Received: 7
th
June-2014 Revised: 20
th
June-2014 Accepted: 24
th
June-2014
Research article
STUDY OF AEROMYCOFLORA IN INDOOR AND OUTDOOR ENVIRONMENT OF NATIONAL
LIBRARY, KOLKATA
Debasmita Ghosh
1*
, Priyanka Dhar
2
, Tanusree Chakraborty
1
, Naim Uddin
1
, Ashok Kumar Das
1
1
Mycology and Plant Pathology Laboratory, Department of Botany, Barasat Govt. College, West Bengal State
University, 24 PGS North, West Bengal, India
2
Defence Institute of High Altitude Research, Defence Research & Development Organisation, C/o 56 APO,
Pin 901205, Leh-Ladakh, Jammu & Kashmir, India
*
Author for correspondence: E-mail: debasmitabotany@gmail.com
ABSTRACT: We aimed at the systematic evaluation of air-borne fungal flora of the National Library, Kolkata for a
period of three months beginning from February to April, 2010 to determine their identification, concentration and
diversity in both indoor and outdoor environment to understand the cumulative aeromycoflora composition. The period
of study was the post winter period followed by pre-summer months that was mild to moderate warm and low to high
humid condition with temperature and humidity ranges of 17.0-38.2°C and 26-92% respectively. Air sample was
collected with interval of two weeks by means of gravitational settling method using petri dishes with Malt Extract Agar
(MEA) media. Fungal colonies that formed after 3-5 days incubation period at 25-28°C were identified on the basis of
micro and macro morphological characteristics and finally percentage contributions of individual fungal species were
calculated. A total of 21 types of fungal spores were identified from indoor environment with 5 sterile hyphae and 13
unidentified spore types. In case of outdoor environment, total number of spore types encountered was 19 along with 12
sterile hyphae and 6 miscellaneous types were recorded under unidentified spore type. The prevailing presence of
Aspergillus niger, Alternaria tenuissima, Cladosporium herbarum and Penicillium sp. were accounted for a high
percentage in indoor environment whereas outdoor environment showed clear dominance of Alternaria alternata,
Asperillus niger, Alternaria tenussima, Cladosporium herbarum, Cladosporium cladosporioides, Curvularia lunata and
Fusarium oxysporum. Among all the fungal spore types the taxonomic group Deuteromycotina showed dominance in
total spore contribution. Biomonitoring of aeromycoflora is a key to open the information of sensitivity towards
bioaerosol in this atmosphere and our findings may be useful with regard to the investigation of corrective measures to
save the library materials from fungal damage and diagnosis and prophylaxis of allergic diseases resulting from
aeromycoflora composition of this environment.
Key words Aeromycoflora. National Library. Indoor and outdoor environment. Fungal spores
INTRODUCTION
Aeromycoflora simply refers to the airborne fungal contributors of the environment. The term aerobiology came in use
since 1930 by the American plant pathologist Fred Cambell Meier to denote the airborne fungal spores, pollen grains
and other airborne microorganisms. It is defined as a discipline in which aerial transport of biological materials is
studied. Fungal spores represent a major fraction of bioaerosol with more than 80,000 species of which the majority are
cosmopolitan in origin [1]. Overall fungal spores have a great response in everywhere for being ubiquitous and the
concentration of aeromycoflora in different environment varies depending on the geographical regions, altitudinal
differences, seasonal variations and atmospheric conditions. A large number of airborne microfungal propagules were
found in indoor and outdoor environments and generally widely distributed in nature [2, 3]. Several microfungal species
have the potential adverse effect to cause allergies, spoilage of foods and many other adverse health effects [4-9].
