Population growth and respiration in the dust mite Dermatophagoides farinae under different temperature and humidity regimes.

Dermatophagoides farinae is an important house dust mite species that causes allergies in humans worldwide. In houses, these mites are commonly found in actively used mattresses and pillows, which provide food (i.e. sloughed skin and microorganisms), moisture, and increased temperature for faster mite development. In mattresses, feeding mites prefer the upper sector, as close as possible to the resting human (temperature 32-36 °C, humidity between 55 and 59%). However, mites that are not actively feeding prefer staying at deeper zones of the mattress. Here, we analyzed mite responses to different temperatures (15-35 °C) and relative humidity (62-94% RH) in terms of their population size growth and respiration (CO2 production) using lab mite cultures. The intrinsic rate of population increase had a single maximum at approximately 28 °C and 85% RH. At 30 °C, there were two respiration peaks at RH 90% (smaller peak) and 65% (larger peak). Therefore, there is a mismatch between the optimal temperature/humidity for the population size increase vs. respiration. We propose preliminary hypotheses explaining the two respiration peaks and suggest that future research should be done to elucidate the nature of these peaks.

Abundance of domestic mites in dwellings of children and adolescents with asthma in relation to environmental factors and allergy symptoms.

Exposure to house dust allergens, mainly from domestic mites, is an important cause of allergic reactions in sensitized asthmatic patients. A total of 63 dust samples were collected from 16 flats in Bytom (south Poland); in each flat a person (age 4-17 years) suffering from bronchial asthma lived with his/her family. Mite density was calculated as the number of specimens per g of dust. The results were compared with household features and the data were statistically analyzed. In total 566 mite specimens were isolated, including 526 members of the family Pyroglyphidae (93%). The dominant species were Dermatophagoides pteronyssinus (60% of the total count) and Dermatophagoides farinae (32%). Pyroglyphids were found in all mite positive samples (68%) of which 35% also contained non-pyroglyphids, including glycyphagids, cheyletids and gamasids. The results suggest associations between the density of some mite taxa (per g of dust) and the following indoor environmental factors: presence of pets, number of inhabitants, coal-stoves as a type of heating, cleaning frequency, higher relative humidity, presence of flowers and PVC windows. The severity of asthma seems to be associated with the numbers of D. farinae, total domestic mites and live mites per g of dust.

Temperature stress response: A novel important function of Dermatophagoides farinae allergens.

Dermatophagoides farinae, an important pathogen, has multiple allergens. However, their expression under physiological conditions are not understood. Our previous RNA-seq showed that allergens of D. farinae were up-regulated under temperature stress, implying that they may be involved in stress response. Here, we performed a comprehensive study. qRT-PCR detection indicated that 26 of the 34 allergens showed differential expression. Der f1 had the most abundant basic expression quantity. Der f 28.0201 (HSP70) and Der f3 had the same regulation pattern in 9 highly expressed transcripts, which only up-regulated at 41 °C and 43 °C, but Der f 28.0201 showed stronger regulation than Der f 3 (19.88-fold vs 6.02-fold). Whereas Der f 1, 2, 7, 21, 22, 27, and 30 were up-regulated under both heat and cold stress, and Der f 27 showed the strongest regulation ability among them. Der f 27 showed more significant up-regulation than Der f 28.0201 under heat stress (23.59-fold vs 19.88-fold), and Der f27 had more obvious up-regulation under cold than heat stress (30.70-fold vs 23.59-fold). The expression of Der f 27, 28.0201 and 1, and D. farinae survival rates significantly decreased following RNAi, indicating the upregulation of these allergens under temperature stress conferred thermo-tolerance or cold-tolerance to D. farinae. In this study, we described for the first time that these allergens have temperature-stress response functions. This new scientific discovery has important clinical value for revealing the more frequent and serious allergic diseases caused by D. farinae during the change of seasons.

Stress response and silencing verification of heat shock proteins in Dermatophagoides farinae under temperature stress.

