Diurnal variation of biological and non-biological particles in the atmosphere of Córdoba, Spain

In this work, the daily distribution of biological and non-biological particles in the atmosphere of the city of Córdoba, Spain, is analysed in order to know at what time of day the concentrations of solid suspended particles in the air is maximum. This knowledge can be useful for all those people who suffer of respiratory diseases and can help them to plan their outdoors activities. The concentrations of non-biological material have been estimated by using spectrophotometrical techniques. With this method, the amount of non-biological material is expressed in percentage of light absorbance in values oscillating from 0.00: absence of particles to 1.00: extreme presence of particles. As the absorption of light depends on the colour of the particle, it is also possible to distinguish dark particles (soot, diesel exhaust particles, sand) from hyaline or light coloured ones (pollen grains and several fungal spores types). The results have shown that the maximum peaks of material are achieved early in the mornings and late in the evenings. Aerobiological methodology has been used when dealing with material of biological origin. In this latter case, pollen grains and fungal spores have been differentiated from the rest of solid material. Due to their importance as agents causing pollen allergy in the area and the high percentage that they represent with respect to the total pollen spectrum, Olea europaea and grass pollen types have been differentiated from the rest of pollen types. The results show that the peaks of non-biological particles in the air throughout the day are related to the activities carried out by men in the city: commercial and working hours or social activities in the different seasons of the year. As regards the biological material, the pattern of fungal spores is clearly influenced by the climatic characteristics of the area, presenting the maximum concentrations in the late afternoon. The pollen grains follow a pattern coinciding with the diurnal maximum temperatures and sunshine hours. The values of the curve oscillate from 200 grains/m3 of air in night hours to maximum over 1000 grains/m3 at midday. The diurnal variation of Olea europaea is quite similar to this general pattern, presenting the maximum concentrations of pollen grains at central hours of day. The grasses show some differences due to the high number of species included in this family.     

Performance of the Coriolis air sampler,a high-volume aerosol-collection system for quantification of airborne spores and pollen grains

The Coriolis δ air sampler manufactured by Bertin Technologies (France) is a continuous air sampler, dedicated to outdoor monitoring of airborne spores and pollen grains. This high-volume sampler is based on patented Coriolis technology delivering a liquid sample. The air is drawn into a conical vial in a whirling type motion using suction; particles are pulled against the wall by centrifugal force. Airborne particles are separated from the air and collected in a liquid medium. This innovative solution allows rapid analysis by several techniques including PCR assay and serological assay in order to measure the antigenicity/allergenicity of pollen grains and fungal spores. Also, traditional counting of pollen grains or taxa identification by optical microscopy can be done. A study has been carried out by the Health Protection Agency (HPA), Porton Down, UK, to measure the physical efficiency of the Coriolis air sampler. The physical efficiency of the sampler for collection of micro-organism-laden particles of various sizes has been compared with that of membrane filter samplers using the techniques described by ISO 14698-1. The Coriolis was operated simultaneously with membrane filter samplers in a controlled room where they were challenged with uniform-sized particles of different diameters containing bacterial spores. For the larger particle sizes, it was found that the physical efficiency of the Coriolis was 92% for 10-μm particles. The biological performance of the Coriolis in the collection of airborne fungal spores and pollen grains was evaluated in comparison with a Hirst spore trap (one-week tape-on-drum type sampler) which is one of the most frequently used traps in the measurement of outdoor pollen grain concentrations. The advantages and limitations of both technologies are discussed. The Coriolis was operated simultaneously with a Hirst spore trap in the sampling station of Réseau National de Surveillance Aérobiologique, France (RNSA); the pollen grain and fungal spore counts were analysed by optical microscopy. The pollen grain count m⁻³ collected was compared for both devices. The dispersion values were obtained and statistical analysis was carried out. This study shows that the Coriolis air sampler provided equivalent recovery of pollen grain and fungal spores compared with the volumetric trap standard method (not significantly different, W test, α = 0.05). Nowadays, the French-led project, acronym MONALISA, with financial support from the European Commission––Life-Environment (LIFE05 ENV/F/000068), is testing this innovative air sampler in order to measure the antigenicity/allergenicity of the main aeroallergen particles, i.e. Betula (birch), Poaceae (grasses), Parietaria (pellitory), Olea spp (olive tree), and Artemisia (mugwort) pollen grains, and Alternaria (fungal spores) to validate a new approach of monitoring instead of quantifying pollen grains by their morphology. The robustness and efficiency of the MONALISA system is being demonstrated at a national level throughout Europe in eight different countries with different bio-climatic and topography characteristics: France, UK, Finland, Poland, Spain, Portugal, Switzerland, and Italy.     

