Anti-viral activity of culinary and medicinal mushroom extracts against dengue virus serotype 2: an in-vitro study
Ellan et al. BMC Complementary and Alternative Medicine
https://doi.org/10.1186/s12906-019-2629-y
(2019) 19:260
RESEARCH ARTICLE
Open Access
Anti-viral activity of culinary and medicinal
mushroom extracts against dengue virus
serotype 2: an in-vitro study
Kavithambigai Ellan1,2* , Ravindran Thayan1, Jegadeesh Raman5, Kazuya I. P. J. Hidari4, Norizah Ismail3 and
Vikineswary Sabaratnam2*
Abstract
Background: Dengue is a mosquito-borne viral infection that has become a major public health concern
worldwide. Presently, there is no specific vaccine or treatment available for dengue viral infection.
Methods: Lignosus rhinocerotis, Pleurotus giganteus, Hericium erinaceus, Schizophyllum commune and Ganoderma
lucidium were selected for evaluation of their in-vitro anti-dengue virus serotype 2 (DENV-2) activities. Hot aqueous
extracts (HAEs), ethanol extracts (EEs), hexane soluble extracts (HSEs), ethyl acetate soluble extracts (ESEs) and
aqueous soluble extracts (ASEs) were prepared from the selected mushrooms. The cytotoxic effects of the extracts
were evaluated by the MTT assay. The anti-DENV-2 activities of the extracts were evaluated in three different assays:
simultaneous, attachment and penetration assays were perfomed using plaque reduction assays and RT-qPCR
assays. The effect of the addition time on viral replication was assessed by the time of addition assay, and
a virucidal assay was carried out to evaluate the direct effect of each mushroom extract on DENV-2. The chemical
composition of glucans, and the protein and phenolic acid contents in the extracts were estimated.
Results: We found that the HAEs and ASEs of L. rhinocerotis, P. giganteus, H. erinaceus and S. commune were
the least toxic to Vero cells and showed very prominent anti-DENV2 activity. The 50% inhibitory concentration (IC50)
values of the ASEs ranged between 399.2–637.9 μg/ml, while for the HAEs the range was 312.9–680.6 μg/ml during
simultaneous treatment. Significant anti-dengue activity was also detected in the penetration assay of ASEs (IC50:
226.3–315.4 μg/ml) and HAEs (IC50: 943.1–2080.2 μg/ml). Similarly, we observed a marked reduction in the
expression levels of the ENV and NS5 genes in the simultaneous and penetration assays of the ASEs and HAEs.
Time-of-addition experiments showed that the highest percent of anti-DENV2 activity was observed when
the mushroom extracts were added immediately after virus adsorption. None of the extracts exhibited virucidal
effect. Chemical composition analysis showed that the major components in the mushroom HAEs and ASEs were
glucan (beta D-glucan) and proteins, however, there was no significant correlation between the anti-dengue
activity and the concentration of glucans and proteins.
Conclusion: These findings demonstrated the potential of mushroom extracts as anti-dengue therapeutic agents
with less toxic effects.
Keywords: Dengue virus, Mushrooms, Anti-dengue activity, Mushroom extracts, Cytotoxicity, Plaque reduction assay
* Correspondence: ;
1
Virology Unit, Infectious Disease Research Centre, Institute for Medical
Research, Ministry of Health, Kuala Lumpur, Malaysia
2
Mushroom Research Centre, Institute of Biological Sciences, Faculty of
Science, University of Malaya, Kuala Lumpur, Malaysia
Full list of author information is available at the end of the article
© The Author(s). 2019 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and
reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to
the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver
(http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
Ellan et al. BMC Complementary and Alternative Medicine
(2019) 19:260
Background
Dengue is a critical mosquito-borne viral disease
that continues to cause substantial public health burdens
in Asia and the Pacific. The reported number of dengue
cases has increased over the past decade. In 2017, there
were 407,199 dengue cases and 874 dengue deaths reported from countries in the Western Pacific Region [1].
In Malaysia, the number of dengue cases was 82,840 and
171 deaths, the third highest among the countries in the
Western Pacific region. Dengue virus (DENV) exists in
nature as four antigenically distinct but closely related
virus serotypes known as DENV-1, 2, 3 and 4, which can
be transmitted to humans by the mosquito species Aedes
aegypti and Aedes albopictus. The clinical syndrome of
DENV infection had been classified into dengue with or
without warning signs and severe dengue. Attempts regarding vaccine development for dengue have been a
continuous challenge for many years due to the inability
of the vaccines to concurrently protect against all four
antigenically distinct dengue serotypes [2, 3]. Effective
antiviral therapies are currently unavailable for DENV.
Dengue patients are usually managed through supportive
therapies until they recover without any specific treatment measures. The search for new antiviral agents is
vital for the prompt treatment of dengue infection to
avoid further development of severe dengue, to control
the spread of the outbreak and to enhance possible vaccination programmes in the future.
Culinary and medicinal mushrooms have great prospects in the drug and nutraceutical industries. They
possess a wide range of pharmacological properties, including antimicrobial, antiviral, antitumour, anti-inflammatory, immunomodulatory, hypoglycaemic and
hepatoprotective properties, and thus can be considered
a functional food. Mushroom antiviral activities have
been reported towards human immunodeficiency virus
(HIV) using lectin isolated from Hericium erinaceus [4]
and laccase isolated from Clitocybe maxima [5]. An
acidic protein bound polysaccharide isolated from Ganoderma lucidium [6] and a protein from Grifola frondosa
[7] inhibited the replication of the herpes simplex virus
(HSV). Influenza viruses have been inhibited by ganomycins A and B from Ganoderma pfeifferi [8] and polysaccharides isolated from Agaricus brasiliensis showed
antiviral activity against polioviruses [9].
In this study, five culinary and medicinal mushrooms
that are commercially grown in Malaysia, Lignosus rhinocerotis (Cooke) Ryvarden, Pleurotus giganteus (Berk) Karunarathna & K.D.Hyde, Hericium erinaceus (Bull) Persoon,
Schizophyllum commune (Fr.) and Ganoderma lucidium
(Curtis) P. Karst were selected for in-vitro screening for
their anti-dengue serotype 2 (DENV-2) activity. Two types
of extraction protocols were used to isolate chemical components from selected mushrooms. Initially, hot aqueous
Page 2 of 12
extracts and ethanol extracts were prepared from the dry
powdered mushroom fruiting bo (...truncated)
