The range of fungal species present in both asthma patients and control subjects was larger than expected. There were also clear differences in the pattern of fungal species between asthma patients and control subjects. The fungi Malassezia pachydermatis, was found in patients with asthma and not the control group. This organism has a known association with atopic conditions including atopic dermatitis . However, there were no other obvious associations were identified in the published literature between asthma and the other fungi found in the pooled samples from the asthma patients. Two of the fungi most commonly found in the sputum of asthma patients (Termitomyces clypeatus and Psathyrella candolleana) represent members of the basidiomycete family . The latter has been found in indoor dust  and one can speculate that fungal spores may have been inhaled within the home. It is possible that most of the fungi identified could have come from a single individual, or a small number of individuals, whose samples were heavily colonised by fungi.
Except for Cladosporium, the species identified in induced sputum are not commonly found in air samples examined using standard culture techniques . Analysis of air samples using molecular techniques may demonstrate that these species are commonly present in the air, but this research has not been undertaken so far. Three out of four species detected in the sputum of asthma patients were from the macromycetes group (commonly known as mushrooms). Although asthma is associated with damp environments that are affected by mould growth, we are unaware of any study that has identified an association between macromycetes and asthma. Future studies should consider analysing air samples from the homes of participants using molecular techniques, so as to take into account the presence of fungi in the ambient environment of participants.
We used universal primers for the eukaryotic 18S rRNA gene and were surprised that no eukaryotes other than fungi were identified in cases or controls. We have considered a number of potential reasons why this may be the case and the most likely was that levels of non-fungal eukaryotic DNA, present in the samples, was below the limits of detection. The PCR primers were chosen after considerable deliberation and a probeCheck test showed that they were universal and matched Homo sapiens’ 18S rRNA gene 100% . However other potential reasons include: a genuine absence of other eukaryotes and unintended removal of DNA from other eukaryotes as part of the processing of the samples.
Individual level analysis of samples was considered, but rejected as it was anticipated that, if the samples from each individual were analysed separately, the number of eukaryotes in each sample would be below the threshold of detection. Samples for this study were therefore pooled to maximise the number of copies of each species in the pooled samples and consequently maximise the probability of detecting all the species that were present.
The study has a number of weaknesses. The sputum was not fresh when it was examined and although every effort was made to prevent contamination of samples by spores in the air, this is a possibility. The sample size is small and therefore may not be representative of asthma patients. Unfortunately, information on pets was not collected in this study and therefore could not be correlated with the presence of absence of particular fungi. It is possible to speculate that the presence of a pet (particularly a dog) in the subjects' house might be associated with the presence of Malassezia pachydermatis in the sputum of the research subjects, as this organism has been identified as a commensal on the skin of dogs and have could contaminated air in the homes of some of the research participants.
The potential significance of these fungi is unclear. There is tentative emerging evidence that microbiota may form part of a complex causal web that results in disease, for example, by their effects on the immune system, without becoming pathogenic in the classical sense. For example, microbial compounds present in sputum may play a role as adjuvant factors and encourage a Th2-biased allergic response [22, 23].