Meteorological phenomena affecting the presence of solid particles suspended in the air during winter

Winter is not traditionally considered to be a risky season for people who suffer from pollen allergies. However, increasing numbers of people are showing symptoms in winter. This prompted our investigation into the levels of solid material in the air, and some of the meteorological phenomena that allow their accumulation. This study showed a possible relationship between the phenomenon of thermal inversion, which occurs when very low temperatures, cloudless skies and atmospheric calms coincide, and an increase in the concentration of solid material in the atmosphere. Frequently, this situation is associated with other predictable phenomena such as fog, dew and frost. This may allow a warning system to be derived for urban pollution episodes. The effect caused by parameters such as wind and rainfall was also analysed. Solid material was differentiated into non-biological material from natural and non-natural sources (e.g. soot, dust, sand, diesel exhaust particles, partially burnt residues) and biological material. The latter mainly comprises pollen grains and fungal spores. Owing to its abundance and importance as a causal agent of winter allergies, Cupressaceae pollen was considered separately. Received: 28 April 1999 / Revised: 16 November 1999 / Accepted: 17 November 1999     

Contribution to the study of fungal spores in honeys of Galicia (NW Spain)

We analysed 30 samples of Galician honey with the aim of quantifying and identifying the fungal spores contained in them. Using an optical microscope, we could identify 40 different types, among them Cladosporium, Penicillium/Aspergillus and the Basidiospores, which were identified in more than 80% of the samples. We calculated the relationship between fungal spores and amount of pollen grains in the samples. The maximum values were obtained in samples 21 (more than 208/100 grains of pollen) and 5 (153 spores/100 grains of pollen).     

The aerobiological results from the 1994 cruise of the Urania (cnr) on the Adriatic. I. Pollen and spore counts on the Mediterranean sea as compared to mainland Italia

In July 1994, we were able to collect airborne fungal spores and pollen grains over the Adriatic Sea from the upper deck of the Oceanographic Ship Urania (CNR). The biological particles were collected using a modified Lanzoni VPPS 1000 sampler (operating at a flux of 10 LPM), on glycerine-gelatine coated microscopic slides. Not only were the airborne concentrations of different organisms estimated, their viability was also tested with a 1% TTC solution. Particles were collected for 60 min (i.e. a volume of 600 liters of air sampled) at every 2 h from 0600–2100 h. Up to 689 pollen grains/m³ and an impressive 48 990 spores/m³ were collected daily. Forty-two fungal taxa were identified and the most abundant spores collected were Cladosporium (82.6%), Smuts (4.8%), Ascospores (2.8%), Basidiospores (2.1%) andAlternaria (1.7%). 20 pollen taxa were identified, and the dominant pollen were Urticaceae (57.9%), Graminaceae (20.7%), Fagaceae (2.4%), Plantaginaceae (1.4%), Pinaceae (1.3%) and Eucalyptus (1.1%). The most abundant captures were done at 0800 and 1000 h (17.8 and 16.7% respectively) and at 1400 and 1600 h (13.2 and 13.8% respectively). Pollen viability per species ranged from 0 to 100%, but for the most abundant taxa, it ranged from 3.8 to 75%, and averaged 27.7%. Maximum viability was found at 0800 and 1200 h. Pollen concentrations were of the same order of magnitude as the ones found on the mainland (Brindisi, Chieti, Matera). However, its specificity was evident. Future work should therefore look more at the pollen transport process which should account for this different assemblage of pollen.     

