Investigation of Aspergillus niger growth and activity in a static magnetic flux density field

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1 Romanian Biotechnological Letters Vol. 16, No. 4, 2011 Copyright 2011 University of Bucharest Printed in Romania. All rights reserved ORIGINAL PAPER Investigation of Aspergillus niger growth and activity in a static magnetic flux density field Abstract 6364 Received for publication, August 12, 2010 Accepted, July 14, 2011 C. MATEESCU, N. BURUNŢEA, N. STANCU National Institute for Research and Development in Electrical Engineering ICPE-CA, 313 Splaiul Unirii Str.., Bucharest-3, , Romania, Tel: /ext. 115, Fax: , carmen.mateescu@icpe-ca.ro Researches focused on the biodegradation of materials have indicated that Aspergillus niger microscopic fungus is destructor of electrical insulations and other materials used in electrical industry in all the climatic zones. Static magnetic fields can exert some influence over microorganisms and this effect depends on many factors such as type and magnitude of magnetic field, type of microorganism, temperature, length/duration of exposure, growth media etc. The aim of the present study was to investigate the effect of the static magnetic induction field on the growth and metabolic activity of Aspergillus niger fungus. The results of the study will give a better understanding of biodegradation processes which occur on materials and electrical equipments. We investigated the growth of the fungus developed on nutritious Czapek-Dox solid media when exposed to a magnetic field of 0.50 T and 0.62 T respectively, for a period of 7 days. The exposed results were compared with those of sham controls, where the fungus developed in the same conditions as the exposed ones but without applying the magnetic field exposure. In order to assess the growing and morphology of the fungus colonies, the cultures have been macroscopically examined after 48, 72, 96 and 168 hours respectively, as well as microscopically at the end of the total exposure period. Keywords: fungus, biodegradation, static magnetic field, electrical equipment Introduction Whereas alternating current (AC) transmission lines produce AC (mostly 50 Hz) electric and magnetic fields, static electric and magnetic fields are produced by direct current (DC) transmission lines which is an economical way for power distribution over long distances. Modern magnetic levitation (maglev) systems use magnetic flux densities around 1 T directly on the rails [1]. It has to be noticed that AC as well as DC electric fields are easily reduced by all kind of obstacles and materials whereas the magnetic field not and is passing through buildings, humans, and most of the materials. Since AC and DC magnetic fields are most likely to penetrate the body, they are the component of electromagnetic fields that are usually studied in relation to the effects on living systems. It is well known that mould might cause damage to materials and equipments and create health problems in warm and humid environments. During the normal growth of fungi, there is an enzymatic secretion that accelerates a chemical transformation of organic substances such as insulation materials, wood derivates (paper), some types of seals, coatings, as well as many other types of materials from the structure of the electrical equipments. Many minerals are also destroyed by the fungus. Aspergillus niger is a fungus so widely distributed that it is ubiquitous in nature. This fungus can spread easily and colonize a wide range of substrates, including wood and plastics in the components of many devices and equipments. Aspergillus niger fungus may also be found in compost and other decaying organic material.