Dermatophagoides farinae is a major exogenous allergen. Its ability to tolerate adverse external temperatures makes it responsible for widespread occurrence of allergies. Heat shock protein (HSP), a recognized temperature stress response gene, but its role in D. farinae remained unclear. Here, we performed a comprehensive study. First, we found that 25 °C was the optimal temperature, and all mites died at 48 or -20 °C for 1 h (LT100). Thus, 41 °C (LT15), 43 °C (LT25), 45 °C (LT45), and -10 °C (LT25) were selected as stress temperatures to perform de novo RNA-seq. Then, 17 main genes of the 47 differentially expressed HSP, were detected by qRT-PCR. Temperature and time gradient versus expression magnitude histogram revealed that HSP70, HSP83-1, HSP83-2, and HSP16-1 showed heat stress response only at 41-43 °C, while HSC71 and HSF played a regulatory role under both heat and cold stress, particularly HSF, with strong intensity, long duration, and quick upregulation at recovery for 10-20 min. Finally, gene expression and D. farinae survival rates significantly decreased following RNAi. These findings indicated that HSPs conferred thermo-tolerance or cold-tolerance to D. farinae. In conclusion, this was the first meaningful exploration that confirmed HSP and HSF playing an important role in temperature resistance of D. farinae.

Screening of reference genes and quantitative real-time PCR detection and verification in Dermatophagoides farinae under temperature stress.

Dermatophagoides farinae is an important source of indoor allergens that shows strong tolerance to external temperatures. However, the regularity and mechanism of tolerance are still unclear. Based on our previous RNA-seq and annotation of D. farinae under temperature stress, it is planned to identify differentially expressed genes (DEGs) involved in the temperature stress response by quantitative real-time PCR (qRT-PCR). However, the lack of reference genes directly limited the detection and confirmation of DEGs. Accordingly, in this study, we have selected six candidates as reference genes in D. farinae: 60S RP L11, 60S RP L21, α tubulin, GAPDH, Der f Mal f 6, and calreticulin, and evaluated their expression stabilities as affected by heat and cold stresses, using geNorm, NormFinder, BestKeeper, comparative ΔCt and RefFinder methods. Then the expression level of 15 DEGs were detected and verified. geNorm analysis showed that α tubulin and calreticulin were the most stable reference genes under heat stress and cold stress of D. farinae. Similar evaluation results were obtained by NormFinder and BestKeeper, in which 60S RP L21 and α tubulin were the most stable reference genes. By comparative ΔCt method and a comprehensive evaluation of RefFinder, α tubulin was identified as the most ideal reference gene of D. farinae under heat and cold stresses. Furthermore, qRT-PCR detection results of 15 DEGs were almost identical to the RNA-seq results, indicating that α tubulin is stable as a reference gene. This study provided technical support for DEGs expression studies in D. farinae using qRT-PCR.

Storage mite concentrations are underestimated compared to house dust mite concentrations.

Dwellings are increasingly well insulated to save energy and this leads to higher humidity and temperature, which improves conditions for mites. Dermatophagoides antigens are the main allergens involved and tested in atopic asthma. We developed three new species-specific quantitative PCR (qPCR) methods for house dust mites (Dermatophagoides pteronyssinus and D. farinae) and storages mites (Acarus siro, Glycyphagus domesticus, Lepidoglyphus destructor). We sampled dust with electrostatic dust collectors, in the bedrooms, under beds and in the kitchens of patients with allergies (n = 24) and healthy controls (n = 18). Mite quantification was carried out with the three new qPCRs and the qPCR previously described for the Dermatophagoides genus. The qPCRs were highly specific and efficient for house dust mite species and the storage mites. Storage mite concentrations were higher than house dust mite concentrations and were higher in dwellings of patients with allergies. Consequently, allergists should test more often patients against the storage mite antigens by prick tests or IgE serology. Dampness is a major factor in storage mite development and the presence of effective mechanical ventilation can reduce storage mite concentrations four-fold. In addition, to limit exposure to dust mites, treatments should be used throughout dwellings and not only in patients' bedrooms.

Feeding Interactions Between Microorganisms and the House Dust Mites Dermatophagoides pteronyssinus and Dermatophagoides farinae (Astigmata: Pyroglyphidae).

The feeding interactions between house dust mites (HDM) and microorganisms are key factors in the survival of mites in human environments. The suitability of different microbes for HDM is not known. Here, microbial species isolated from HDM cultures were offered to HDM in food preference tests under laboratory conditions. The microbial species were added to the rearing diet of mites to reach 7% of Saccharomyces cerevisiae and 10% of each tested microorganism. The suitability of each microbe-supplemented diet for Dermatophagoides pteronyssinus and Dermatophagoides farinae was compared in terms of mite population growth and mite preference in a cafeteria test. The effect of mite feeding on the respiration of microorganisms in the diet was observed in microcosms. HDM were able to feed and reproduce on some bacterial and fungal species, but the suitability of microbial species differed. Increasing the yeast Sa. cerevisiae in the diet from 7 to 17% appeared the most suitable for both mite species. Staphylococcus spp. bacteria were preferred for D. farinae and were suitable for reproduction. The population growth and feeding preferences of D. pteronyssinus and D. farinae with respect to microorganisms indicate that D. farinae can develop on a diet with bacterial (Staphylococcus nepalensis and Staphylococcus kloosii) additions, whereas D. pteronyssinus was successful on a diet with fungal (Aspergillus jensenii and Aspergillus ruber) additions. The bacteria Kocuria rhizophila and Bacillus cereus decreased population growth in D. pteronyssinus, whereas the yeasts Hyphopichia pseudoburtonii, Hyphopichia burtonii, and Candida ciferrii decreased population growth in D. farinae. These results indicate that some microorganisms are an important food source for HDM.

