Antiviral activity of medicinal plant

ANTIVIRAL ACTIVITIES OF MEDICINAL PLANTS B.Pharma Final Year

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VIRAL INFECTIONS

• LIST OF VIRUSES AND VIRAL DISEASES:

A: Adenovirus

AIDS: Cuteneous lesion associated with AIDS

AIDS related malignant tumours

Alphaviruses causing Encephalitis

Arenavirus

Argentine hemorrhagic fever

Arthropod-borne viral encephalitis

Avian Influenza (Bird Flu)

B:

Bolivian Hemorrhagic Fever

C: Chickenpox

Chikungunya

Coxsackievirus Infection

Crimean-Congo Hemorrhagic Fever

Cytomegalovirus infection

D:

Dengue

E: Eastern equine encephalitis Ebola Virus Infection

Echovirus Infection

Epstein - Barr virus infection

Epstein-Barr Virus Related Malignant Tumors


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F: Fifth Disease

Filovirus

Flavivirus

G:

German measles

H: Hand, foot & mouth disease

Hemorrhagic fever with renal syndrome

Herpes Virus(Herpesviridae)

Herpes Simplex Virus Infection

Herpes Zoster Virus (Shingles)

Human Papilloma Virus Associated Epidermal Lesions

Human Papilloma Virus in Cervical Cancer

I: Infectious Mononucleosis

Influenza

J:

Japanese Encephalitis

K: Kaposi Sarcoma

Korean Hemorrhagic Fever

Kyasanur Forest Disease

L: Lassa Fever

Lymphocytic choriomeningitis


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M: Marburg Virus Disease

Measles

Molluscum Contagiosum

Mumps

Murray Valley encephalitis

N:

Norwalk Virus related Diarrhea

O: Omsk hemorrhagic fever

Orthomyxoviruses

P: Parainfluenza Virus Infection

Paramyxovirus

Parvovirus B19 Infection

Picornavirus

Poxviruses

R: Rabies

Respiratory syncytial virus infection

Rift Valley Fever

Rotavirus diarrhea

Rubella

Rubeola

S: Smallpox

St. Louis Encephalitis

T:

Tick-borne Encephalitis


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V: Varicella

Variola

Venezuelan equine encephalitis

Viral hemorrhagic fevers

Viruses in Leukemia and Lymphoma

W: Western equine encephalitis

West Nile Virus disease

Y:

Yellow Fever


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� BRIEF DISCUSSION ABOUT THE INFECTIONS :

Adenovirus

Adenoviruses were first isolated in 1953 by Rowe and co-workers from human adenoids

removed at surgery.

There are 51 immunologically distinct human adenovirus serotypes - (6 species: Human

adenovirus A through F). Many serotypes are not linked to a specific disease.

Human adenoviruses have a capsid with icosahedral symmetry.

Rodlike structures with knobs at the ends protrude from the capsid. The genome of the

virus is a double-stranded DNA linear molecule.When infecting cells in vitro,

adenoviruses are capable of lytic infection, latent infection, and transformation.

Because of the ability to transform cells and to produce tumours in rodents, they have

been considered possible human tumour viruses, but up to now there has been no

evidence that links adenoviruses to human tumours.

AIDS: Cutaneous lesion associated with AIDS

The acquired immune deficiency syndrome (AIDS) is caused by human immune

deficiency virus (HIV) infection, damaging the cell mediated immune system.

Skin is the most commonly affected organ in HIV infection. Cutaneous lesions in HIV

positive patients serve as a marker of HIV infection and also indicates the stage of the

disease.

Opportunistic infection patterns are different in different parts of the world and change

as people migrate.

Opportunistic infections in HIV positive patients have decreased since introduction of the

therapy. Following therapy there is fall in viral titre and increase in CD4 cells. The


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cutaneous conditions improve or decline in incidence after the therapy. In many

developing countries HAART is still not widely available.

Viral:

Molluscum contagiosum: In HIV- positive patients this lesion presents as persistent

umbilicated or verrucous papules, commonly on the head and neck region.

Histopathology reveals cup-shaped lesion containing molluscum bodies. Giant and warty

verrucous mollusca contagiosa are markers of advanced HIV infection.

Herpes Simplex Usually occurs in perianal, genital and orofacial skin. In HIV and HSV

coinfection the lesions last for more than one month. Histopathology reveals numerous

intranuclear and intracytoplasmic nuclear inclusions. Extensive ulceration and

intraepidermal acantholytic vesicles are noted .

