Abstract
Background
In spite of the significant progress in modern medicine, viruses continue to be a formidable challenge to human health. The use of plants for the remediation of viral-borne diseases stretches back to the very dawn of mankind. Among bulbous plants, the Amaryllidaceae is one of the most popular families exploited in the traditional remediation of infectious diseases.
Methods
This account details the findings from a literature search carried out on the antiviral properties of the Amaryllidaceae. The keyword engaged in the search was “antiviral” in combination with the words “Amaryllidaceae,” “Amaryllidaceae specie,” and “Amaryllidaceae alkaloid.”
Results
Thirty-six taxa from 13 countries, notably in Africa and parts of Asia, have been cited as traditional remedies for viral diseases. Alcoholic bulb extracts of 18 species were evaluated against 23 different pathogens from 13 viral families. A wide range of activities was observed, with the whole-plant methanol extract of Zephyranthes candida seen to be the most striking (IC50 0.0019 µg/mL against poliovirus). The active principles in the main were isoquinoline alkaloids, of which lycorine impressed against the Avian influenza virus (strain H5N1). The mechanisms underlying the antiviral effects were seen to be related to the inhibition of DNA, RNA, and protein synthesis as well as inhibitory effects toward reverse transcriptase and protease enzymes.
Conclusion
Amaryllidaceae provides a richly diverse platform for antiviral drug research. Such endeavors have been fortified by the significant amounts of information emerging from indigenous knowledge systems. Ongoing studies will continue to target the active entities, particularly from taxa with verifiable ethnomedicinal backgrounds.
Introduction
Diseases caused by viruses have been prevalent throughout the course of history. 1 They initiate a broad spectrum of diseases that range from common colds and flu to serious ailments such as Ebola and AIDS. 1 Some have had a devastating impact on society, such as “Spanish flu,” which was responsible for upwards of 50 million deaths during 1918 to 1920. 1 In recent times, COVID-19 has brought to fore the impact of a full-scale pandemic, which has touched almost every facet of contemporary society. 2 Over 600 million cases and in excess of 6 million deaths were tracked globally by the third year of its spread. 3 This in spite of phenomenal efforts in search of vaccines and drugs against SARS-CoV-2, the virus behind COVID-19. 2
The Baltimore classification is recognized as the authoritative system in the taxonomic consideration of viruses. 1 The 4 key factors used to identify viruses via this system are nucleic acid makeup, strandedness, sense, and method of replication. 1 The 7 viral groups recognized by the system are ssDNA viruses, dsDNA viruses, dsRNA viruses, (+)ssRNA viruses, (−)ssRNA viruses, ssRNA-RT viruses, and dsDNA-RT viruses. 1 The host engages the immune system through either innate or adaptive responses as a primary defense against viruses. 4 The inhibition of viral replication by inhibiting viral RNA replication may be viewed as an innate response, while the adaptive response involves the key antibodies IgM and IgG. 4 Cell-mediated immunity on the other hand invokes the use of interferon, macrophages, and “killer” T-cells, among others, for the detection and destruction of pathogens as well as for the annihilation of infected cells. 4 Complementing the natural host response are vaccine schedules and antiviral drug targets that have had a poignant influence on viral epidemiology. 4
There has been notable interest in the exploitation of plants for the management of viral-borne diseases through the course of history.5,6 This has been bolstered by their vast chemical resources, which reflect attractive structural diversities and interesting biological activities.
7
The family Amaryllidaceae J.St.-Hil. is reputed for its geophytic flowering plants, which produce a wide array of biologically active isoquinoline alkaloid entities.
8
Of note here are its representatives galanthamine (
Antiviral principles identified in the plant family Amaryllidaceae.