Risk of exposure of a selected rural population in South Poland to allergenic mites. Part I: indoor acarofauna of one-family houses.

The aim of this study was to investigate the occurrence of mites in dust samples from houses in agricultural areas of South Poland, with particular reference to allergenic and parasitic species as a potential risk factor of diseases among people. A total of 250 dust samples from 50 single-family houses situated in Stryszawa and vicinity (Malopolskie province) were examined for the presence of domestic mites. Dust was taken from beds, floors in bedrooms, upholstery furniture, floors in family rooms and from floors in kitchens. Mites were found in 74.8% of samples collected. A total of 5340 mite specimens were isolated, including 2771 members of the family Pyroglyphidae (51.9%). Dominants were Dermatophagoides pteronyssinus (36.4% of all mites) and Gohieria fusca (25.1%), followed by Chortoglyphus arcuatus (18.3%) and D. farinae (15.1%). Dermatophagoides pteronyssinus was also the most frequent species (53.2% of the total count of samples examined), followed by G. fusca (42.4%), D. farinae (37.2%) and C. arcuatus (36.4%). Lepidoglyphus destructor was found more frequently than Glycyphagus domesticus in the examined samples. Density of D. pteronyssinus was associated with beds, presence of pets, coal stoves used for heating, lower number of rooms, higher cooking frequency, higher washing frequency, working housewife, open kitchen, wooden floors in kitchens, lower cleaning frequency, type of upholstery furniture in living rooms (arm chairs), lower humidity and higher temperature.

Reproductive biology of Euroglyphus maynei with comparisons to Dermatophagoides farinae and D. pteronyssinus.

The reproductive biology of the house dust mite, Euroglyphus maynei, is not well studied. This mite is usually less common in homes than Dermatophagoides farinae and D. pteronyssinus. When it is present, it usually co-inhabits with the Dermatophagoides spp. and is more restricted in geographical distribution. In this study, the duration of the life cycle (egg to adult) at 23 and 30 °C at 75% relative humidity (RH) and fecundity at 23 °C and 75% RH were determined for E. maynei and the data were compared to similar data for D. farinae and D. pteronyssinus. Adults hatched from eggs after 28 days at 23 °C and 20 days at 30 °C. Females produced 1.4 eggs/day during a reproductive period of 24 days at 23 °C. Euroglyphus maynei has a shorter life cycle than D. farinae and D. pteronyssinus at 23 °C but a longer life cycle at 30 °C. Euroglyphus maynei has a shorter reproductive period and produces fewer eggs than both D. farinae and D. pteronyssinus.

Fungal food choices of Dermatophagoides farinae affect indoor fungi selection and dispersal.

House dust mite (HDM) feces and molds are the main allergens involved in allergic asthma. Differences exist between the housing fungal biome of allergic patients and standard or unhealthy housing. House dust mite (HDM) feed off spores and transport them on their bodies, but do they have fungal food preferences? We observed Dermatophagoïdes farinae in vitro with 16 mold menus and repeated the experiment 10 times. This observation led us to define Alternaria alternata, Cladosporium sphaerospermum, and Wallemia sebi as "tasty" molds and Penicillium chrysogenum, Aspergillus versicolor, and Stachybotrys chartarum as "repulsive" molds. The food preferences of D. farinae may play a role in the following two phenomena: a decrease in spore numbers due to HDM consumption and a scattering of spores that stick to the bodies of HDMs. The extent of these two phenomena should be estimated in future studies for other common domestic HDM species.

Essential oil components from Asarum sieboldii Miquel are toxic to the house dust mite Dermatophagoides farinae.