Varicela zoster virus infection Clues to HIV and VZV coinfection - The lesions usually

occur in younger patients. These are more severe lesions and of longer duration.

Human papillomavirus infection there is a high incidence of common and anogenital wart

in HIV positive patients. Condylomata acuminata may occur in HIV infected homosexual

men. There is risk of dysplasia in perianal condyloma. Grossly, these lesions may

present as smooth sessile plaques to exophytic cauliflower plaques. Verrucae vulgaris,

multiple plantar warts, flat and filiform warts may be noted in HIV infected patients.

Common warts are frequently present on the bearded area of the face in HIV positive

patients.

Cytomegalovirus: Almost 90% HIV positive patients develop CMV infection.

Histological examination reveals CMV inclusions in endothelial cells and fibroblasts

together with areas of epidermal necrosis .

Oral hairy leukoplakia: Poorly defined projections are noted on the lateral borders of the

tongue.This lesion indicates advanced immunosuppression. Causative organisms include

Epstein-Barr virus, human papillomavirus or candida. Histological examination reveals

some acanthosis and parakeratosis. Large pale staining cells resembling keratinocytes are

present.


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AIDS related malignant tumours

Patients with HIV infection are at increased risk for developing Kaposi's sarcoma, non-

Hodgkin's lymphoma, and several other cancers.

Accumulating data suggest that HIV-infected patients also are at increased risk for

developing Hodgkin's lymphoma, cervical carcinoma in situ (CIS), other anogenital

neoplasms (invasive cancer and CIS), leiomyosarcoma, and conjunctival squamous cell

carcinoma.

AIDS is caused by the human immunodeficiency viruses HIV-I (present globally) and

HIV-II (mainly restricted to West Africa and contacts with that zone).

It is generally assumed that HIV-1 infection play a passive role in cancer development by

impairing the host immune surveillance and increasing the risk of oncogenic virus

infection. Recent insights, however, indicate that HIV-1 infection more actively promotes

cancer growth.

HIV is not per se oncogenic, but a wide range of tumours is now described with increased

incidence among HIV-infected people and/or with a more aggressive clinical course,

compared with HIV-uninfected people.

Kaposi’s Sarcoma (KS):Kaposi’s Sarcoma (KS):Kaposi’s Sarcoma (KS):Kaposi’s Sarcoma (KS):

One of the earliest opportunistic diseases that defined AIDS (even before the etiological

link of AIDS with infection by HIV was recognized) was KS in skin and viscera.

Whether KS is a genuine neoplasm or a reactive hyperplasia of endothelium is still

disputed.

The incidence of KS among HIV-infected people varies with risk-factors for HIV

transmission and geography ; high in homosexual men and African adults and children,

low in haemophiliacs; more frequent in east than West Africa. Epidemiological blood and

tissue studies support its etiological role in development of KS.


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Lymphoma:Lymphoma:Lymphoma:Lymphoma:

High-grade B-cell lymphoma, particularly in the central nervous system, is a major cause

of morbidity and mortality in HIV/AIDS.

Tumours are usually extra-nodal and destructive.

They are commonly associated with Epstein-Barr Virus infection.

Incidence is inversely related to CD4+ T-cell count, which may explain why it is less

common in patients in Africa and India.

T-cell lymphomas, Hodgkin’s disease and myeloma have a slightly increased incidence

compared with HIV-uninfected people.

ConjuncConjuncConjuncConjunctival carcinoma:tival carcinoma:tival carcinoma:tival carcinoma:

The incidence of conjunctival squamous cell carcinoma is closely related to exposure to

UV light. In equatorial, though not elsewhere, there is an epidemic of this malignancy

among HIV-infected adults. A proven co-factor in a proportion of cases is HPV-16.

Other Tumours:Other Tumours:Other Tumours:Other Tumours:

Small series and case reports of cancers in HIV-infected patients strongly suggest an

increased incidence and more aggressive behavior of other epithelial tumours. These

include skin squamous cell carcinoma and basal cell carcinoma and melanoma. They

are generally present in the later stages of HIV disease. The relative risks for the most

common epithelial cancers in the general population -lung, breast, colon/rectum,

stomach, liver, and prostate - are not increased substantially in people with AIDS,

however.