The aerobiological results from the 1994 cruise of the Urania (cnr) on the Adriatic. I. Pollen and spore counts on the Mediterranean sea as compared to mainland Italia

In July 1994, we were able to collect airborne fungal spores and pollen grains over the Adriatic Sea from the upper deck of the Oceanographic Ship Urania (CNR). The biological particles were collected using a modified Lanzoni VPPS 1000 sampler (operating at a flux of 10 LPM), on glycerine-gelatine coated microscopic slides. Not only were the airborne concentrations of different organisms estimated, their viability was also tested with a 1% TTC solution. Particles were collected for 60 min (i.e. a volume of 600 liters of air sampled) at every 2 h from 0600–2100 h. Up to 689 pollen grains/m³ and an impressive 48 990 spores/m³ were collected daily. Forty-two fungal taxa were identified and the most abundant spores collected were Cladosporium (82.6%), Smuts (4.8%), Ascospores (2.8%), Basidiospores (2.1%) andAlternaria (1.7%). 20 pollen taxa were identified, and the dominant pollen were Urticaceae (57.9%), Graminaceae (20.7%), Fagaceae (2.4%), Plantaginaceae (1.4%), Pinaceae (1.3%) and Eucalyptus (1.1%). The most abundant captures were done at 0800 and 1000 h (17.8 and 16.7% respectively) and at 1400 and 1600 h (13.2 and 13.8% respectively). Pollen viability per species ranged from 0 to 100%, but for the most abundant taxa, it ranged from 3.8 to 75%, and averaged 27.7%. Maximum viability was found at 0800 and 1200 h. Pollen concentrations were of the same order of magnitude as the ones found on the mainland (Brindisi, Chieti, Matera). However, its specificity was evident. Future work should therefore look more at the pollen transport process which should account for this different assemblage of pollen.     

An aerobiological study of pollen grains and fungal spores of Barcelona (Spain)

Summary A study of concentration of airborne pollen grains and fungal spores has been carried out in Barcelona (Spain) during 1989–90. The volumetric method of filtration, previously described for airborne pollen analysis (Suarez-Cervera and Seoane-Camba, 1983) has been used. In this case, the filters have also been cultivated in Czapecdox-agar, Sabouraud-agar and Sabouraud-agar with streptomycin for the identification of the fungal colonies. Analysis of the number of fungal spores growing on the filter shows that the maxima of colonies of spores developed in culture per m³ of air filtered, correspond to September–December. Pollen and spore concentrations start from November–December, reach a maximum in March–April and decline progressively until September–October. Therefore, in the city of Barcelona, the greatest concentration occurs in spring and the lowest in autumn.     

Airborne pollen and spores on the Arctic island of Jan Mayen

A survey of airspora collected on Jan Mayen, an isolated North Atlantic island (71°N, 8°30′W), using a Burkard seven-day volumetric trap from 24th April to 31th August, 1988, revealed only very small concentrations. A total of 10 different pollen types were recorded, constituting a seasonal sum of 29 pollen grains. The local pollen season was confined to July, with Oxyria digna and Salix as the most numerous pollen types recorded. Exotic pollen grains, namely Betula, Pinus and Castanea type, were recorded in three periods during June and July. Studies of back trajectories indicate North America and/or Iceland and Greenland as possible source areas for the Betula pollen. There were more diatoms than pollen in the local airspora. Fungal spores mainly occurred in late July and August. Cladosporium constituted less than 5% of the total seasonal sum of fungal spores, while basidiospores contributed nearly 12%. The highest diurnal average of Cladosporium was 27 spores m^?3 air. The seasonal maximum of unidentified fungal spores reached a diurnal average of 639 spores m^?1 air on 27th August.     

Banksia pollen in the diet of Australian mammals

Mammals are frequent visitors to flowers in some Australian plant communities Nectar is usually considered to be the food sought, because pollen is thought to be of low digestibility This study compared the abilities of four non-flying mammal species to extract the protoplasts from Banksia pollen grains and quantified the amount of pollen in the diets of the species over 16 months All four species were capable of extracting the protoplast from a large proportion of the pollen grains that they ingested the mean percentage of empty Banksia pollen grains in the faeces ranged from 37% to 66% Banksia pollen was a major component in the faecal samples from two species, Cercartetus nanus and Petaurus breviceps , between March and October It occasionally comprised a large proportion of Amechmus stuartti faeces but was rare in the faeces of Rattus fuscipes Other major components of faeces included invertebrates and plant material other than pollen and fungal spores As the four mammal species tested were all from separate families and all capable of extracting the protoplast from a large proportion of the pollen grains, it is concluded that the extraction of pollen protoplasts by mammals may be widespread and may require no special adaptations     