In our effort to develop novel plant-derived acaricides, we examined the contact and fumigant toxicity of Asarum heterotropoides (Asarum sieboldii Miquel) essential oil constituents to the house dust mite Dermatophagoides farinae (Acari: Pyroglyphidae). Ten constituents, including methyl eugenol (relative amount 42.18 %), were identified by gas chromatography-mass spectroscopy (GC-MS) in the A. sieboldii Miq. essential oil. In contact toxicity tests, methyl eugenol (4.2 µg/cm(2), 24 h LD50) was most toxic to D. farinae, followed by benzyl benzoate (9.1 µg/cm(2)), A. sieboldii Miq. essential oil (37.7 µg/cm(2)), and dibutyl phthalate (DBP 57.9 µg/cm(2)). The potency of methyl eugenol and A. sieboldii Miq. essential oil was higher than benzyl benzoate and DBP, with mortalities of 100, 100, 94.6, and 13.2 %, respectively, after 2.5 h of exposure. In the vapor phase mortality bioassay, methyl eugenol and A. sieboldii Miq. essential oil resulted in 100 % mortality in closed containers after 24-h exposure, but only 4.7 and 7.9 %, respectively, in open containers, indicating that the toxicity in these tests was largely due to the vapor phase. Methyl eugenol and A. sieboldii Miq. essential oil merit further study as potential D. farinae control compounds.

Interference in foraging behaviour of European and American house dust mites Dermatophagoides pteronyssinus and Dermatophagoides farinae (Acari: Pyroglyphidae) by catmint, Nepeta cataria (Lamiaceae).

The European and American house dust mites, Dermatophagoides pteronyssinus and D. farinae, have a huge impact upon human health worldwide due to being the most important indoor trigger of atopic diseases such as asthma, rhinitis and atopic dermatitis. Preceding studies have shown that the behavioural response of house dust mites towards volatile chemicals from food sources can be assessed using a Y-tube olfactometer assay. In the current study, we used this assay to investigate, for the first time, the ability of the essential oil of the catmint plant, Nepeta cataria (Lamiaceae), known to repel other ectoparasites affecting human and animal health, to interfere with the attraction of D. pteronyssinus and D. farinae towards a standard food source (fish flakes). Two distinct chemotypes (A and B), enriched in the iridoid compounds (4aS,7S,7aR)-nepetalactone and (4aS,7S,7aS)-nepetalactone, and the sesquiterpene (E)-(1R,9S)-caryophyllene, were used. Initial assays with a hexane extract of fish flakes (FF extract) confirmed attraction of mites to this positive control (P < 0.001 and P < 0.05 for D. pteronyssinus and D. farinae respectively), but when presented in combination with either N. cataria chemotype, tested across a range of doses (10, 1, 0.1 and 0.01 µg), decreasing attraction of mites to their food source was observed as the dose augmented. Our study shows that N. cataria, enriched in iridoid nepetalactones and (E)-(1R,9S)-caryophyllene, exhibits potent repellent activity for house dust mites, and has the potential for deployment in control programmes based on interference with normal house dust mite behaviour.

Allergenic Dermatophagoides mites causing asthma among schoolchildren at Ain-Shams District, Cairo, Egypt.

The present study was performed on house dust samples collected from ten homes of schoolchildren suffering from asthma at Ain-Shams district, over a period of two years (2008 & 2009). The data revealed that the total annual density of the two allergenic mites (Dermatophagoides pteronyssinus (Trouessart) & Dermatophagoides farinae (Hughes) was 202 individual mites with 22 asthmatic children in the first year, and individual mites increased to 268 with 36 asthmatic children in the second year. The two house dust mites were more abundant in bedrooms than in living ones (292 & 187 mites, respectively). The prevalence of the house dust mites on the mattresses and furniture were higher in bedrooms and living rooms than on the floors (340 & 140 mite, respectively). Winter season recorded the highest prevalence for both mites in the first and second year (87 & 110, respectively). Summer represented the lowest values (19 & 25, respectively).

The fauna and distribution of house dust mites in residential homes of Bandar Abbas District, Southern Iran.

This study was designed to determine the occurrence, distribution and abundance of house dust mites (HDM) in residential homes in Bandar Abbas (Hormozgan Province), because of numerous complaints of allergies in this oriental city. The study area was divided in five sampling zones based on population density and geographical distribution. In each sampling zone 10 houses were randomly selected. A total of 50 home dust samples were collected using a portable vacuum cleaner for 2 min from 1 m(2) of the surface of mattresses, carpets, sofas and furniture in residential houses. After collection, samples were immediately frozen. Mite species were identified and counted using standard methods and keys. Of the sampled houses 88% (44 houses) were contaminated with at least one HDM species. Three species were identified: Dermatophagoides pteronyssinus (63.1%), D. farinae (32.8%) and D. evansi (4.1%) (Pyroglyphidae). Our findings indicate a relationship between HDM density and moisture and temperature of residential places. The high contamination rate of residential houses (88%) and the favourable environmental conditions for these arthropods stress that they should be considered as important allergic causing agents.