Most cancers seen in the AIDS setting are related to oncogenic virus infections, such as

Epstein-Barr virus (EBV), Kaposi's sarcoma (KS)-associated herpesvirus (KSHV) and

human papillomavirus (HPV).


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Clinical evidence suggests that the oncogenicity of HPV is altered by the presence of

HIV-1 infection irrespective of host immune status.

Avian Influenza (Bird Flu)

Avian influenza in birds:

It is an infection caused by avian (bird) influenza (flu) viruses. These viruses occur

naturally among wild birds worldwide in their intestines, but they do not get sick. It is

very contagious among birds including domestic birds eg. Chickens, ducks & turkeys and

kill them.

Virus are excreted from infected birds in their saliva, nasal secretions, & feces. Birds

including domestic ones become infected through direct contact with other infected

poultry, or through contact with infected surfaces (such as dirt or cages) or materials

(such as water or feed).

It causes two main forms of diseases in poultry.

Human infection :

“Human influenza virus” is those subtypes that spread widely among humans. There are

only three known a subtypes of influenza viruses (H1N1, H1N2, and H3N2).Viruses are

constantly changing, and they adapt over time to infect and spread among humans.

Avian Influenza A (H5N1):

Subtype Influenza A “H5N1 virus” that occurs mainly in birds, is highly contagious and

are deadly to them. Infections with these viruses have also occurred man.


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Bolivian Hemorrhagic Fever

Bolivian hemorrhagic fever is an acute febrile disease with hemorrhagic manifestations

that are almost identical to those of Argentine hemorrhagic fever . It is a disease endemic

to northeastern Bolivia.

The etiological agent is Machupo virus (MACV, Arenaviridae).

The virus is similar to that which causes Argentine hemorrhagic fever, and its reservoir is

also wild rodents.

Bolivian hemorrhagic fever was first recognized in rural areas of of northern Bolivia

(1959), and there were two major epidemics in Bolivian villages during 1962-64.

Chickenpox

Varicella (chickenpox) is an acute vesicular exanthem caused by the varicella-zoster

virus, an agent that has a worldwide distribution and for which humans are the only

known host. There are no animal reservoirs.

VZV (HHV3) is a neurotrophic alpha-herpesvirus.

VZV is named after : Varicella, an alternative name for chickenpox (the primary VZV

infection) Zoster, another name for shingles (reactivation of latent VZV infection).

Although all age groups are susceptible, in temperate zones chickenpox affects mostly

children and in the tropics mostly young adults. The virus, which is spread through

inhalation of droplets or by direct contact is highly contagious from about 24 hours

before the initial eruption to a week of more thereafter. Although infection with varicella-

zoster virus establishes lifelong immunity and chickenpox does not recur, the latent viral

genome may be activated years later to cause shingles.

No immune persons (usually children) are susceptible to primary infection with varicella-

zoster virus.


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Chickenpox begins as a “silent” infection of the nasopharynx, with local replication of

varicella-zoster virus.

After an incubation period of 10 to 23 days, the virus enters the bloodstream, causing

viremia and a sudden onset of fever, malaise, and anorexia.

In the circulation it seeds reticuloendothelial cells, an effect that leads to secondary waves

of viremia. The virus then disseminates to skin and viscera, and within 24 hours a red

maculopapular eruption develops, usually on the upper trunk and face.

The papules rapidly become clear vesicles with an erythematous base.

In the next 24 hours, the vesicles become cloudy, the eruption begins to itch, and

scratching may rupture the vesicles. Separate crops appear for 3 to 6 days and spread

peripherally. After the last crop, the scabs heal without scarring.

Though vesicles of the skin are generally painless, painful lesions may develop on

mucous membranes, such as the cornea and tympanic membrane.

Complications include pneumonia, encephalitis, hepatitis, carditis, keratitis, orchitis,

arthritis, hemorrhages and acute encephalopathy with fat accumulation in the viscera

(Reye’s syndrome).

Histologically, the skin lesions initially show ballooning of epidermal cells.

Later, unilocular vesicles containing proteinaceous fluid, degenerating cells and syncytial

giant cells are seen.

Cowdry type A intranuclear inclusions are seen in epidermal cells, endothelial cells of

superficial capillaries, reticuloendothelial cells, and fibroblasts.

The affected organs in chicken pox exhibit spherical foci of coagulative necrosis.

At the margin of these necrotic foci, surviving cells contain intranuclear inclusions.