Herbivory in Spiders: The Importance of Pollen for Orb-Weavers

Orb-weaving spiders (Araneidae) are commonly regarded as generalist insect predators but resources provided by plants such as pollen may be an important dietary supplementation. Their webs snare insect prey, but can also trap aerial plankton like pollen and fungal spores. When recycling their orb webs, the spiders may therefore also feed on adhering pollen grains or fungal spores via extraoral digestion. In this study we measured stable isotope ratios in the bodies of two araneid species (Aculepeira ceropegia and Araneus diadematus), their potential prey and pollen to determine the relative contribution of pollen to their diet. We found that about 25% of juvenile orb-weaving spiders’ diet consisted of pollen, the other 75% of flying insects, mainly small dipterans and hymenopterans. The pollen grains in our study were too large to be taken up accidentally by the spiders and had first to be digested extraorally by enzymes in an active act of consumption. Therefore, pollen can be seen as a substantial component of the spiders’ diet. This finding suggests that these spiders need to be classified as omnivores rather than pure carnivores.     

Some components of the air spora in Jamaica and their possible medical application

A knowledge of the pollen and fungal spores which comprise the air spora is useful as a preliminary approach to the problem of respiratory allergy. Therefore, this study of the qualitative and quantitative aspects of the air spora was done. Fungal spores were found to be numerically dominant, comprising 97.73% whilst pollen comprised 0.40% of the total material observed. A small number of types made up the majority of the fungal air spora, namely, Cladosporium, the Sporobolomycetaceae group, Diatrype, Glomerella, hyaline and coloured basidiospores, and septate fusiform spores. Seasonal periodicity studies on twenty-five fungal types showed that a high number of spores were trapped for sixteen during wet months, four during cooler months, and that five showed no seasonal trends. Mean diurnal periodicity studies for the year on the same twenty-five spore types showed that all had a maximum number of spores trapped at some time during the day. Investigation of the effect of rainfall on the numbers of spores released showed that the amount and duration of rainfall, the time of day rain occurs, and the length of the dry period preceding rain were of varying importance to particular spore types.     

Relationship between indoor and outdoor airborne fungal spores, pollen, and (1→3)-β-D-glucan in homes without visible mold growth

In this exploratory study, indoor and outdoor airborne fungal spores, pollen, and (1→3)-β-D-glucan levels were determined through long-term sampling (24-h) using a Button Personal Inhalable Aerosol Sampler. The air samples were collected in five Cincinnati area homes that had no visible mold growth. The total count of fungal spores and pollen in the collected samples was conducted under the microscope and Limulus Amebocyte Lysate (LAL) chromogenic assay method was utilized for the determination of the (1→3)-β-D-glucan concentration. For the combined number concentration of fungal spores and pollen, the indoor and outdoor geometric mean values were 573 and 6,435 m⁻³, respectively, with a geometric mean of the Indoor/Outdoor (I/O) ratio of .09. The geometric means of indoor and outdoor (1→3)-β-D-glucan concentrations were .92 and 6.44 ng m⁻³, respectively, with a geometric mean of the I/O ratio equal to .14. The I/O ratio of (1→3)-β-D-glucan concentration was found to be marginally greater than that calculated based on the combined number concentration of fungal spores and pollen. This suggests that (1→3)-β-D-glucan data are affected not only by intact spores and pollen grains but also by the airborne fragments of fungi, pollen, and plant material, which are ignored by traditional enumeration methodologies. Since the (1→3)-β-D-glucan level may elucidate the total exposure to fungal spores, pollen, and fungal fragments, its I/O ratio may be used as a risk marker for mold and pollen exposure in indoor environments.     

Airborne Pollen and Fungal Spores in Florist Shops in Worcester and in Bristol, UK: A Potential Problem for Occupational Health

Persistent allergies are common in workers in florist shops but little research has been done on the reasons for this. This paper reports an investigation of occupational exposure of florists to pollen and spores in three florist shops over a 2-week period in the autumn of 2000. In each shop three sampling methods were used: Burkard continuous volumetric samplers, deposition plates and low-tac tape for surface samples, including hands of the florists and leaves of a selection of the plants. The florists kept a record of the type of work they undertook each day and of the stock amounts of flowers in the shops. The volumetric traps collected 80 pollen and spore taxa. The average concentrations recorded through the working day were generally low but short-term peak (one hour mean) concentrations of some types were found to be relatively high or very high. For a few taxa these concentrations equalled or exceeded those typical for short-term peaks in wind dispersed pollen types in the ambient air. Fungal spore concentrations of several known allergenic types were also very high for peak periods. This was most notable for Aspergillus spp. which reached extremely high concentrations in one shop, compared with typical peak concentrations in the ambient atmosphere. Low-tac tape leaf samples demonstrated that the flowers' foliage is a major source of the fungal spores. Few pollen grains or fungal spores were found on the hands.     