Identification of neryl formate as the airborne aggregation pheromone for the American house dust mite and the European house dust mite (Acari: Epidermoptidae).

The American house dust mite, Dermatophagoidesfarinae Hughes, and European house dust mite, Dermatophagoides pteronyssinus Trouessart, are major pests of medical importance throughout the developed world, causing atopic diseases such as asthma, rhinitis, and atopic dermatitis. Previous studies in our laboratory have shown that the behavioral responses of house dust mites toward volatiles from food sources could be assessed using a Y-tube olfactometer assay. The current study used this Y-tube assay to investigate house dust mite pheromones. A hexane extract of D.farinae, along with fractions of the extract prepared by microscale liquid chromatography over Florisil, were tested for behavioral activity. One of the chromatographic fractions was shown to be significantly attractive (P < 0.05) for D. farinae, compared with a solvent control. Coupled gas chromatography-mass spectrometry analysis of this behaviorally active fraction indicated that neryl or geranyl formate was the major component. Peak enhancement by gas chromatography, using authentic samples of the neryl and geranyl isomers prepared in high purity by chemical synthesis, confirmed the identity of the major peak as neryl formate. In Y-tube assays, male and female D. farinae and D. pteronyssinus both were significantly attracted to synthetic neryl formate at doses of 100 and 10 ng, respectively (P < 0.05). No significant differences were found for D. farinae and D. pteronyssinus when synthetic neryl formate and house dust mite extracts containing natural neryl formate were tested at the same level. Dynamic headspace collection of D. farinae and D. pteronyssinus colonies showed that neryl formate was released as a volatile organic compound by both species. Our study shows that neryl formate is an aggregation pheromone for D. farinae and D. pteronyssinus, and has the potential to be used as part of a novel lure-and-kill system for house dust mite control.

Olfactory responses of medically and economically important mites (Acari: Epidermoptidae and Acaridae) to volatile chemicals.

Dermatophagoides farinae Hughes (Acari: Epidermoptidae), the American house dust mite, and Tyrophagus putrescentiae (Schrank) (Acari: Acaridae), the mold mite, are medically and economically important but controlling them has proved difficult, and recolonization is commonplace. Their behavioral responses to different sources of volatile chemicals are still not fully elucidated. For the first time, the Y-tube olfactometer, which is an enclosed bioassay to resolve responses to test and control volatiles, has been successfully used with these mites. Mites were tested individually, and both T. putrescentiae and D. farinae responded to food volatiles. Y-tube olfactometers may be used to test for potential semiochemicals, thereby increasing knowledge of our behavior of astigmatic mites.

Mating and fecundity of Dermatophagoides farinae.

Studies of the life cycle of cultured Dermatophagoides farinae found that after an initial mating D. farinae females lived for 63.3+/-64.6 (SD) d after their egg production period ended (Arlian and Dippold 1996). The long period after cessation of egg production for D. farinae suggested D. farinae females could mate multiple times and produce eggs continuously for a longer period. The purpose of this study was to determine if female D. farinae could mate at least two times, and subsequently increase the production of viable eggs over a longer period of time compared to a single mating. Female D. farinae were randomly selected from thriving cultures and isolated in cages. When the females had ceased to lay eggs a male was added to the cage. Fifty-seven percent of the isolated females mated again and produced a second batch of viable eggs. In natural or culture populations, females have continuous availability of males. Therefore, in another experiment, females that emerged from the tritonymphal stages were continuously exposed to fresh males and fecundity and lengths of the reproductive and post reproductive periods were determined. These females had a 11 d longer reproductive period and produced 30.7% more eggs compared to females that only mated one time after they emerged from the tritonymphal stage. However, the post reproductive period was still long (58.6+/-11.4 [SE] d) the significance of which is not clear. In conclusion, this study revealed that D. farinae females are capable of more than one successful mating that results in increased egg production compared to that of a single mating. It is likely that females mate multiple times in natural and culture populations. It was observed that females actively attract males during the reproductive period but not afterward even though she continues to live a long time.