Chikungunya

Chikungunya virus (CHIKV) is a toga virus belonging to the genus alpha virus,

indigenous to tropical Africa and Asia, where it is transmitted to humans by the bite of

infected mosquitoes, usually of the genus Aedes.

Chikungunya (CHIK) fever, the disease caused by CHIKV, was first recognized in

epidemic form in East Africa during 1952-1953.


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Since the beginning of 2006, this crippling mosquito-borne disease (Chikungunya virus

infection) has shown an explosive emergence in nations in the Indian Ocean area. By

March 7, 2006, 157,000 people had been infected in the French island La Reunion, and

the disease had spread to the islands of Seychelles, Mauritius, and Mayotte (French).

Subsequently, the disease appeared in India, China, and European countries.

Infection in Aedes aegypti salivary glands is asymptomatic and lifelong. It is transmitted

by Aedes aegypti to human, there is no human to human transmission.

The disease is highly infectious.

Its main symptoms are sudden onset of chills, fever, headache, rash, and debilitating

arthralgia . The symptoms may persist for several weeks.

The word "chikungunya" is thought to derive from description in local dialect of the

contorted posture of patients afflicted with the severe joint pain associated with this

disease.May rarely cause hemorrhagic fever, specially in children.Incubation period is 1

to 12 days. Fever lasts for 3 - 7 days, arthralgia may persit for upto 3 weeks.

Because CHIK fever epidemics are sustained by human-mosquito-human transmission,

the epidemic cycle is similar to those of dengue and urban yellow fever.

Large outbreaks of CHIK fever have been reported recently on several islands in the

Indian Ocean and in India.

Though fetal contamination risks appear to be rare before 22 weeks of gestation, they are

potentially dangerous. After 22 weeks gestation, newborns infection occurs if the mother

is viremia positive at delivery. Transplacental transmission is suspected, but the

pathogenic mechanism remains unknown.

Warm, humid climates and water reservoirs serve as an excellent breeding ground for the

vector of the virus, Aedes mosquitoes. With an increase in temperature, susceptibility of

mosquitoes to CHIKV increases.

Although the disease is self-limiting, the risk to non-immune travellers from other parts

of the world to areas with a chikungunya epidemic, continues to exist and should be

included in the differential diagnosis of travellers returning home with fever.

Due to similarities in clinical presentation with dengue, limited awareness, and a lack of

laboratory diagnostic capability, CHIKV is probably often under diagnosed or

misdiagnosed as dengue .


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Treatment is supportive. Analgesic and anticonvulsants are often used.The prognosis is

generally good, although some patients experience chronic arthritis. There are very few

recorded fatalities.

With no vaccine or antiviral available, prevention and control depends on surveillance,

early identification of outbreaks, and vector control. CHIKV should be borne in mind in

sporadic cases, and in patients epidemiologically linked to ongoing local or international

outbreaks or endemic areas.

Dengue

Dengue the most prevalent arthropod-borne viral (Arborvirus) disease of humans caused

by four serotypes of dengue virus (DENV 1-4) of the genus Flavivirus.

It is transmitted to man by mosquito Aedes aegypti. It is common in tropical and

subtropical countries, especially in coastal areas. (World distribution of dengue viruses

and their mosquito vector, Aedes aegypti : CDC)

Source: Man is infective to mosquito and mosquito transmits the disease to man.

Echovirus Infection

Echoviruses have been subdivided on the basis of antigenic relationships and differences

in host range into polioviruses , coxsackie viruses groups A and B, and echoviruses.

Echoviruses are RNA viruses of the genus Enterovirus and the family Picornaviridae.

The echoviruses (enterocytopathogenic human orphan viruses) were isolated from faecal

specimens of healthy children.

They produce cytopathic effects in primate cell cultures, but at isolation they were

nonpathogenic for suckling mice or primates.

They are immunologically distinct from polioviruses and have been associated with a

variety of diseases including nonspecific febrile illnesses with or without respiratory

symptoms, aseptic meningitis, paralysis and encephalitis, exanthema, generalized disease


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of new-born and neonatal diarrhea.They also have been associated with chronic meningo-

encephalitis in immaglobulinemic patients.

Flavivirus

The Flaviviridae include almost 70 viruses, nearly half of which have been associated

with human disease. Flavivirus is a genus of the family Flaviviridae.

Flavivirus share a common size (40-60 nanometres), symmetry (enveloped, icosahedra

nucleocapsid), nucleic acid (positive-sense, single stranded RNA approximately 10,000-

11,000 bases), and appearance in the electron microscope.