Considerations in the grouping of plant and fungal taxa for an epidemiologic study

Although exposure to airborne pollen grains and fungal spores has been implicated as a causative factor for acute exacerbation of asthma, the few epidemiologic studies that have attempted to evaluate the relationship between these bioaerosols and asthma have used only total counts (ignoring the relative importance of different taxa) or a few predominant pollen or spore types (ignoring less abundant but potentially relevant groups). This paper reports the development of hypothesis‐driven exposure metrics (based on known aeroallergen associations with allergic asthma and other hypersensitivity diseases, pollen allergen cross‐reactivity, and the presence of local sources in the city of Fresno, California, USA) for a 3.5 year epidemiologic study of childhood asthma. Outdoor regional and neighborhood concentrations of pollen and spores were measured using Hirst‐type, 7‐day samplers. Indoor and outdoor residential concentrations were measured at 84 selected homes with similar 24‐hour slit impactors. All pollen and spore concentrations were recorded in 2‐hour intervals to assist in understanding diurnal fluctuations in aeroallergen concentrations, identify exposures during the time periods that children are outdoors, and study interaction between aeroallergens and other air contaminants, which were the primary focus of the study. The 124 pollen taxa that were observed were reduced to 15 categories and the 66 fungal and algal taxa were reduced to five categories that will be used in microenvironmental models to generate individual daily exposure estimates for each of the 315 children. These new exposure metrics will allow examination of health effects for taxa traditionally associated with allergy and those with locally elevated concentrations in combination with exposures to other indoor and outdoor air contaminants.     

Pollen molecular biology: Applications in the forensic palynology and future prospects: A review

Palynology, which is the study of pollen and spores in an archaeological or geological context, has become a well-established research tool leading to many significant scientific developments. The term palynomorph includes pollen of spermatophytes, spores of fungi, ferns, and bryophytes, as well as other organic-walled microfossils, such as dinoflagellates and acritarches. Advances in plant genomics have had a high impact on the field of forensic botany. Forensic palynology has also been used and applied more recently to criminal investigation in a meaningful way. However, the use of pollen DNA profiling in forensic investigations has yet to be applied. There were earlier uses of dust traces in some forensic analyses that considered pollen as a type of botanical dust debris. Pollen grains can be studied for comparative morphological data, clues to unexpected aspects relating to breeding systems, pollination biology and hybridization. This can provide a better understanding of the entire biology of the group under investigation. Forensic palynology refers to the use of pollen and other spores when it is used as evidence in legal cases to resolve criminal issues by proving or disproving relationships between people and crime scenes. This overview describes the various contributions and the significance of palynology, its applications, different recent approaches and how it could be further employed in solving criminal investigations.     

Pollen and Spore Incidence and Phenology in the Stockholm Area during 1972

Records of air-borne pollen and spores were made during a six month period in the Stockholm area in 1972. This paper presents phenological data and volumetric measurements obtained by means of a Burkard trap. Attention was paid to both arboreal pollen and non-arboreal pollen and spores, identified to family, genus or species level. Spores of certain common Pteridophytes were also included in the investigation, but not Bryophytes or fungal spores.     