Traditional Plant Usage
Although contemporary medicine has had a dominant influence on the well-being of society today, traditional medicine still plays a significant function in the management of diseases particularly on the African continent and parts of Asia. 12 The number of plants that have been used for viral infections runs into several thousand.5,6 A global survey of the Amaryllidaceae indicated that as many as 36 of its members have been cited for such functions in TM (Table 1). Of the 18 identified in Africa, 14 were noted for their origins in South Africa. This is not surprising given that over a third of all Amaryllidaceae taxa are to be found in South Africa.13,14 They are among the most popular bulbous plants exploited for medicinal use by the indigenous Xhosa, Sotho, Venda, and Zulu peoples.13,14 The genera represented in South Africa for viral infections were Boophone, Brunsvigia, Clivia, Crinum, Cyrtanthus, Gethyllis, Haemanthus, Nerine, and Scadoxus. Globally, Crinum emerged as the most popular genus, with 12 of its species reported for such purposes in 8 different countries. India came second to South Africa in its use of Amaryllidaceae plants (8) for antiviral purposes, the majority of which emanated from the genus Crinum. Also noteworthy was the exploitation of Haemanthus and Zephyranthes (4 species each) in 4 different countries including, South Africa, India, Nigeria, and Argentina. Eight areas were identified wherein Amaryllidaceae plants have found use against viral-borne diseases, which ranged from colds, fever, and flu to STDs, smallpox, and measles as well as polio and HIV/AIDS. The most common usage was for the alleviation of colds and fever. Since the Amaryllidaceae are bulbous geophytic plants it was not surprising to see that the majority of the cases involved the use of bulbous materials, water, or alcohol extracts of which were taken orally. However, there were cases involving the use of flower, leaf, whole plant, and root extracts as indicated in Table 1.
Amaryllidaceae Plants Documented in the Traditional Remediation of Viral-Borne Diseases.
Plant parts used: 1bulb; 2leaf; 3whole plant; 4root; and 5flower.
Activities Against Arenaviridae
Arenaviruses, as members in this family are referred to, are generally spread by rodents.
1
These RNA viruses have a segmented genome containing 2 single-strand ambisense RNA molecules.
1
As with all negative-sense RNA viruses, genomic RNA alone is not infectious so that the virus must replicate to allow infection of the host.
1
The first member to be identified in this family was lymphocytic choriomeningitis (LCM) virus, which is responsible for LCM in rats and aseptic meningitis in humans.
1
Over 70 pathogens have since been identified in the family, causing diseases mostly in rodents but also in humans.
1
Narcissus tazetta was the first Amaryllidaceae plant to be screened for antiviral effects, appearing amongst a collection of around 200 Chinese medicinal plants that were screened for effects on the rodent viral disease LCM caused by LCM virus.
35
A bulb H2O extract exhibited a maximum nontoxic dose (MNTD) of 0.6 mg relative to 5-methyltryptophan (0.4 mg) during a screen for its effect on the proliferation of LCM in human KB epithelial cells.
35
Furthermore, no cytopathic effects were observed for the extract at concentrations up to and including its MNTD.
35
In vivo, there was only a 37% mortality rate in LCM-infected mice exposed subcutaneously twice-daily for 5 days to Narcissus tazetta at its MNTD, compared to untreated controls (86%).
35
The extract (30%) also improved the mortality rate relative to methotrexate (70%) in infected mice subjected to the respective MNTD regimens.
43
These effects were shown to be dose-dependent over the injected dosage range 0.5, 1.0, 2.0, and 4.0 mg.
43
A total alkaloid fraction (at 1 mg) prepared from the original H2O extract via partitioning with acid and base followed by the elimination of neutral material was seen to further improve mortality to 20%.
44
However, its 2 isolated principles narciclasine (
Activities Against Retroviridae
A retrovirus is characterized by its ability to insert a copy of its RNA genome into the DNA of the host, where it uses reverse transcriptase to produce DNA from RNA.
1
An integrase enzyme catalyzes the incorporation of the new DNA into the host genome, a point at which the retroviral DNA is referred to as a provirus.
1
The viral DNA is then recognized by the host as a component of its own genome, allowing it to transcribe and translate viral genes as well as its own genes and produce the proteins needed to make new copies of the virus.
1
Further studies on Narcissus tazetta saw its bulb H2O extract (at 100 and 200 mg/kg) increase the prolongation of median survival time (MST) to 26% relative to controls in BALB/c mice that were infected with MCDV-12 cells.
45
The ascites cell line MCDV-12, a syngeneic tumor, is known to develop in BALB/c mice following exposure to Rauscher leukemia virus (RLV).
