The effects of ultraviolet B beams on programmed cell death activities in Staphylococcus epidermidis

Journal of Microbiology and Infectious Diseases / JMID 2015; 5 (1): 21-24 doi: 10.5799/ahinjs.02.2015.01.0169 RESE ARCH ARTICLE The effects of ultraviolet B beams on programmed cell death activities in Staphylococcus epidermidis Payam Behzadi1, Elham Behzadi1, Reza Ranjbar2 1 Department of Microbiology, College of Basic Sciences, Shahr-e-Qods Branch, Islamic Azad University, Tehran, Iran 2 Molecular Biology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran ABSTRACT Objectives: Bacterial skin diseases caused by drug-resistant Staphylococcus epidermidis are known as a big problem in the field of treating management of infectious diseases. Progression of resistant strains has led to use phototherapy in parallel with pharmacotherapy. In this short survey, we tried to obtain a logic Ultraviolet Radiation protocol to induce the process of programmed cell death in irradiated Staphylococcus epidermidis. Methods: The samples of Staphylococcus epidermidis were classified in 4 categories. Each plate which is known as a category included well-grown colonies of bacteria. One plate was taken as a control sample and the left three plates were irradiated by UVB (302 nm) in 10 minutes from the distance of 8 cm. The irradiated plates were kept in a dark cell and for 1, 24 and 72 hours respectively. Then, total genomic DNA molecules pertaining to all of the colonies comprising control and irradiated samples were harvested by DNP kit and the extracted DNA molecules were running upon the 1% agarose gel together with ethidium bromide. Results: Control and irradiated samples were studied for probable changes in their macroscopic, microscopic characteristics and then the DNA pattern relating to each group was detected for any variation including smear or DNA laddering bands. No changes were observed in different bacterial properties. No apoptosis were observed in irradiated samples. Conclusion: UVB is a strong apoptosis stimulus in organisms. However, authors of the present study strongly reject the apoptotic effect of UVB irradiation in the format of the present protocol. J Microbiol Infect Dis 2015;5(1): 21-24 Key words: In vitro, programmed cell death, Staphylococcus epidermidis, UVB UVB ışınlarının Stafilokok epidermidisdeki programlanmış hücre ölüm faaliyetleri üzerine etkileri ÖZET Amaç: İlaca dirençli Staphylococcus epidermidis neden olduğu bakteriyel deri hastalıkları, enfeksiyon hastalıklarının tedavi alanında büyük bir problem olarak bilinmektedir. Dirençli suşların artması farmakoterapiyle beraber fototerapinin de kullanılmasına yol açmaktadır. Bu kısa araştırmada, ışına maruz kalmış Staphylococcus epidermidisin programlanmış hücre ölüm sürecini başlatacak anlamlı bir Ultraviole Radyasyon protokolünü oluşturmaya için uğraştık. Yöntemler: Staphylococcus epidermidis örnekleri 4 kategoride sınıflandırılmıştır. Bakteri kolonilerinin çoğalma durumuna göre her plak bir kategoriye dahil edildi. Bir plak kontrol olarak ve diğer 3 plağa ise 8cm uzaktan 10 dakika süreyle ultraviolet B (UVB) (302 nm) ışını uygulandı. Işına maruz kalan plaklar sırayla 1saat, 24 saat ve 72 saat süreyle karanlık bir hücre tutuldu. Daha sonra, kontrol ve ışınlanmış örnekler içeren tüm kolonilerin toplam genomik DNA molekül parçaları DNP kiti ile çoğaltıldı ve ekstre edilmiş DNA molekülleri, etidyum bromür ile birlikte % 1 agaroz jel üzerinde yüklenip yürütüldü. Bulgular: Kontrol ve ışınlanmış örneklerde makroskopik, mikroskopik özelliklerindeki olası değişiklikler için çalışma yapıldı ve sonra her gruba ilişkin DNA merdiven bantları veya yaymalarını da içeren DNA örnekleri üzerinde herhangi bir değişiklik olup olmadığı belirlendi. Farklı bakteriyel özelliklerde herhangi bir değişiklik gözlenmemiştir. Işınlanmış örneklerde hiçbir apoptoz gözlenmemiştir. Sonuç: UVB organizmalarda güçlü bir apoptoz uyarıcıdır Ancak, bu çalışmanın yazarları bu protokol formatında UVB ışınlamasının apoptotik etkisinin olmadığını şiddetle reddetmektedir. Anahtar kelimeler: in vitro, programlanmış hücre ölümü, Staphylococcus epidermidis, UVB Correspondence: Payam Behzadi, Department of Microbiology, College of Basic Sciences, Shahr-e-Qods Branch, Islamic Azadi University, Tehran, Iran Email: Received: 01 October 2014, Accepted: 05 December 2014 Copyright © Journal of Microbiology and Infectious Diseases 2015, All rights reserved 22 Behzadi P, et al. Apoptosis in Staphylococcus epidermidis INTRODUCTION The human skin encompasses a vast diversity of microbial normal flora, including Coagulase-Negative Staphylococci (CNS). CNS comprises the gram positive bacterium of Staphylococcus epidermidis as the most common colonizer of the human body which involves armpits, inner surfaces of nares. The predominant strains of S.epidermidis support health of the skin against pathogenic bacteria like Staphylococcus aureus. However, some opportunistic pathogenic strains of S.epidermidis are recognized as device-related bacterial infectious agents. Besides, a wave of Methicillin-Resistant S.epidermidis (MRSE) has been distinguished among nosocomial infectious isolates.1-6 Today, the multi-drug-resistant strains are significantly increasing around the world. Thus, the use of UV-phototherapy may be a good choice for replacing or administrating in parallel with chemotherapy.2,7 minimum volumes. According to our protocol, the three irradiated plates were maintained in a dark cell for respectively 1, 24 and 72 hours while the control plate did not undergo UVB-exposure. In the next step, the total genomic DNA of the four samples, including control and irradiated colonies were extracted by DNP kit through following company’s protocol (50T, CinnaGen Inc.).4,9,10 In the following step, the harvested DNA molecules relating to each studied group were running on agarose gel electrophoresis comprising ethidium bromide 1%. Furthermore, a DNA weight marker III produced by CinnaGen Company was applied to compare and analyze the created bands. The density of RNA molecules was ignorable and not significant. In the final stage, the created DNA molecules bands belonging to control and UVB-exposed S.epidermidis bacterial cells were assessed for probable DNA laddering bands as a symbol pattern of programmed cell death (Figure 1).4,8-16 The use of UV beams may help to induce the programmed cell death (apoptosis) in irradiated bacterial cells. For this purpose, the agarose gel electrophoresis seems a suitable tool for observing apoptosis feature in the format of DNA laddering bands. The appearance of apoptosis may lead to use the protocol as an in vivo treating management for different microbial organism.4,8-14 The aim of this survey is to detect programmed cell death (apoptosis) activities in the irradiated bacterial cells of S.epidermidis via a determined UVBirradiation protocol designed by the authors. METHODS The bacterial samples of S.epidermidis were provided from the microbial resources of the microbiology laboratory, Islamic Azad U (...truncated)