis and targeted antifungal therapy. Morphological and biochemical identification methods are time-consuming and require trained experts. Alternatively, molecular methods, such as DNA barcoding, a powerful and easy tool for rapid monophasic identification, offer a practical approach for species identification and less demanding in terms of taxonomical expertise. However, its wide-spread use is still limited by a lack of quality-controlled reference databases and the evolving recognition and definition of new fungal species/complexes. An international consortium of medical mycology laboratories was formed aiming to establish a quality controlled ITS database under the umbrella of the ISHAM working group on "DNA barcoding of human and animal pathogenic fungi." A new database, containing 2800 ITS sequences representing 421 fungal species, providing the medical community with a freely accessible tool at http://www.isham.org and http://its.mycologylab.org/ to rapidly and reliably identify most agents of mycoses, was established. The generated sequences included in the new database were used to evaluate the variation and overall utility of the ITS region for the identification of pathogenic fungi at intra-and interspecies level. The average intraspecies variation ranged from 0 to 2.25%. This highlighted selected pathogenic fungal species, such as the dermatophytes and emerging yeast, for which additional molecular methods/genetic markers are required for their reliable identification from clinical and veterinary specimens.This study was supported by an National Health and Medical Research Council of Australia (NH&MRC) grant [#APP1031952] to W Meyer, S Chen, V Robert, and D Ellis; CNPq [350338/2000-0] and FAPERJ [E-26/103.157/2011] grants to RM Zancope-Oliveira; CNPq [308011/2010-4] and FAPESP [2007/08575-1] Fundacao de Amparo Pesquisa do Estado de So Paulo (FAPESP) grants to AL Colombo; PEst-OE/BIA/UI4050/2014 from Fundacao para a Ciencia e Tecnologia (FCT) to C Pais; the Belgian Science Policy Office (Belspo) to BCCM/IHEM; the MEXBOL program of CONACyT-Mexico, [ref. number: 1228961 to ML Taylor and [122481] to C Toriello; the Institut Pasteur and Institut de Veil le Sanitaire to F Dromer and D Garcia-Hermoso; and the grants from the Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq) and the Fundacao de Amparo a Pesquisa do Estado de Goias (FAPEG) to CM de Almeida Soares and JA Parente Rocha. I Arthur would like to thank G Cherian, A Higgins and the staff of the Molecular Diagnostics Laboratory, Division of Microbiology and Infectious Diseases, Path West, QEII Medial Centre. Dromer would like to thank for the technical help of the sequencing facility and specifically that of I, Diancourt, A-S Delannoy-Vieillard, J-M Thiberge (Genotyping of Pathogens and Public Health, Institut Pasteur). RM Zancope-Oliveira would like to thank the Genomic/DNA Sequencing Platform at Fundacao Oswaldo Cruz-PDTIS/FIOCRUZ [RPT01A], Brazil for the sequencing. B Robbertse and CL Schoch acknowledge support from the Intramural Research Program of the NIH, National Library of Medicine. T Sorrell's work is funded by the NH&MRC of Australia; she is a Sydney Medical School Foundation Fellow.info:eu-repo/semantics/publishedVersio

Etude des flux microbiens dans les étables de production laitière de Franche-Comté et de Bavière (intérêts dans les domaines de l'asthme et de l'allergie)

BESANCON-BU Médecine pharmacie (250562102) / SudocSudocFranceF

Performance of MALDI-TOF MS platforms for fungal identification

International audienceMatrix-assisted laser desorption/ionisation mass spectrometry (MALDI-TOF MS) is increasingly used by clinical microbiology laboratories to cope with the need for rapid, cost-effective and accurate identification of microorganisms. Several research teams have recently succeed in identifying moulds using MALDI-TOF MS, which was first adapted to bacteria, then to yeast identification. Since 2004, different commercial firms have released several ready-to-use MALDI-TOF MS platforms. This review describes the similarities and differences between the commercially available systems. In two parts, we first describe and compare the preprocessing and identification steps between the platforms and then compare the identification efficacy of yeast, moulds and dermatophytes species