45
Studies aimed at identifying the anti-RLV component of Narcissus tazetta led to the identification of pseudolycorine (
A screen for anti-HIV effects of Korean medicinal plants indicated only 2 plants (out of 49) to be active against HIV-1, which were Crinum asiaticum var. japonicum Baker (Amaryllidaceae) and Veratrum patulum O.Loes. (synonym Veratrum oxysepalum Turcz.) (Liliaceae).
47
The MeOH root extract of Crinum asiaticum var. japonicum was active against HIV-1 multiplication in human MT-4 cells (ED50 12.5 µg/mL) (Table 2), whose effect was seen to be selective by a factor of 16.
47
In terms of how it manifested this action, at half its cytotoxic dosage (100 µg/mL) the root extract inhibited HIV-1 RT by as much as 70.8%.
47
The chief principle of Crinum asiaticum lycorine (
Activities of Amaryllidaceae Plant Extracts Against Various Viral Pathogens.
At MNTD values.
By contrast, Hippeastrum glaucescens via its CH2Cl2 bulb, flower, leaf, and root extracts was seen to be inactive against HIV-1 that was propagated in Vero cells.
55
Similarly, the EtOH extract of bulbs from Crinum macowanii Baker (at its CC90 value) had little effect on the viabilities of HIV-1-infected lymphoblastic CEM cells, neither did it affect HIV-1 p24 antigen levels.
51
In spite of this, Crinum macowanii has been reported to be used against STDs, presumably including HIV-AIDS, in South Africa and Zimbabwe.13,27 The African medicinal plant Pancratium trianthum Herb. was screened for effects against HIV-1.
61
Its dried bulb MeOH extract dose-dependently (0.09-3.125 µg/mL) inhibited viral proliferation of THP-1 monocytes, whereby weakly cytotoxic concentrations of 0.09 and 0.19 µg/mL significantly reduced infectivity by 28% and 52%, respectively.
61
At its EC50 (0.17 µg/mL) dose, the viability of host cells was 65% that of the control standard NITD008.
61
Trisphaeridine (
Activities Against Herpesviridae
Comprising around 130 members, Herpesviridae represents a large family of DNA viruses.
1
The most common of these are herpes simplex virus, herpes zoster virus (shingles), and varicella zoster virus (chickenpox).
1
These viruses are known to cause illnesses in amphibians, birds, fish, mammals, molluscs, and reptiles.
1
The EtOH bulb extract of Narcissus tazetta was examined for effects on a further 2 animal viral pathogens, infectious bovine rhinotracheitis (IBR) and equine rhinopneumonitis (ERP) both from the family Herpesviridae.
59
No viral-induced plaque formation was observed in IBR-infected BECs subjected to treatments with Narcissus tazetta at 1.0 and 2.0 mg/mL, respectively.
59
Similar effects were observed for the extract in ERP-infected BECs but at markedly higher dosages of 250 and 500 mg/mL.
59
These activities interestingly ensued without palpable deleterious effects on the BECs.
59
A screen of 100 higher plants (from around 40 families) against 6 human viral pathogens indicated the Amaryllidaceae representatives to exhibit the best overall activities.
49
EtOH leaf extracts of the 5 plants at MNTDs, including Clivia × cyrtanthifolia (Lindl. ex K.Koch & Fintelm) T.Moore, Clivia miniata (Lindl.) Bosse, Hymenocallis littoralis (Jacq.) Salisb., Narcissus pseudonarcissus L., and Narcissus tazetta, exhibited viral titer reduction factors (in multiples of 10) that ranged from 1 to 10 000.
49
These were determined to be 10 000, 10 000, 100, 100, and 100 for the 5 plants against herpesvirus in Vero cells, respectively, for which the viral titer was estimated to be 105 TCID50/mL.
49
Out of these plants, Clivia miniata is known to be used for the remediation of HIV in South Africa, and there is a likelihood that its usage may also involve other STDs (such as herpes).
15
Of the 4 alkaloids isolated from Clivia miniata, lycorine (
Activities Against Picornaviridae
The Picornaviridae is comprised of nonenveloped (ie, lacking an outer lipid bilayer) RNA viruses that target vertebrates such as mammals, birds, and fish.
1
Poliomyelitis, meningitis, and hepatitis are some of the diseases caused by viruses in this family.