Airborne pollen, spores, and dust across the East Mediterranean Sea

Airborne pollen and spores, as well as airflow directions, were continuously monitored during a cruise across the East Mediterranean from Tel Aviv, Israel, to Istanbul, Turkey. In spite of the fact that a high-altitude dust cloud moved, at that, time from North Africa, across the East Mediterranean, only a few dust particles were monitored on the boat. The numbers of counted airborne pollen along the cruise path were rather small. This is, in part, because the trip was taken after the main flowering season in the East Mediterranean region. Nevertheless, airborne pollen grains were still found, either as a result of remnant pollen releases by late-flowering plants or because of secondary lift-up of previously settled pollen. The presented pollen counts are average pollen counts /m³ air /6 h. The counts ranged between ∼5 pollen/m³ of air in mid-sea (July 16th–July 17th) or ∼6 pollen/m³ of air on the Israeli coast (July 16–July 17th), and 30 pollen/m³ of air near the coasts of Turkey and of the Greek Islands (July 18th–July 19th) and some 18 taxa of pollen were identified, most of them at the family level. Some 30 taxa of different spores were recorded. The numbers of airborne spores were relatively low in mid-sea (300–750 spores/m³ air), but were high near the coasts of Turkey (1,200–2,400 spores/m³ air) and of Israel (340–1,695 spores/m³ air).     

Dispersal of spores and pollen from crops

Fungal spores and pollens can be dispersed in a number of ways: by animals and insects; by water; by wind or by rain. This paper concentrates on the effects of wind on the dispersal of spores and pollen grains and the effects of rain on spore dispersal. For dispersal to be successful particles must complete three phases: removal, dispersal through the air and deposition. The biology of the organism and its environment can affect all three phases, however, once released the fate of all airborne particles largely depends on the laws of physics which govern the motion of the air. Many types of spore are actively ejected into the air while others are simply blown from the host surface. Particle size and shape affects dispersal and deposition phases. Local environmental factors such as temperature, humidity and light, as well as wind or rain, can play a key role in the removal of spores. Wind speed and turbulence or rainfall, largely determine spore dispersal, but, the size and shape of the particle, the nature of the plant canopy and the way the particles are released into the air may also be important. Particle deposition depends on both environmental and biological factors. This paper briefly considers these processes using examples and how they can be modelled.     

Terminal settling velocity and physical properties of pollen grains in still air

Numerical simulation of wind pollination requires knowledge of pollen grain physical parameters such as size, shape factor, bulk density, and terminal settling velocity. The pollen grain parameters for Japanese cedar, Japanese cypress, short ragweed, Japanese black pine, and Japanese red pine were assessed for dry condition. Terminal settling velocities of dry pollen grains in still air were measured using image analysis of scattered light tracks in a dark settling tube. The measurement system was validated by comparing results to those obtained for standard microspheres of known size and density. Dry pollen grain shape factors indicate the resemblance of particles to spheres, except for pine pollen. Circularity factors of dry pine pollen grains were 0.90–0.86, suggesting more irregular shape than those of other pollen species. Aerodynamic diameters of dry pollen grains were calculated based on the terminal settling velocity. Aerodynamic diameters of Japanese cedar, Japanese cypress, and short ragweed closely resembled the projected area equivalent diameters, suggesting that aerodynamic behaviors of these pollen grains can be managed simply in numerical simulations. However, aerodynamic diameters of dry pine pollen grains were nearly 30 % smaller than projected area equivalent diameters. Sacci on dry pine pollen can reduce the terminal settling velocity through low density and shape effects attributed to their non-sphericity, engendering aerodynamic diameter smaller by more than 10 µm from area equivalent diameters.     

Status of airborne spores and pollen in a coir factory in Kerala,India

The prevalence of airborne fungal spores and pollen grains in the indoor and outdoor environments of a coir factory in Thiruvananthapuram district of Kerala state, India was studied using the Burkard Personal Sampler and the Andersen 2-stage Sampler for 2 years (September 1997 to August 1999). The concentration of pollen grains was remarkably lower than that of fungal spores (ratio of 1:28). There was no large difference in the concentrations and types of fungal spores between the indoor and outdoor environments, with 26 spore types found to be present indoors and 27 types outdoors; of these, 22 were common to both the environments. Aspergillus/Penicillium, Cladosporium, ‘other basidiospores’ and ascospores were the dominant spore types. The total spore concentration was highest in February and lowest in September, and it was significantly higher in 1998–1999 than in 1997–1998. Twenty viable colony-forming types were isolated from inside the coir factory. The most dominant viable fungi isolated were Penicillium citrinum, Aspergillus flavus and Aspergillus niger. The total pollen concentration was higher in the outdoor environment of the coir factory than indoors, with 15 and 17 pollen types, respectively. Grass and Cocos nucifera pollen types were dominant. The dominant spore and pollen types trapped in the two environments of the coir factory are reportedly allergenic and, consequently, workers are at risk of catching respiratory/allergic diseases.