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Early detection of pathogens is crucial for the effective surveillance of diseases. Many efforts have been made to explore methods which can detect these pathogens within a short period of time without requiring a tedious protocol. However, these developed methods have disadvantages such as they are relatively time-consuming or require specialized laboratory facilities. In this work, we have developed an integrated microfluidic system for rapid and automatic detection of viruses by direct analysis from fresh Phalaenopsis orchid leaves. The entire protocol, including ribonucleic acid (RNA) purification, reverse transcription loop-mediated-isothermal-amplification (RT-LAMP) and optical detection by measuring changes in turbidity was performed on a single chip. This is the first time that an integrated microfluidic system for the detection of viruses infecting the Phalaenopsis orchid has been demonstrated. The sensitivity of the developed system was also explored in this study to validate its performance., From the Clinical Editor: In this study, the authors report the development of an integrated microfluidic system for rapid and automatic detection of viruses by direct analysis of fresh Phalaenopsis orchid leaves, performing the 3-step protocol using a single chip. Similar methods may find clinical application for fast and accurate detection of viral infections., (Copyright © 2013 Elsevier Inc. All rights reserved.)

• PCR-based techniques are the most widely used diagnostic tools for virus indexing. • Out of 681 total genera, only 49 genera were reported for virus infection. • Cypripedioideae is the most affected among the sub-family in Orchidaceae. • Production of Virus-free orchids using in vitro techniques is a desirable approach. The maximum number of genera in the family Orchidaceae are well represented by their floricultural importance and are exported globally for trade exchanges. However, economic loss due to viral diseases has a profound impact on the orchid trade industry worldwide. Further, enduring phytosanitary certificates and undergoing stringent quarantine measures for virus-free orchids pose a major setback during the exchange of breeding materials between regions or countries. Fortunately, the spread of orchid viruses can be effectively controlled if proper diagnosis of viral diseases is made at an early stage and strict application of appropriate eradication methods are deliberated in a holistic approach. This bibliographic review highlights the substantial advancements in the diagnostic and eradication techniques of orchid viruses during the past few decades. The review shows that PCR-based techniques, which are represented as the "gold standard" method, are the most widely used diagnostic tools because of their accuracy, simplicity, and sensitivity. Considering the large diversity of orchids, only 7.2% of the genera belonging to the sub-families Epidendroideae, Orchidoideae, Cypripedioideae, and Vanilloideae have been detected for virus infection. The tribes Podochileae, Thaieae, Nervilieae, Gastrodieae, Xerorchideae, Triphoreae, Wullschlaegelieae, Tropidieae, Sobralieae, and Neottieae of the family Orchidaceae are under-represented. In vitro techniques, including meristem culture, thermotherapy, chemotherapy, and their combinations, represent the most desirable and effective approaches for orchid viral elimination. The use of cryotherapy, a simple technique based on cryopreservation, should have a high priority for viral elimination from infected orchids. This review could pave the way forward for comprehensive research on the production of virus-free orchids. [ABSTRACT FROM AUTHOR]

Societal Impact Statement Orchid viruses are capable of causing flower deformities and death, which can severely impact the horticultural industry and wild orchid conservation. Here we show how two of these quickly evolving viruses display few genetic differences since their first emergence, across countries and host plants. This is concerning as, despite biosecurity regulations to control the movement of orchids and their related pathogens, these patterns are suggestive of rapid and regular international movement of horticultural material. Poor biosecurity practices could threaten the orchid horticultural industry and result in the accidental translocation or reintroduction of infected plant material intended to recover wild populations.

Societal Impact Statement: Orchid viruses are capable of causing flower deformities and death, which can severely impact the horticultural industry and wild orchid conservation. Here we show how two of these quickly evolving viruses display few genetic differences since their first emergence, across countries and host plants. This is concerning as, despite biosecurity regulations to control the movement of orchids and their related pathogens, these patterns are suggestive of rapid and regular international movement of horticultural material. Poor biosecurity practices could threaten the orchid horticultural industry and result in the accidental translocation or reintroduction of infected plant material intended to recover wild populations. [ABSTRACT FROM AUTHOR]