These viruses are among the most important arthropod-borne viruses worldwide and

include dengue, yellow fever, and Japanese encephalitis viruses.

Morbidity and mortality caused by these viruses vary, but collectively they account for

millions of encephalitis, hemorrhagic fever, arthralgia, rash, and fever cases per year.

Most of the members of this family are transmitted between vertebrate hosts by arthropod

vectors, most commonly mosquitoes or ticks. Transmission cycles can be simple or

complex depending on the hosts, vectors, the virus, and the environmental factors

affecting both hosts and viruses.

German measles

German measles, or rubella, is a mild, systemic infection of childhood.

It is a pleomorphic RNA virus in the Togaviridae family of the genus Rubivirus.

In older children and adults, especially women, it may be more severe, with joint

involvement and purpuric rash. Except for superficial similarity in the name, the rubella

virus is not related to the measles (rubeola) virus.

A worldwide disease, rubella is characterized by measles-like rash, low-grade fever, and

swollen posterior auricular and occipital lymph nodes. The congenital rubella syndrome

(CRS) involves multiple organ systems and has a long period of active infection and virus

shedding in the postnatal period.


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Subclinical infections are common. The incubation period is 12 to 21 days, and the mild

rash and other symptoms resolve within 3 days. Initially, swollen, nontender posterior

auricular and occipital lymph nodes appear. A day or two later the nonpruritic rash begins

on the face and rapidly spreads over the rest of the body, sparing the palms and soles.

The virus is very contagious and is shed from the nasopharynx. The virus is also

transmitted through the placenta, and rubella infections in pregnant women are a serious

public health concern, because intrauterine infection causes spontaneous abortion, fetal

death and a variety of congenital abnormalities. Infection during the first trimester is most

serious. The classic triad of congenital rubella includes ocular abnormalities, heart

disease, and deafness.

The principal congenital abnormalities are patent ductus arteriosus, pulmonary and aortic

stenosis, coarctation of the aorta, defects of the atrial or ventricular septum, ocular lesions

(cataracts, glaucoma, and chorioretinitis), deafness, microcephaly, mental retardation, and

retarded growth. Congenital rubella produces a classic salt-and-pepper pigmentation of

the fundus.

Herpes Simplex Virus Infection

Herpes simplex virus can be divided into two types on the basis of antigenicity,

pathogenicity and genetic properties.

Herpes simplex type II is the major cause of urogenital infections, whereas Herpes

simplex type I is more often isolated from nongenital infections.

Herpes simplex viruses have a worldwide distribution, and direct contact with infected

secretions is the principal mode of spread. Herpes simplex causes disease of the skin and

mucosa such as acute gingivostomatitis, recurrent stomatitis, oral ulcers (cold sores),

herpetic keratoconjunctivitis, herpetic esophagitis, and genital herpes.It also causes

systemic disease as in disseminated herpetic infection of infants, acute necrotizing

encephalitis of the adult, and infections in the immunodeficient host.


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Cytopathologic changes in Herpes infection:

Two types of cytopathology seen in herpes simplex infections are rounding and

degeneration of the cells with inclusion body formation and cell fusion with formation of

syncytia. The intranuclear inclusion is the characteristic cellular lesion, and it consists of

a single, eosinophilic, well-demarcated inclusion body surrounded by a halo and

marginated chromatin.

By electron microscopy the inclusion body consists of paracrystalline arrays of viral

capsids and electron-dense glycoprotein.

Herpetic infection of skin and mucosa:

Herpetic infection of skin and mucosa is manifested by the appearance of red papules that

quickly become vesiculated.

Histologically there is degeneration of the cells in the stratum malpighii and the stratified

squamous epithelium.

In early stages one observes ballooning of the epidermal cells and clearing of the nuclear

chromatin. There is acantholysis and formation of a unilocular vesicle. The typical

inclusion bodies (Cowdry type A) can be seen in the epidermal cells.The upper dermis or

submucosal portions of the affected regions show an inflammatory infiltrate around

capillaries.

The histologic lesion of herpes simplex in skin and mucosa is practically

indistinguishable from that caused by herpes zoster. Immunohistochemistry using

fluorescent antibodies or immuno-peroxidase can, however, make the distinction between

these viruses and even allows typing of the herpesvirus.

