Jan A. Wendt1 Liliana Indrie2 Paula Dejeu3 Adina Albu2 Dorina Camelia Ilieș4 Monica Costea5 Tudor Caciora4,* Gabriela Ilieș6 Nicolaie Hodor7 Ioana Josan4 Zharas Berdenov8 Vasile Grama4 Bahodir Safarov9 Natural Sources in Preventive Conservation of Naturally Aged Textiles DOI: 10.5604/01.3001.0014.9309 Abstract Natural antimicrobials can eliminate fungi and prevent the aging of cotton fabrics. While fungi can cause severe infections to the fabric user etc, natural antimicrobials have the advantage of not being toxic to humans. The present study showed that the essential oils of lemon (Citrus limon), lavender (Lavandula angustifolia) and mint (Mentha piperita) have inhibitory effects on yeast and mould spores on a piece of textile from Romanian cultural heritage, “ie”, stored in a space within an ethnographic museum., inhibitory action against Botrytis sp., the inhibitory effect of lemon essential oil on Cladosporium sp. and that of peppermint essential oil on yeast species Rhododtorula mucilaginosa were demonstrated, respectively. Being environmentally friendly, these sources, can be tested on a large scale. Key words: essential oils, cotton, antibacterial agents, health, prevention and conservation. 1 Gdansk University, Faculty of Oceanography and Geography, Institute of Geography, 4 Bażynskiego Street, 80309, Gdansk, Poland, e-mail: jan.wendt@ug.edu.pl 2 University of Oradea, Faculty of Energy Engineering and Industrial Management, Department of Textiles, Leather and Industrial Management, 4 Barbu Stefanescu Delavrancea Street, 410058, Oradea, Bihor County, Romania, e-mail: lindrie@ uoradea.ro, aalbu @uoradea.ro 3 Medical Laboratory service M.D., Bethany Medical Clinic Oradea, 12 Menumorut Street, 410004, Oradea, Bihor County, Romania, e-mail: office@betania-centrumedical.ro 4 University of Oradea, Faculty of Geography, Tourism and Sport, Department of Geography, Tourism and Territorial Planning, 1 Universitatii Street, 410087, Oradea, Bihor County, Romania, e-mail: dilies@uoradea.ro, tudor.caciora@yahoo.com, ijosan@uoradea.ro, vgrama@uoradea.ro 5 University of Oradea, Faculty of Environmental Protection, 26 General Magheru Boulevard, 410035, Oradea, Bihor County, Romania, e-mail: mcostea @uoradea.ro 6 Babes-Bolyai University, Sighetu Marmației extension, 6 Avram Iancu Street, 435500, Sighetu Marmației, Maramures County, Romania, e-mail: gabriela.ilies@geografie.ubbcluj.ro 7 University “Babes-Bolyai” Cluj Napoca, Faculty of Geography, Street Clinicilor no 5-6, Cluj Napoca, Cluj County, Romania, e-mail: nicolaie.hodor@ubbcluj.ro 8 L. N. Gumilyov Eurasian National University, Department of Physical and Economic Geography, Faculty of Science, 2 Satpayev Street, 010008, Nur-Sultan, Republic of Kazakhstan, e-mail: berdenov-z@mail.ru 9 Samarkand State University, Faculty of Human Resources Management, Department of Digital Economy, Samarkand, Uzbekistan, 40104, University blv. 15, Samarkand, e-mail: saforvb@rambler.ru 80 Introduction Among the many elements of tangible cultural heritage specific to the Beius Depression, Romania, the traditional women shirt called ‘IA’ stands out. It is a part of the cultural heritage with implications for shaping place-specific identity [1-4]. ‘IA’ is also an indisputable historical landmark which provides information about the people who made and wore such garments, their social status, the level of knowledge and understanding of the universe in which they lived and worked, etc. [5-7]. Museums, especially ethnographic museums with their clothes, fabrics, throws, scarves, etc., classic for the region, play an important role in education and preserving the original cultural heritage of each country and region [8]. Therefore, the preventive conservation of naturally aged textiles is of fundamental importance for the protection and preservation of the material heritage of the region. The high collections presented are also important in shaping regional tourist centers [9] and the potential for cultural heritage marketing [10-12] influencing the choice of destinations preferred by tourists. Textiles are exposed to many challenges, such as indoor temperature and relative humidity fluctuations, light impact, insects, dust and particulate matter, nonstandard storage and display, housekeeping methods, very poor restoration and conservation etc [13]. In this context, the present study aimed at non-invasive analysis in order to biore- mediate an ‘ia’ clothing element located in the depository of the Beius Municipal Museum (Figure 1). The importance of the study lies in the need to preserve and protect such elements of tangible heritage of national and even international importance and of inestimable value. Antiquity together with time and the simultaneous action of some factors related to microclimate, microorganisms and the ‘human’ represent causes that endanger the integrity of such textile creations. Hence, the need arises to carry out