2 C. MATEESCU, N. BURUNŢEA, N. STANCU This paper aimed to investigate the growth and activity of Aspergillus niger fungus by exposing it under two magnetic flux densities (B-field), for a total exposure period of 168 hours. Since the present study is quite original data on a similar issue are scarce in literature. Manoliu et al (2007) [3] investigated the development of Aspergillus niger exposed to a static B-field varying from T. The results showed a times faster growth rate than in sham controls. Moreover, it has been showed that the B-field exposure can have an effect on the biodegradability of materials by enhancing the growth rate and the aggressiveness of the fungus. It has also been noticed that a B-field exposure of more than 350 T may delay the growth of microscopic fungus. The influence of the B-field exposure on cellulases and catalase activity in cellulolytic fungi Chaetomium globosum and Trichoderma viride cultivated on media with waste from industry of panification has been investigated too. It has been shown that the activity of these enzymes was influenced by both studied cellulolytic species and the exposure length to the B- field [4]. Besides Aspergillus niger fungus, the effect of static magnetic fields has also been studied at the molecular and cellular levels in bacteria species, yeasts and molds. Moreover, the growth rate, colony size, gas production, viability and mutation effects, enzyme activity and germination of spores [4] have also been investigated. The motivation of our study is double. Firstly, because the Aspergillus niger fungus is widely spread on electrical equipment in various environmental conditions and secondly because there is a lack of scientific data on the effects of static magnetic fields on this fungus. Materials and Methods For Aspergillus niger growth it has been used Czapek-Dox agar, a synthetic solid medium, containing sucrose as carbon source and nitrate as nitrogen source. On Czapek-Dox agar, colonies of Aspergillus niger consist of a compact white or yellow basal felt covered by a dense layer of dark-brown to black colonial heads. The culture medium was prepared by dissolving of the following components into one liter of distilled water: sucrose 30 g, sodium nitrate 3 g, potassium chloride 0.5 g, magnesium sulfate heptahydrate 0.5 g, iron (II) sulfate heptahydrate 0.01 g, di-potassium hydrogen phosphate 1 g and agar 15 g. The suspension was brought to the boil in order to dissolve completely all the ingredients and then was sterilized by autoclaving at C for 15 minutes [5]. The molten medium was poured into three 7 cm diameter Petri dishes. The plates were inoculated with Aspergillus niger using a sterile wire. The fungus which we used for testing was 30 days old and was obtained from our own collection. The environment conditions for fungus growth were room temperature of 25 ± 2 0 C, natural light and the humidity provided by the culture medium [6]. For studying the efficiency of the B-field, one sham control and two irradiated (exposed) inoculation plates were used. Whereas the sham inoculated plate was treated in the same way as the irradiated ones but without applying the B-field, the two other inoculated plates were exposed to a B-field of 0.5 T and 0.62 T respectively. Though the total exposure time was 168 hours, the cultures were periodically analyzed after 48, 72, 96 and 168 hours respectively. After incubations all the cultures were microscopically analyzed as regards the Aspergillus niger sporulation growth, using an optical inverted microscope type Nikon Eclipse Ti-E fitted with a confocal system Eclipse C1si. The microscopic examination of the fungus was performed in bright field with a magnitude of 40 X. In order to make a comparative analysis of Aspergillus niger growth under different static magnetic flux densities, two magnetic systems have been calculated and designed for Romanian Biotechnological Letters, Vol. 16, No. 4,

3 Investigation of Aspergillus niger growth and activity in a static magnetic flux density field performing exposures to B-field of 0.5 T and 0.62 T. These magnetic systems consisted of the following components: a. Magnetic circuit made of steel type OL 35; b. Permanent magnets, made of magnetic alloy based on NdFeB, having the following magnetic field characteristics: Br = 1.2 T, Hcb = 970 ka/m, (BH) max = 35 MGsOe, respectively; c. Polar pieces made of steel type OL 35. The exposure strength of the B-field in air has been performed by simulation of magnetic fields using a magnetic calculation soft type FEM (Finit Element Method). Results and discussions For the investigations of the effect of a static B-field on the mycelial growth and conidia formation, the plates were examined after an incubation period of 48, 72, 96 and 168 hours respectively. Figures 1-4 (a,b,c) show the Aspergillus niger growth without B-field exposure (sham control), under a B-field exposure of 0.5 T and of 0.62 T, for 48, 72, 96 and 168 hours respectively. During the first 48 hours we observed that the non-exposed fungus has grown faster than the two other samples which were exposed to the B-field. It means that after an incubation of 48 hours, the growth of Aspergillus niger was characterized by the development of small but compact colonies with dense sporulation, spread from the inoculated point to the whole surface of the culture medium (Figure 1 a). The fungus exposed to the B-field produced less but larger colonies, with strong sporulation. The colonies are not spread on the whole surface of the culture medium (Figures 1b, 1c). Fig 1. (a,b,c) Aspergillus niger growth after 48 hours of incubation (a -without magnetic field; b magnetic field of 0.5T; c magnetic field of 0.62T) After a B-field exposure of 72 hours, the mould followed an atypical growth, characterized by about 20 larger and bombastic colonies having white-yellowish normal aspect and very few dark-brown colonial heads. A stronger B-field exposure resulted in a larger but rarer colony formation in the culture medium. Thus, for the plate exposed to a B- field of 0.5 T (Figure 2b), we observed more but smaller colonies which tend to join each other, comparing to the sample exposed in B-field of 0.62 T (Figure 2c). By comparing the 0.5 T (Figure 2b) and the 0.62 T exposure results, we observe in the first case more but smaller colonies which tend to stick together than in the second case Romanian Biotechnological Letters, Vol. 16, No. 4, 2011