Disseminated herpetic disease:

Disseminated herpetic infection of the newborn affects neonates in the first 4 weeks of

life as well as an older group of infants in whom it causes hepatoadrenal necrosis.

Autopsies of infants dying from disseminated herpetic disease show multiple military

foci of necrosis surrounded by hemorrhagic borders in the liver, adrenal gland, and brain.

Microscopically these foci consist of a zone of central coagulative necrosis surrounded by

an area of hyperemia with a sparse mononuclear inflammatory inflammatory reaction.

Inclusion bodies characteristic of the herpetic infection are found in nuclei of the

parenchymal cells surrounding the necrotic zone.


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Pathologic combination may be complicated by the presence of gram-negative

septicemia. Morphologic manifestations of disseminated intravascular coagulation (DIC)

may be also prominent. The virus can be easily easily demonstrated in the lesions by

electron microscopic examination.

Acute necrotizing encephalitis:

Herpes simplex virus has been shown to be a cause of acute necrotizing encephalitis of

the adult. Patients with this syndrome may display a picture of encephalitis or a clinical

picture of an expanding lesion, often of the temporal lobe.

In fatal cases the brain shows asymptomatic necrosis particularly prominent in the

temporal lobes but also involving the hippocampus and the posterior occipital cortex.

Hemorrhage may be prominent. In the early stages of the disease the histopathologic

picture is that of acute necrosis associated with a diffuse meningoencephalitis.

Inclusion bodies within the nuclei of neurons and glial cells can be seen but on occasion

are difficult to find.Reactivation and dissemination of herpetic lesions are not uncommon

in immunocompromised hosts. Esophageal and upper respiratory tract lesions are

commonly found at autopsy in patients who have undergone intense chemotherapy.

Dissemination occasionally occurs, and a picture similar to the disseminated disease in

newborns can be seen.

Dissemination with fatal outcome also occurs in some severely burned patients.

Evidence is accumulating that herpes simplex virus (HSV) infection is implicated in

oncogenesis. HSV antigens have been observed in some oral cancers.

Influenza

Orthomyxoviruses (influenza A, B, and C viruses) are enveloped viruses that contain

single-stranded RNA in a helical array of nucleoprotein. Inserted in the lipid envelop are

two glycoproteins- the hemagglutinin (HA) and the neuraminidase (NA).

Influenza viruses have the unusual capacity of changing the antigenic identities of their

HA and NA polypeptides, thus creating numerous antigenic variants. The host of

different types of virus give their names to the disease, as in, for example, human, swine,

and avian influenza.


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Lassa Fever

The arenavirus Lassa causes a severe illness called Lassa fever. Lassa fever was first

recognized in herdsmen’s village in Nigeria in 1969. Lassa fever is known to be

endemic in Guinea (Conakry), Liberia, Sierra Leone and parts of Nigeria, but probably

exists in other West African countries as well. Lassa fever has been seldom investigated

outside of a few hyperendemic regions, where the described epidemiology may differ

from that in areas of low or moderate incidence of disease.

Lassa fever is a serious, highly infectious, viral, hemorrhagic disease characterized by

high fever, prostration, generalized hemorrhages, abdominal pain, vomiting, diarrhea,

severe pharyngitis, dyspnea, serous effusions, facial edema, and shock. It is fatal in

almost half the cases. Some studies indicate that 300 000 to 500 000 cases of Lassa fever

and 5000 deaths occur yearly across West Africa.

The Lassa virus emerged suddenly and may be a virulent mutant of the lymphocytic

choriomeningitis virus.

Measles

Measles is a highly infectious, systemic infection of childhood, caused by measles virus.

Up to 6 months of age, infants are protected, due to antibodies from mothers, but the

reafter are vulnerable. Measles vaccine has almost eliminated measles.

Mode of infection: Humans are the only natural host.

i) Infection is most commonly transmitted by inhalation of contaminated droplets.

ii) May also be passed through the placenta.

Incubation period: About 14 days (10-21 days).


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Mumps

Mumps is an acute but usually mild viral infection of childhood, characterized by swollen

and inflamed salivary glands (most often the parotids). Less often the virus attacks the

pancreas, ovaries, testes, and other organs. Mump virus is a highly contagious

paramyxovirus that is commonly transmitted in respiratory droplets. A vaccine made

from attenuated live mumps virus has reduced the frequency. Most adults are immune.

Nonimmune adults who acquire the infection may have a painful and debilitating illness.