interdisciplinary studies (by conservators, chemists, microbiologists, geographers, etc.), as non-invasively as possible, in order to preserve these artefacts for posterity. An assessment of contamination and the effects of essential oils on potentially existing fungi on the surface of the material was conducted. The microscopic appearance of threads in the area with the most damaged macroscopic appearance, precisely the inner left fibre, was further evaluated. Microscopic images are shown in Figure 1. The microscopic images (x 20; x 40) (Figure 1) of threads taken from the inner left sleeve respectively show the deterioration of their quality. Due to single broken fibres, the arrangement of fibres visible under the microscope is disordered. Fibres do not form a bundle, indicating their location of mechanical damage (left picture). Visible is a change in its arrangement, probably under the influence of chemical factors [14]. Several cracks oblique to the fibre axis can be observed, and there are cracks propagating from the constriction in both directions of the fibre (Figure 1, right down). Wendt JA, Indrie L, Dejeu P, Albu A, Ilieș DC, Costea M, Caciora T, Ilieș G, Hodor N, Josan I, Berdenov Z, Grama V, Safarov B. Natural Sources in Preventive Conservation of Naturally Aged Textiles. FIBRES & TEXTILES in Eastern Europe 2021; 29, 5(149): 80-85. DOI: 10.5604/01.3001.0014.9309 existing fungi on the surface of the material was conducted. acteristics and giving-off of a good smell [25]. Gutarowska et al. [1] mentions in Hammer et al. [15] tested 52 natural veg- her research paper the short disinfection etal oils and plant extracts, showing very effect of essential oils; however, most good antimicrobial and preservation ac- importantly, the tests conducted were tivity. Mimica-Dukic et al. [16] analysed environmentally friendly. In Romania, the antimicrobial activities of essential oil Iordache et al. [26] showed the great pofrom Mentha Sp; Bozin et al. [17, 18] dis- tential of Rosemary and Orange oils for played the antioxidant and antimicrobial obtaining very good antimicrobial textile activities of essential oils such as Oci- effects [27]. Hercules and Papadopoulou mum basilicum L., Origanum vulgare L., [28] analysed the antimicrobial proprieand Thymus vulgaris L., as well as Rose- ties of Basil, Oregano, and Thyme essenmary, Sage (Rosmarinus officinalis L. tial oils. Nazzaro et al. [29] describes the and Salvia officinalis L., Lamiaceae). antifungal properties of essential oils, as Abdel-Kareem [19] tested the preven- well as their importance in the commution/inhibition of biodeterioration and nicative inhibition of the cellular system, the improvement of textiles properties in the creation of biofilms, and in the prousing polymers in combination with bi- duction of mycotoxins. Andra et al. [30] ocides, the results of which being among obtained crude extract from Pangamic the best. The effects of cleaning mate- pinnate and evaluated its antibacterial rials containing natural dyes were also activity against E. coli, upon being coattested for silk fabric stored in unsuitable ed on cotton fibre. Elsayed and Shabana conditions in the Islamic Museum [20]. [31] studied the effects of some essential Mahesh et al. [21] studied antimicrobial oils on the fungi contamination (Aspertextile finish using natural plant products. gillus niger and Alternaria alternate) of Four species of Laminaceea: Pogostemon archaeological heritage objects. Othman cablin, Lavandula angustifolia, Melissa et al. [27] concludes in his/her study that officinalis, and Salvia officinalis, native the essential oils tested can be successto Pakistan, were tested To assess the by Hussain et fully used for fungicidal preparations for al. [22], and showed antioxidant action. the disinfection of biodeteriorated linen Some biotreatments and fungal washing and papyrus artefacts. Pereira et al. [32] of heavy metals in wastewaters were ex- ``demonstrated that the phenolic comamined by Gupta et al. [23].[3 The paper ] of pounds in olive leaves have antimicrobial Pannu [24] analysed and proposed natu- action (Olea europaea L. Cv. Cobrançoral solutions for antimicrobial finishes. sa)”, [3 even ]in lower concentrations. Stan et al. [33] and Thilagavathi and Bala Csapek Dox sterile culture media, Lavender, thyme, and clove essential oils [34] mention the use of the microencapare extracted from natural sources and sulation of essential oils in textiles with API® AUX kit fabrics for yeast identification, used in the20C treatment of cotton promising antioxidant and antibacterial for the attainment of antibacterial charresults. 