1
The screen of Clivia miniata, Clivia × cyrtanthifolia, Narcissus pseudonarcissus, Narcissus tazetta, and Hymenocallis littoralis for antiviral effects also involved 2 representatives of the Picornaviridae; polio and coxsackie virus (cultured in HeLa cells).
49
The best activities were those determined for both Clivia species against coxsackie and Clivia miniata against poliovirus (viral titer reduction factors of 10 000, respectively) (Table 2).
49
A follow-up study resulted in the isolation of the 4 alkaloidal substances lycorine (
Given its reputed use in Nigerian traditional medicine, a MeOH extract of Zephyranthes candida (Lindl.) Herb. prepared from whole-plant material was examined for effects on poliovirus.
40
It exhibited potent activity (IC50 0.0019 µg/mL) against viral infection of rhabdomyosarcoma, with its effect on host cells shown to occur at a far higher concentration (CC50 0.293 µg/mL).
40
Partitioning of the crude between CHCl3, EtOAc, and Hex and evaluation of the resultant extracts indicated that only the former (IC50 0.0012 µg/mL) was capable of improving the activity of the primary extract.
40
Activities for the EtOAc and Hex extracts were far lower as indicated by IC50 values of 3.78 and 1.62 µg/mL, respectively.
40
Nonetheless, respective selectivity indices of 7.3 and 18.3 indicated an overwhelming antiviral effect for these 2 extracts.
40
These effects could be explained in regards to the Zephyranthes candida component lycorine (
Activities Against Reoviridae
Viruses in this family contain double-stranded RNA molecules and are known to infect a wide range of hosts including, vertebrates, invertebrates, plants, fungi, and even protists. 1 Without possession of a lipid envelope, they utilize multilayered capsid structures to package their segmented genome. 1 Of the Reoviridae, only Simian rotavirus (strain SA-11) was engaged in studies against the Amaryllidaceae. 54 The EtOH bulb extract of Haemanthus albiflos manifested its action against this virus by inhibiting RNA synthesis by 42% in MA-104 monkey kidney host cells. 54 However, the event ensued at a much higher concentration of the extract (25 µL/mL) than that observed above against poliovirus. 53 Viral RNA synthesis was also inhibited (46%) by the extract in the presence of 4 µg/mL actinomycin D. 54 This response occurred at a much lower inhibitory level (46%), higher dose (25 µL/mL), and over a longer duration (20 h) relative to the outcomes observed for poliovirus (90%, 0.2 µL/mL, 3 h).53,54
Activities Against Togaviridae
This family of small, enveloped viruses possesses single-strand, positive-sense RNA genomes that are typically 10 to 12 kb in size.
1
The 2 genera constituting the family are Alphavirus and Rubivirus, of which Rubella virus from the latter and Chikungunya virus from the former are of considerable concern to humans.
1
Alphavirus also comprises several pathogens that are of veterinary relevance, such as Venezuelan equine encephalitis virus, western equine encephalitis virus, and eastern equine encephalitis virus.
1
The investigation of van den Berghe et al in 1978 outlined above also involved an examination of semliki forest virus (grown in HeLa cells).
49
In this instance, viral titer reduction factors for Hymenocallis littoralis, Clivia × cyrtanthifolia, Clivia miniata, Narcissus tazetta, and Narcissus pseudonarcissus ranged from 100 to 10 000, with both Clivia species shown to be the most effective.
49
The viral titer measured for semliki forest virus amounted to 107 TCID50/mL.
49
The constituents from Clivia miniata lycorine (
Activities Against Paramyxoviridae
Paramyxoviridae is a family of negative-strand RNA viruses which infect vertebrates and include common human diseases such as measles and mumps.
1
Van den Berghe et al in their broad screen of higher plants against human viruses included measles virus as a representative of the Paramyxoviridae.
49
The Amaryllidaceae members Clivia miniata, Clivia × cyrtanthifolia, Narcissus pseudonarcissus, Hymenocallis littoralis, and Narcissus tazetta were all active against the pathogen (cultivated in Vero cells), with viral titer reduction factors of 100 to 10 000 (Table 2).
49
The viral titer for measles virus was determined to be 106 PFU/mL, with Clivia miniata and Narcissus pseudonarcissus shown to be the pick of the 5 plants.