Anthropogenic impact on environmental filamentous fungi communities along the Mediterranean littoral

International audienceWe hypothesised that anthropogenic influences impact the filamentous fungi community structure and that particular species or species patterns might serve as markers to characterise ecosystems. This study aimed to describe the filamentous fungi community structure in various biotopes along the Mediterranean shore that were exposed to various levels of anthropogenic influence. We sampled filamentous fungi from yellow-legged gull faecal samples at five study sites along the Mediterranean littoral in southern France. The sites were characterised by variable anthropogenic influence, ranging from building rooftops in two cities to a natural reserve. The sites also included two suburban ecoclines, one of which was exposed to sewer pollution. Filamentous fungal colonies were quantified and identified via MALDI-TOF mass spectrometry. Interestingly, we found that both fungal diversity and abundance were low in urban areas compared with suburban ecocline or environments little affected by anthropogenic influence. Furthermore, some fungal species were clearly associated with particular environments. In particular, Mucor circinelloides was associated with a natural environment with little anthropogenic impact and distant from human settlements. Whereas, Scedosporium apiospermum was associated with an ecocline polluted by sewage. Our findings indicate that particular fungal species or species combination might be used as surrogate markers of ecosystems exposed to anthropogenic pollution

Assessment of Dust Sampling Methods for the Study of Cultivable-Microorganism Exposure in Stables▿

Studies have shown a link between living on a farm, exposure to microbial components (e.g., endotoxins or β-d-glucans), and a lower risk for allergic diseases and asthma. Due to the lack of validated sampling methods, studies of asthma and atopy have not relied on exposure assessment based on culture techniques. Our objective was therefore to compare several dust sampling methods for the detection of cultivable-microorganism exposure in stables. Sixteen French farms were sampled using four different methods: (i) active air sampling using a pump, (ii) passive dust sampling with a plastic box, (iii) dust sampling with an electrostatic dust fall collector (wipe), and (iv) dust sampling using a spatula to collect dust already settled on a windowsill. The results showed that collection of settled dust samples with either plastic boxes or wipes was reproducible (pairwise correlations, 0.72 and 0.73, respectively) and resulted in highly correlated results (pairwise correlation between the two methods, 0.82). We also found that settled dust samples collected with a plastic box correctly reflected the composition of the samples collected in the air of the stable when there was no farmer activity. A loss of microbial diversity was observed when dust was kept for 3 months at room temperature. We therefore conclude that measurement of viable microorganisms within a reasonable time frame gives an accurate representation of the microbial composition of stable air

Gut yeast communities in Larus michahellis from various breeding colonies

International audienceYellow-legged gulls have been reported to carry antibiotic-resistant Enterobacteriaceae; however, the gut mycobiota of these birds has not yet been described. In this study, we analyzed the gut yeast communities in five yellow-legged gull breeding colonies along the Mediterranean littoral in southern France. Gull fecal samples were inoculated onto four types of culture media, including one supplemented with itraconazole. Yeast species richness, abundance, and diversity were estimated, and factorial analysis was used to highlight correspondences between breeding colonies. Yeast grew in 113 of 177 cultures, and 17 distinct yeast species were identified. The most frequent species were Candida krusei (53.5%), Galactomyces geotrichum (44.1%), C. glabrata (40.9%), C. albicans (20.5%), and Saccharomyces cerevisiae (18.1%). Gut yeast community structure in the gulls at both Pierre-Blanche Lagoon (PB) and Frioul Archipelago (F) were characterized by greater species richness and diversity than in those at the two cities of La Grande-Motte (GM) and Palavas-les-Flots (PF) as well as Riou Archipelago (R). Gulls in these latter three sites probably share a similar type of anthropogenic diet. Notably, the proportion of anthropic yeast species, including C. albicans and C. glabrata, in the gull mycobiota increased with gull colony synanthropy. Antifungal resistance was found in each of the five most frequent yeast species. We found that the gut yeast communities of these yellow-legged gulls include antifungal-resistant human pathogens. Further studies should assess the public health impact of these common synanthropic seabirds, which represent a reservoir and disseminator of drug-resistant human pathogenic yeast into the environment