The incubation period is 2 to 4 weeks, after which fever, headache, malaise, and swelling

and inflammation of the salivary glands follow.Swelling and other acute symptoms

usually subside within 2 weeks. Mumps is usually diagnosed clinically from swollen

salivary glands and confirmed by finding rising titers to mumps virus in the serum of

convalescent patients.

Norwalk Virus related Diarrhea

The prototype “Norwalk agent” was discovered in an outbreak of diarrhea and vomiting

among the elementary school students in Norwalk, Ohio. The virus has icosahedral

symmetry, contains single-stranded RNA, and thought to be a Calicivirus.

Norwalk-like viruses have been associated with gastroenteritis, primarily in adults and

older children. Diarrhea and vomiting are of short duration, and dehydration is rare (as

compared with rotavirus diarrhea ).

The outbreaks have occurred at all times of the year and have been associated with

contaminated water and foods (eg. oysters) infected food-handlers, and person-to-person

spread.


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Poxviruses

The causative viruses are large with a DNA core and a surrounding capsid. Poxvirus

infection occurs in human and animals. These are members of Poxviridae family. There

are two subgroups, based on the morphological features of the virus. The viruses of

molluscum contagiosum and orf are oval or cylindrical in shape and measure

approximately 150x300 nm. The remaining viruses are brick-shaped and range in size in

size from 250 to 300nm x200 to 250 nm.

Clusters of these poxviruses can be identified in hematoxylin and eosin-stained sections

as intracytoplasmic eosinophilic inclusions. The poxviruses fall into different subgroups

depending on antigenic relationships.

Rabies

Rabies viruses are a RNA viruses of the genus Lyssavirus and the family

Rhabdoviradae.

Rabies remains as one of the most feared zoonotic diseases in the world. In humans it is a

lethal encephalitis. Carnivores, such as the dog, wolf, fox, and skunk, are the principal

reservoirs, but the infection extends to bats and to some domesticated animals, such as

cattle, goats, and swine. The infectious agent is transmitted to humans through

contaminated saliva introduced into the wound of a bite.The virus enters a peripheral

nerve and is transported by centripetal axoplasmic flow to the spinal cord and brain.

The latent interval varies in proportion to the distance of transport, being as short as 10

days or as long as 3 months. Centrifugal intra-axonal transport of the virus contaminates

visceral organs, particularly the salivary glands, which in turn contaminate the saliva.


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Rotavirus diarrhea

Rotaviruses contain double-stranded RNA, resemble a wheel, have icosahedral

symmetry, and are “double-shelled” (with inner and outer capsid). Rotavirus particles

have been found in the duodenal mucosa of children with acute gastroenteritis and in

diarrheal stool specimens. In developed countries rotavirus is the most common pathogen

of childhood diarrhea. Rotavirus-induced diarrhea is an endemic problem throughout the

world, the organisms being identified in half the children in developing countries.

In temperate countries rotavirus diarrhea usually has a seasonal winter peak, but in

tropical countries high rates are observed throughout the year.

Nosocomial infections are common, and shedding of rotavirus has been found in some

(usually asymptomatic) newborns in communal obstetric nurseries within 3 to 4 days of

birth.Antibodies to the rotavirus in colostrums and breast milk protect against infection.

The highest incidence of symptomatic infection is in children aged 7 to 24 months. Most

children have rotavirus antibodies by end of the third year, and nearly all infections in

adults are subclinical.

Smallpox

Before its eradication, smallpox (variola) was an acute, highly contagious, exanthematous

viral infection. The virus contains a double-stranded DNA and produces a typical plaque,

or “pock”, when cultured on chorioallantoic membrane of embryonated chicken eggs.

Jenner’s pioneering work in 1796, a similar virus - vaccinia, the causative agent of

cowpox - has been used for “vaccination” to protect against small pox.Smallpox was

evidently an ancient disease; a rash resembling smallpox was found in the mummified

remains of the Egyptian Pharaoh Ramses V, who died in 1160 BC. The disease once had

worldwide distribution in both urban and rural areas, afflicting persons of both genders

and all ages, but particularly children.In 1967, the World Health Organization began its

uniquely successful campaign to eradicate smallpox.


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By then, smallpox had already been controlled in most developed countries but was still

endemic in the less developed world. In 10 years the vaccination campaign eradicated the

disease.