49
Interestingly, Clivia miniata has been cited in Zulu medicinal tradition specifically for the remediation of measles.
16
Phytochemical studies of the plant followed by screening against measles virus indicated that of the isolates only lycorine (
Activities Against Adenoviridae
Adenoviruses are medium-size, nonenveloped viruses of about 90 to 100 nm in diameter, with double-strand DNA genomes that are housed within an icosahedral nucleocapsid. 1 Originally derived from human adenoids in 1953, there are now over 50 distinct adenoviral serotypes. 1 Targeting mainly vertebrate hosts, they can cause conditions such as respiratory disease (adenovirus B and C), conjunctivitis (adenovirus B and D), gastroenteritis (adenovirus F and G), and obesity (adenovirus A, C, and D). 1 Adenovirus (no strain indicated) was the only member of the Adenoviridae engaged in the study on the antiviral effects of higher plants. 49 Grown in HeLa cells, it proved to be weakly susceptible from exposure to EtOH leaf extracts of Clivia miniata, Clivia × cyrtanthifolia, Narcissus pseudonarcissus, Hymenocallis littoralis, and Narcissus tazetta (viral titer reduction factors of 10-100). 49 A viral titer of 103 TCID50/mL was measured for adenovirus. 49
Activities Against Rhabdoviridae
Rhabdoviridae constitutes a family of negative-strand RNA viruses with a wide variety of hosts including, vertebrates and invertebrates, plants, fungi, and protozoa.
1
Diseases associated with these viruses include rabies encephalitis (after infection with rabies virus) and flu-like symptoms that afflict humans (after exposure to some vesiculoviruses).
1
The family has 40 genera, most assigned to 3 subfamilies; alpharhabdovirinae, betarhabdovirinae, and gammarhabdovirinae.
1
Indiana vesiculovirus represents the only member of the Rhabdoviridae that has been screened against the Amaryllidaceae.
58
In this regard, EtOH bulb extracts of Narcissus pseudonarcissus and Narcissus poeticus L. were screened against Indiana vesiculovirus.
58
Narcissus poeticus (MIC 5 µg/mL) was twice as active as Narcissus pseudonarcissus, with their cytotoxic concentrations against host Vero cells ascertained as 63 and 25 µg/mL, respectively.
58
The Narcissus pseudonarcissus principle lycorine (
Activities Against Flaviviridae
Flaviviridae represent positive-strand, enveloped RNA viruses that mainly afflict birds and mammals and are spread via arthropod vectors such as mosquitoes and ticks.
1
There are 89 species in the family which are divided into 4 genera: Hepacivirus, Flavivirus, Pestivirus, and Pegivirus.
1
Of note, the flaviviruses West Nile, Dengue, Zika and Yellow fever viruses can have life-threatening outcomes in humans.
1
The Hepacivirus representative hepatitis C virus can likewise be fatal against which, unlike hepatitis A and B, there is no vaccine to prevent infection.
1
A bulbous MeOH extract of Crinum macowanii was active against Japanese encephalitis virus (IC50 5.12 µg/mL) and Yellow fever virus (IC50 4.33 µg/mL).
27
A 32 µg/mL dose was sufficient to reduce by 100% and 70%, respectively, the cytopathic effect of each virus, without any deleterious effects being manifested toward the host Vero epithelial cells.
27
The basis for the action against Yellow fever virus may reside with the Crinum macowanii constituent lycorine (
The EtOAc bulb extract of Crinum jagus following acid and base partitioning steps was screened against Dengue virus, where it dose-dependently relative to NITD008 inhibited viral replication at concentrations between 0.078 and 2.5 µg/mL.
50
The EC50 was determined to be 0.25 µg/mL, with the corresponding cytotoxic effects on Huh-7 host cells shown to be minimal at all tested concentrations.
50
The adjoining facet of the study was the demonstration that lycorine (
Activities Against Phenuiviridae
Phenuiviridae is a family of negative-strand RNA viruses with ruminants, camels, humans, and mosquitoes serving as their natural hosts.
1
Its members have enveloped viruses that have helical capsid morphology and whose glycoproteins display icosahedral symmetries.