100μL. Literature review Figure 2. Traditional Romanian “ie”. FIBRES & TEXTILES in Eastern Europe 2021, Vol. 29, 5(149) Three Figure 1. Microscopic assessment of the quality of twisting fibres in the inner left fibre. Material and method To assess the presence of viable fungi on the surface of an examined area (Figure 2) before and after the application of essential oils, the following materials were required: a delimiting frame, sterile swabs, sterile distilled water, three essential oils with antifungal effects [35-40], namely Lavandula angustifolia – lavender, Mentha piperita – mint and Citrus limon – lemon [39],glass containers with a sprayer, Saboruaud and Csapek-Dox sterile culture media, a microbiological hood, an incubator, glass slides and slides, microbiological handles, KOH dissociative solution, an optical microscope, an API® 20C AUX kit for yeast identification, a densitometer, and a 10 volumetric automatic pipette -100μL. sprayer, Saboruaud and Working technique Three areas of dimensions of 5/5 cm were examined, as shown in Figures 2 and 3: inner left sleeve, medial-upper right face, and left shoulder. Figure 2. Traditional Romanian ``ie`` 81 2/3: inner left Three Three 2/3:inner innerleft left 2/3: the use of the API® 20 C AUX6 kit for identification of PI® 20 C AUX principl µL of fresh yeast suspension calibrated at a turbidity of 2 McFarland at temperatures of 29 ⁰C ± 2 ⁰C, and readings done after 48h and 72h by comparing each with the negative control. On the results sheet, the nding to the digit 0. The final result obtained at 72h is a 7 profile interpreted using ApiwebTM computer software. Figure 3. Areas examined (from left to right: left inner sleeve, right medial-upper front, left shoulder). The The sampling samplingstep stepwas wasperformed performedusing usingsterile sterileswabs. swabs.The Thesize sizeofofthe theparts parts the thedelimiting delimitingframe. frame.Samples Sampleswere weretaken takenbefore beforeand and Figure 4. From left to right: the area examined (inside the left sleeve), the Csapek-Dox environment and Sabouraud environment. The sampling step was performed using sterile swabs. The size of the parts examined was 5/5 cm in accordance with the delimiting frame. Samples were taken before and after 15 minutes from the application of essential oils according to Table 1. 5 drops (150 µL) of each essential oil were applied to the corresponding area of the fabric. To make their application as accurate as possible, glass containers with a sprayer were used, in which 50 drops of sterile distilled water, the equivalent of 7.5 mL, were previously added. After application, a break of 15 minutes was made before the second set of samples was taken. The samples were seeded on Sabouraud and Csapek-Dox culture media. Petri dishes with the seeded culture media were incubated for 7 days at 28 °C. After 3 days of incubation, fungal colonies became macroscopically visible. On the seventh day, three different types of colonies could be identified: two as a mould species and one as a yeast species . Identification of the fungi was performed by evaluation of macroscopic and microscopic characteristics in the case of moulds, respectively, and with the use of the API® 20 C AUX6 kit for identification of the yeast species. API® 20 C AUX principly evaluates the biochemical characteristics of yeasts by Table 1. Areas of traditional cloth ”ie” examined and essential oils applied. Area examined 82 Essential oil applied Area examined Essential oil applied Left inner sleeve Citrus limon (lemon) Upper medial face Lavandula angustifolia (lavander) Left shoulder Mentha piperita (mint) following 19 assimilation reactions. In the microwells of the API gallery, 100 µL of fresh yeast suspension calibrated at a turbidity of 2 McFarland was pipetted. Incubation was performed at temperatures of 29 °C ± 2 °C, and readings done after 48h and 72h by comparing each with the negative control. On the results sheet, the positive reactions are denoted by +, corresponding to the digit 1, and the negative ones by –, corresponding to the digit 0. The final result obtained at 72 h is a 7-digit numerical profile interpreted using ApiwebTM computer software. Results and discussions All fungal colonies were developed from samples before the application of essential oils. Figures 4, 5 and 6 represent the areas examined and fungal colonies developed on the culture media. Two different kinds of mould (Cladosporium sp. and Botrytis sp.) and a yeast species (Rhodotorula mucilaginosa) deFIBRES & TEXTILES in Eastern Europe 2021, Vol. 29, 5(149) Figure 5. From left to right: area examined (medial-upper right front), Csapek-Dox environment and Sabouraud environment. Two different kinds of mould ( [ ] Th Figure 6. From left to right: area examined (left shoulder), the Csapek-Dox environment and Sabouraud environment. veloped on the surface of the Petri dishes Table 2. Types of moulds identified. Two different kinds ofandmould ( [41]. The different moulds identified Images the effect of the essential oil on them are shown in Table 2. Since Cladosporium belongs to mould fungi, its spores are particularly troublesome for allergy sufferers, and its presence in the external