International Journal of Plant, Animal and Environmental Sciences Page: 663
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Debasmita Ghosh et al Copyrights@2014 IJPAES ISSN 2231-4490
Books, papers and other documents preserved in libraries are important and valuable cultural heritage for a nation and
knowledge of all kinds in these articles are passed continuously to future generations. These are considered as precious
legacies as they have the capacity to remind people about their culture, religion and traditional ethnicity [10]. Therefore
it should be of prime importance to maintain and conserve these materials in libraries and archival settings. The
dispersing nature of fungi makes them one of the leading agents of contamination of any type of substrate in the books
of library. The aeromycoflora in the library environment causes bio-deterioration and damage of books and other
materials [11-15]. Environmental variables such as temperature, relative humidity and rainfall play vital role in the
occurrence of fungal spores in indoor air of library [10, 16]. The predominance of airborne fungi in diverse
environmental conditions especially indoor and outdoor environments of different work places and their health hazards
has recently been reviewed by Khan and Karuppayil, 2012 [9]. Investigations by previous researchers showed that some
fungi like Aspergillus candidus, A. niger, A. versicolor, Cladosporium cladosporioides, C. herbarum, Penicillium
brevicompactum and P. chrysogenum, can elevate intense allergic reactions [17] and these bio-components may cause
eye and sinus irritation, headache, tiredness, sore throat, general weakness, wooziness and severe asthma [18, 19]. To
determine the degree of precipitation of airborne fungi particles and the potential role of fungi as allergic contaminants
in book collections of libraries and archival settings, aeromycoflora of different university libraries has been studied in
different parts of the world and a diverse number of fungal species was found in these investigations [20-23]. In the
Indian scenario, the survey of airborne fungal spores in libraries of different regions of this country were carried out
with conventional techniques and a diverse number of fungal genera were identified and isolated [10, 24-30].
With this previous background, we aimed at the systematic quantification of indoor and outdoor fungal flora of the
National Library, Kolkata, India for a period of three months beginning from February to April, 2010 (Fig. 1). The aim
of this study was to determine the aeromycoflora, their identification, concentration and diversity in both indoor and
outdoor environment to understand the cumulative aeromycoflora composition in the library environment. We
hypothesized that the library environment may possess both beneficial and harmful fungal flora. In the case of beneficial
fungal species, they could be cultured and used further to produce useful substances. In contrast, in the case of harmful
fungi, their effects could be studied more critically and appropriate precautionary measures could be taken. To the best
of our knowledge, this is the first report regarding the distribution of fungal population in the National Library from
eastern India.
MATERIALS AND METHODS
Study site
The present study was conducted in the National Library, Kolkata (formerly the Calcutta Public Library, latitude 22.53°
N, longitude 88.33° E), first public library in India established in 1836. It is the largest library in India by volume and
India's library of public record. It is an institution of national importance under the Ministry of Culture, Government of
India. The library is designated to collect, disseminate and preserve the printed material produced in the country.
However, till date, there is no published record on microflora studies of this particular place which may have a
significant effect as the concentration and contamination of pathogens are related to the quality of air, ventilation,
cleanliness and maintenance of the library buildings and books along with the seasonal and environmental variations.
Although the library entrance area is more or less equivalent to natural outdoor environment but indoor section contains
a closed environment system and specially the rare books section have a typical community.
Media preparation
A suitable substrate or culture medium supporting nutritional needs of fungi was required for our study. It is possible to
establish the necessary conditions in vitro to support the optimal growth of a fungal species. For this purpose, Malt
Extract Agar (MEA) media, containing malt extract 20 gmL
-1
and agar 20 gmL
-1
of distilled water was prepared
aseptically. Then the liquid media was poured into sterile petridishes following aseptic techniques. The media was
allowed to solidify and then the junctures of the petridishes were sealed by selotape. The cool petridishes were wrapped
with brown paper and taken to the site of investigation.
Sampling procedure
The petridishes were taken to the selective site to trap the fungal composition. Air samples were obtained from six
different sites within and around the library. One petridish was used for each sampling site for each visit. The sampling
sites were majorly divided into i. indoor environment (comprising of five sampling sites viz. a. Language Division,
Bengali of Bhasha Bhavan; b. Bhasha Bhavan Reading Room; c. Central Sorting Room of Bhasha Bhavan; d.
Preservation Room of New Annexe Building and e. Ground Floor of Annexe Building, Map Section) and ii. Outdoor
environment of the National Library building.
International Journal of Plant, Animal and Environmental Sciences Page: 664
Available online at www.ijpaes.com
Debasmita Ghosh et al Copyrights@2014 IJPAES ISSN 2231-4490
The samples were collected at fifteen days intervals between the months of February to April, 2010. Samples were taken
in the afternoon (12:30 p.m.-1:30 p.m.). The Petriplate Gravitational Method was used for the isolation of fungi [31-34].
Petridishes were exposed to the air for 15 min and then covered by the lid and again sealed by sellotape. Next, these
petriplates were brought into the laboratory within 2 hr and incubated for 3-5 days at room temp (25ºC-28ºC). The
meteorological parameters like temperature (°C), relative humidity (%) and rainfall had intense effect on air-borne
fungal species both qualitatively and quantitatively and these were also recorded on each sampling date (Table 1).