1
The study of Crinum macowanii involved 2 Phenuiviridae members: Sandfly fever-Sicilian virus and Punta Toro virus.
27
In contrast to the effectiveness of its MeOH bulb extract against the Flaviviridae above, no inhibitory effects were observed on the Vero cell-cultured Phenuiviridae viruses at doses lying below the cytotoxic concentration (30 µg/mL).
27
Nonetheless, lycorine (
Activities Against Coronaviridae
Coronaviridae is a family of positive-strand, enveloped RNA viruses that utilize mammals, birds, and amphibians as their hosts.
1
Included in the group are the subfamilies Letovirinae and Orthocoronavirinae, the latter of whose members are referred to as coronaviruses.
1
They are associated with respiratory conditions that range from the common cold to serious illnesses such as MERS (caused by MERS-CoV), SARS (caused by SARS-CoV-1), and COVID-19 (caused by SARS-CoV-2).1,2 A scan of over 200 representatives from the Chinese medicinal flora database against Severe Acute Respiratory Syndrome–associated coronavirus (SARS-CoV) indicated Lycoris radiata (L’Hér.) Herb. to be the most potent.
56
Its EtOH stem extract exhibited an EC50 of 2.4 µg/mL against SARS-CoV propagation in Vero cells that was, relative to interferon-α, over 14 times lower than that observed for the second most active plant (Artemisia annua L., Asteraceae).
56
Furthermore, this effect was shown to be selective as the cytotoxic concentration (CC50) of Lycoris radiata was ascertained to be as high as 886.6 µg/mL.
56
Accompanying this discovery was the identification of lycorine (
Activities Against Orthomyxoviridae
Orthomyxoviridae is a family of negative-sense RNA viruses containing 7 genera, which are Isavirus, Thogotovirus, Quaranjavirus, Alphainfluenzavirus, Betainfluenzavirus, Gammainfluenzavirus, and Deltainfluenzavirus.
1
The latter 4 of these, as their names suggest, are notorious for causing influenza in mammals and birds.
1
A further Lycoris species examined for antiviral effects was the Korean medicinal plant Lycoris squamigera Maxim.
57
Its leaf, stem, and bulb MeOH extracts were subjected to the hemagglutinin test for Avian influenza virus (H9N2) (Orthomyxoviridae) infection of chicken red blood cells.
57
Positive tests were obtained for each dilution of the bulb extract, in the range 400, 300, 200, 100, 50, 25, and 12.5 µg/mL.
57
The leaf extract was only active at the 400 and 300 µg/mL levels, while the stem extract remained active up to the 50 µg/mL mark.
57
Furthermore, no compounding cytotoxic effects were observed with any of the extracts at the highest tested concentration.
57
The Lycoris squamigera extractive lycorine (
Conclusions
Interest in viruses and viral pathogenesis has intensified in recent times due to the COVID-19 pandemic. Plants have long served as an attractive platform for antiviral drug discovery, affording not only broad diversity and appealing chemical traits but also close alignment to traditional systems of medicine. The plant family Amaryllidaceae has met all of these criteria with aplomb. Indigenous knowledge has to a significant extent guided the antiviral screening measures, which have led to the identification of the active extracts some of which displayed effects at the submicrogram per milliliter level. These have in turn guided the isolation and identification of the active principles, which largely evolved out of the isoquinoline class of Amaryllidaceae alkaloids. Since only 18 species, out of nearly 1000 in the family, have to date been screened, going forward there is an abundance of plant materials to be explored. Intriguingly, the majority of taxa that have documented uses in TM for viral diseases remain to be investigated. Together, these should fortify the procurement of both known and novel alkaloid entities, whose chemical characteristics should allow for facile modifications to be made to their structures. Although useful information has emerged out of in silico models of study, much of the data gleaned from such enterprises is yet to be tested at the laboratory level.
Footnotes
Acknowledgements
Financial support by the University of KwaZulu-Natal is duly acknowledged by both authors.
Author Contributions
JvS was responsible for the conceptualization and design of the study. JJN undertook the literature review, analysis of data, and drafting of the text. Proofreading and critical revision of the text were carried out by JvS.
Declaration of Conflicting Interests
The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.
Funding
The author(s) received no financial support for the research, authorship, and/or publication of this article.