4 C. MATEESCU, N. BURUNŢEA, N. STANCU Fig 2. (a,b,c) Aspergillus niger growth after 72 hours of incubation (a -without magnetic field; b magnetic field of 0.5; c magnetic field of 0.62T) After 96 hours of incubation, some notches or cuts became visible on the swollen surface of the colonies. They are more explicit for the stronger B-field of 0.62 T (Figure 3c). These notches did not arise when the B-field was not applied. Fig 3. (a,b,c) Aspergillus niger growth after 96 hours of incubation (a -without magnetic field;b magnetic field of 0.5T; c magnetic field of 0.62T) After an incubation period of 168 hours (at the end of testing), we observed a relatively homogenous growth of the fungus in the plate not exposed to the B-field (Figure 4a). The colonies became completely black on the whole surface of the culture medium. The sample exposed to the strongest B-field of 0.62 T (Figure 4c) showed intensively black colored and large colonies. In this case the black colonial heads are denser as compared to the fungus developed in cultures not exposed to a B-field. This atypical growth does not cover the entire surface of the Petri dish. The sample exposed in the B-field of 0.5 T (Figure 4b) shows an intermediary aspect between the two plates. Fig 4. (a,b,c) Aspergillus niger growth after 168 hours of incubation (a -without magnetic field; b magnetic field of 0.5T; c magnetic field of 0.62T) Romanian Biotechnological Letters, Vol. 16, No. 4,

5 Investigation of Aspergillus niger growth and activity in a static magnetic flux density field Figure 5 shows a microscopic photograph of radiate conidial head and brown-colored round-shaped unicellular conidia of Aspergillus niger. Fig. 5. Micrograph of Aspergillus niger conidia with a full-field on the left and a detail shot on the right Conidia in chains or detached and dispersed can be observed in this microscopy image. Single or paired conidia may resemble yeast cells. The size of spores of Aspergillus niger ranges from 2 to 4 microns. Conclusions The effect of a static magnetic flux density on the growth and activity of Aspergillus niger have been macroscopically and microscopically investigated by means of two exposure strengths. It is concluded that the static magnetic field produces an atypical growth of the fungus that is characterized by less and swollen, bombastic colonies which did not spread on the entire surface of the culture medium. From the comparison between the exposed and the sham, we conclude that the magnetic field is an efficient inhibitor of the surface growth of the fungus. The validation or practical application of the present study lies in the fact that the results can be used for developing magnetic field sources/methods that are able to reduce or eliminate bio-damage of components of electrical equipments or installations and materials which are sensitive to Aspergillus niger and perhaps to other fungi. References 1. ICNIRP 13/2003. Exposure to Static and Low Frequency Electromagnetic Fields. Biological Effects and Health Consequences (0-100 khz). Publication of the International Commission on Non-Ionizng Radiation Protection. Printed by Märkl-Druck, München. 2. S. A. SEMENOV,KLARA Z. GUMARGALIEVA,GENNADIĬ EFREMOVICH ZAIKOV, Biodegradation and durability of materials under the effect of microorganisms, VSP BV, ISBN , pp , (2003). 3. A. MANOLIU, L. OPRICK, D. CREANGA, The influence of the static magnetic field (SMF) on some biochemical parameters in cellulolytic fungi Chaetomium globosum and Trichoderma viride cultivated on media supplemented with panification industrial wastes, Rom. Journal Biol, Vol , 2007, pp P.E KOVACS., R.L VALENTINE., P.J. ALVAREZ, The effect of static magnetic fields on biological systems: Implications for enhanced biodegradation, Crit. Rev. Environ. Sci. Technol., 27, 1997, pp zapek_dox_agar.pdf 6. M. MITITIUC, C. TANASE, Micologie, Editura Univ. Al. I. Cuza, Iasi, 2000, pp Romanian Biotechnological Letters, Vol. 16, No. 4, 2011