•
*
KOLKAT A
•
ab
cd
ALIPORE
Fig. 1 Survey and sampling site of the study. a. India map showing location of Kolkata. b. Kolkata map (partial)
showing location of Alipore. c. Satellite map (Google map) showing location of the National Library in Alipore. d.
The National Library campus
Table 1. Temperature, humidity and rainfall on different sampling dates
Date
Temperature Humidity Rainfall
Maximum Minimum Maximum Minimum
09.02.10 30.5°C 17.5°C 89% 43% -
22.02.10 31.2°C 17.0°C 87% 26% -
11.03.10 34.0°C 22.4°C 77% 29% -
25.03.10 34.0°C 26.3°C 90% 60% -
06.04.10 38.2°C 27.7°C 92% 42% -
23.04.10 35.6°C 28.7°C 91% 59% -
Source: India Meteorological Department, Regional Meteorological Centre, Alipore, Kolkata-700027
International Journal of Plant, Animal and Environmental Sciences Page: 665
Available online at www.ijpaes.com
Debasmita Ghosh et al Copyrights@2014 IJPAES ISSN 2231-4490
Identification of fungal strains
The fungal colonies were counted based on their macro-morphological properties. The colonial features of fungal
colonies were studied minutely. Then compound microscope was used to determine the morphological structures of
fungi after mounting in lactophenol and cotton blue covered with cover slip on slides. Fungal types were analyzed for
each day. The species were identified on the basis of micro and macro morphology, reverse and surface colouration of
colonies grown on MEA media. Identification of fungi was carried out as described by previous investigators [35].
Percentage contributions of individual fungal species were calculated as per the following formula:
% Contribution = (Total number of colonies of one species / Number of colonies of all species) × 100
RESULTS AND DISCUSSION
A number of literatures on airborne fungal spores and its link with the working environments have been published from
India and abroad [36-41]. However, fungal spores in National Library has not been estimated or reported so far. In our
study our main purpose was to collect the accurate information about the fungal occurances in such a worthy site.
The results of the three months aeromycofloral survey in the National Library showed that indoor and outdoor
atmosphere of this library was never free of fungal spores. A total of 21 types of fungal spores were identified from
indoor environment with 5 sterile hyphae and rest 13 spore types which were not identified are grouped under
unidentified spore types (Table 2). In case of outdoor environment, total number of spore type encountered was 19
along with 12 sterile hyphae and 6 miscellaneous types were also recorded under unidentified spore type (Table 3).
According to their occurrence in the exposed petriplate samples, the total population in terms of percentage occurrence
were also presented in Fig 2. Alternaria tenussima (indoor-11.111% and outdoor-11.111%), Aspergillus niger (indoor-
7.936% and outdoor-13.114%) and Cladosporium herbarum (indoor-10.33% and outdoor-8.39%) were of high
occurrence in the library environment. The degrees of difference between indoor and outdoor counts of aeromycoflora
composition was calculated as indoor/outdoor ratio as suggested by previous investigators [20] and displayed in Table
4. Ratios > 1 indicate higher indoor counts whereas ratios < 1 indicate elevated outdoor concentration of fungal spore
types (Fig. 3). Alternaria humicola, Alternaria sp., Arthobotrys sp., Aspergillus ochraceus, Aspergillus sp., Chaetomium
globosum, Cladosporium sp., Drechslera sp., Geotrichum candidum, Penicillium herquei and Sirosporium sp. were
isolated only from indoor. On the other hand, Alternaria alternata, Alternaria brassicicola, Alternaria dianthi,
Alternaria dianthicola, Aspergillus candidus, Aspergillus fumigatus, Aspergillus sp., Curvularia sp., Fusarium
oxysporum, Fusarium sp. were found to be present only in outdoor environment. Alternaria tenuissima, Aspergillus sp.,
Humicola sp., Penicillium sp. and unidentified fungal spore types were present more frequently in indoor than outdoor.
Alternatively, Aspergillus flavus, Aspergillus niger, Cladosporium
cladosporioides, Cladosporium herbarum,
Curvularia lunata, Curvularia pallescens and sterile hyphae spore forms were found to be more prevalent in outdoor
section than indoor (Fig. 2, 4). Among all the fungal spore types the taxonomic group Deuteromycotina showed
dominance in the total spore contribution.