and internal environment makes it difficult to prevent. However, it develops mainly in the external environment, often on dead plants, but also indoors [42-44]. In addition to moisture, its multiplication is favoured by dust and rare airing, typical of museum exhibitions, despite the use of humidity and air conditioning controls. Hence, due to its frequent presence in exhibits and display cases. Cladosporiums has a strong allergic effect [45]. The main symptoms of allergic reactions in the case of skin are most often atopic inflammation, with dry and cracked skin and itching. If it enters the respiratory tract, symptoms may include sneezing, itching of the nasal mucosa, larynx, and dry and FIBRES & TEXTILES in Eastern Europe 2021, Vol. 29, 5(149) [ ] The Name Effect of essential oils Cladosporium sp. Citrus limon (lemon) – inhibitory effect Botrytis sp. Citrus limon (lemon) – inhibitory effect Lavandula angustifolia (lavander) – inhibitory effect Mentha piperita (mint) – inhibitory effect 7 Table 3. Yeast species identified Images Name Rhodotorula mucilaginosa profil API: 6642272 – profil API: 6642272 Since Effect of essential oil Mentha piperita (mint) – inhibitory effect 83 [42 ] irritating cough. Allergic effects on the mucous membranes of the larynx cause a dry cough, and in the case of the mucous membranes of the eyes, the effect is often conjunctivitis on the rim of the eyelids, discharge from the eyes etc. More than ten years ago Rhodotorula fungi, due to the increasing number of immunocompromised patients, was classified by some researchers as pathogens [46]. This is owing to the high resistance to some medicines and the high adaptability of this species. In humans, it primarily causes bloodstream infections, but hypersensitivity pneumonitis is also possible [47]. Risk factors increasing its presence and activity include solid and haematologic malignancies in patients who receive corticosteroids and cytotoxic medicines, the presence of CVC, and the use of broad-spectrum antibiotics [48]. Unlike Cladosporium and Rhodotorula, fungi of the Botrytis species are not reported as pathogenic in humans. However, in the case of people with primary or acquired immunodeficiency and structural lung disease, they may experience systemic fungal infections [49]. This may apply to staff working in museum rooms, at exhibitions, and in places where fungi of the Botrytis family have good conditions to thrive. In such cases, systemic symptoms such as weight loss and/or a recurrent cough, shortness of breath etc. appear [50-52]. Botrytis in immunocompromised people, such as patients with the hepatitis C virus, AIDS, and organ transplant patients, can cause local infections, causing pathologies of the eye, brain, heart, peritoneum, or onychomycosis. It is highly resistant to conventional and most modern antifungal agents when its concentration is elevated or the body’s resistance is low [53, 54]. From the present study, we can state that the essential oils of lemon (Citrus limon), lavender (Lavandula angustifolia) and mint (Mentha piperita) have inhibitory effects on yeasts and mould spores. As well as this, the inhibitory action of lemon, lavender and mint essential oils against Botrytis sp that of lemon essential oil against Cladosporium sp. and that against the yeast species Rhododtorula mucilaginosa were demonstrated. Conclusions The research conducted allowed to identify the main species of developing fungi 84 in the material studied: Cladosporium, Rhodotorula and Botrytis. The first two are definitely pathogenic, while Botrytis, which is generally not hazardous to health, may show pathogenic features in selected, specific cases. As museum rooms and exhibitions are characterised by specific climatic conditions, also generated by CVC, they often become a place for fungal growth, especially on plant-based materials, such as cotton. Due to the time of creation of the exhibit (fabrics) as well as the storage and climatic conditions in the exhibition rooms, microorganisms (fungi, yeasts etc) often develop there. Fungi can easy contaminate clothing exhibits stored in museums and improper exhibition halls. They are one of the factors favouring the process of damage to the fabrics from which they are made. The fungi detected on the samples, depending on the species, may cause many allergic reactions and serious diseases. They are primarily exposed to museum workers who stay in rooms with fungal spores for several hours a day. Yeasts can also be dangerous to eldery people or those which have a weakened immunological system. Bearing in mind these risks, and all of the above, to try and prevent them, testing with essential oils should be implemented on a large scale, which, as shown by the results of the research, can be considered natural agents for biodesinfection to preserve museum exhibits and are non-toxic for humans. 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