Our results showed a similar pattern with the previous studies by other researchers in different parts of the world
regarding indoor and outdoor environment [42, 43]. In our case study we got the clear picture of the diversity of fungal
spores present in indoor and outdoor environment of the library. The library was always occupied with the activity of
various types of fungi. The group Deuteromycotina or the fungi imperfecti represent the species which have thick spore
walls that may promote them to remain viable in the dust. These spores may enter into indoor environment from the
outdoor atmosphere through doors and windows, ventilation and air conditioning systems, and different fungal types
and concentrations from settled dust in normal residences were also reported [44]. The cellulose materials of the library
like books, wooden racks, cardboards etc. along with the other conditions favor the abundance of Aspergillus. Most of
the fungi imperfecti are known producer of mycotoxin and they have the ability to transmit contamination. These
contaminants had a correlation with the library atmosphere. Presence of the fungal spore types may be because of the
deficiency of cleanliness which favours the conditions for growth of aeromycoflora in the stored books and the dust
present on the books. It also depends on the meteorological parameter like temperature and relative humidity. It is a
proven association between the fungal contribution and atmospheric factors. The anthropogenic action may also be
responsible which had been reported in some early cases [45]. The concentration of airborne spore and resulting air
quality depend on the overall condition and cleanliness of the atmosphere, humidity and temperature, access to light,
ventilation, oxygen-water and other allied factors [46-51]. Additional factors like building age and mean age of books
may also have contributed to the aeromycofloral diversity that follow the same pattern reported in many libraries [20].
Many aerobiological survey showed that the existence of airborne bio-components in indoor working environments is a
major health problem in India and other parts of the world [9, 52, 53].
International Journal of Plant, Animal and Environmental Sciences Page: 666
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Debasmita Ghosh et al Copyrights@2014 IJPAES ISSN 2231-4490
The high variety of spores carries the potential of risk of many respiratory diseases. We can assume from the reference
works that the staff members and the library visitors are continuously exposing to a growing level of aero allergenic
fungal spores. In addition, the increased spore diversity is proportional to the occurrence of allergic disorders such as
allergic rhinitis, bronchial asthma, atopic dermatitis and many more heath issues related to fungal contamination. The
contaminants may provoke variety of respiratory diseases and other health problems to the staff members and persons
exposed to this environment [53]. In general, our observation showed substantial higher concentration of fungal spores
in indoor and outdoor environment of the National Library which should be emphasized for more study in the hazards
associated with human respiratory tract.
In accord to our results, previous studies in India and abroad also showed the presence of similar types of airborne
fungal species in the library atmosphere [10, 20, 24, 25, 30]. These aeromycofloral surveys were conducted with the
objective to determine the diversity of airborne fungal spores in a particular period of time and/or to estimate the
seasonal variation of aeromycoflora in the library atmosphere. Our study is unique in the sense that this is the first ever
study for the assessment of aeromycofloral diversity in the National Library of Eastern India. We have conducted the
study to monitor the distribution of airborne fungal spore types in one of the important cultural heritage of our country
to ascertain risk associated with these harmful fungal species causing damage of books, articles and other invaluable
documents of cultural ethnicity.
Table 2. Total count and percentage contribution of fungal colony from indoor environment of the National
Library, Kolkata
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Spore types
Day I
09.02.10
Day II
22.02.10
Day III
11.03.10
Day IV
25.03.10
Day V
06.04.10
Day VI
23.04.10
Total
% count of
fungal colony
Alternaria humicola
- - - - 1 - 1 1.587
Alternaria sp.
- - - 1 - - 1 1.587
Alternaria tenuissima
- 3 3 1 - - 7 11.111
Arthobotrys sp.
1 - - - - - 1 1.587
Aspergillus flavus
- - - - 1 - 1 1.587
Aspergillus niger
1 - 1 1 - 2 5 7.936
Aspergillus ochraceus
- - - - - 2 2 3.174
Aspergillus sp.
- - - - - 4 4 6.345
Chaetomium globosum
- - - - - 1 1 1.587
Cladosporium
cladosporioides
1 - - - - - 1 1.587
Cladosporium herbarum
6 - - - - - 6 9.523
Cladosporium sp.
1 - - - 1 - 2 3.174
Curvularia lunata
- - - - - 1 1 1.587
Curvularia pallescens
- - 1 - - - 1 1.587
Drechslera sp.
- - - - - 1 1 1.587
Geotrichum candidum
- - 1 - - - 1 1.587
Humicola sp.
1 - - - - 2 3 4.761
Penicillium herquei
- - - - - 1 1 1.57
Penicillium sp.
- - - 1 - 2 3 4.761
Sirosporium sp.
- - - 1 - - 1 1.587
Sterile hyphae
4 - - 1 - - 5 7.936
Unidentified
3 1 1 4 - 4 13 20.634
Total 18 4 7 11 3 20 63 99.991