Viral hemorrhagic fevers

Viral hemorrhagic fevers are a group of distinct acute viral infections that cause varying

degrees hemorrhage and shock, and sometimes death. There are many similar viral

hemorrhagic fevers in different parts of the world, often named for the area where they

are first described.

Yellow Fever

Yellow fever (YF) is an infectious, non-contagious disease caused by an RNA virus of

the family Flaviviridae (that cause dengue and other hemorrhagic fevers), which is

transmitted to man by the bite of hematophagous mosquitoes. First described in the

Caribbean, yellow fever is the oldest known viral hemorrhagic fever.It is a re-emerging

infectious disease that currently is at risk of urbanization due to the advance of the Aedes

aegypti vector.

Geographic distribution: The disease affects about 200,000 individuals annually, mainly

in tropical Africa and South America, including both jungle and urban settings.It is a

significant hazard to unvaccinated travelers to these endemic areas. Recent increases in

the density and distribution of the urban mosquito vector, Aedes aegypti, as well as the

rise in air travel increase the risk of introduction and spread of yellow fever to North and

Central America, the Caribbean, the Middle East, Asia, Australia, and Oceania.

Mode of Infection: The usual reservoir is tree dwelling monkeys, virus being passed

among them in the forest canopy by mosqitoes. These monkeys are a good reservoir

because the virus neither kills them nor makes them ill . Humans acquire jungle yellow

fever by entering the forest and being bitten and inoculated by Aedes mosquitoes. Felling

trees increases the risk because mosquitoes are brought down with the tree. On returning

to the village or city, the victim becomes the reservoir for epidemic yellow fever in the

urban setting , where Aedes aegypti is the vector.


23

Hence, virus transmission occurs between humans, mosquitoes, and monkeys. The

mosquito, the true reservoir of YF, is infected throughout its life, and can transmit the

virus transovarially through infected eggs. Man and monkeys, on the other hand, play the

role of temporary amplifiers of the virus available for mosquito infection.


24

� List of plants which extract used as antiviral medicine

Species Common name Claim

Acanthopanax gracilistylus

Prickly ginseng, Wujiapi Aids digestion, cures hepatitis C, lowers blood pressure, increases stamina.

Achillea millefolium Yarrow

Eaten to counter poisoning, but must be eaten quickly

Agaricus subrufescens

Agaricus blazei

May enhance immune system and have anti-cancer properties (Reviewed by Hetland) ]

Allium sativum Garlic

Antibiotic (in vitro)/stops infectionNicole Johnston (April 2002). "Garlic: a natural antibiotic". Modern Drug Discovery

Cardiovascular health

Aloe ferox

Anethum graveolens Dill and Dill oil used to soothe the stomach after meals

Amorphophallus konjac

Konjac

Atopic dermatitis

high cholesterol

Arnica montana Arnica

Used for strains, sprains, and bruises. The roots contain derivatives of thymol,[14] which are used as fungicides and preservatives and may have some anti-inflammatory effect.

Aquilaria agollocha Eaglewood

Artemisia annua L. Sweet sagewort

Help to prevent the development of parasite resistance,it also has anti-malarial properties, and has anti-cancer properties

Artemisia absinthium L.

Wormwood Removal of internal parasites

Aristolochia rotunda Smearwort

Arum maculatum Lords and Ladies

Astragalus membranaceus

Astragalus

Cannabis Sativa

Cannabis, Cannabis sativa, Marijuana, Hashish

Pain relief, hunger stimulation, wasting caused by HIV/AIDS, Glaucoma, nausea, | anticarcinogenic through angiogenesis inhibition through various


25

agents such as tetrahydrocannabinol, cannabidiol, hexahydrocannabinol, quinone, arachidonyl ethanolamide

Citrus aurantium ssp. bergamia

Bergamot orange Malaria

Crataegus spp. L. Hawthorn Nervous tension

Cydonia oblonga Quince

Cymbopogon flexuosus

Lemon grass

Cymbopogon schoenanthus

Fever grass

Digitalis lanata Digitalis, Balkan Foxglove Antiarrhythmic agent and inotrope

Echinacea purpurea

Purple coneflower, and other species of Echinacea

Reduce the severity and duration of symptoms associated with cold and flu.

Filipendula ulmaria (Spiraea ulmaria)

Meadowsweet

Fevers and inflammations. Pain relief. Ulcers. Bacteriostatic. Listed as therapeutical in 1652 by Nicholas Culpeper. In 1838, salicylic acid was isolated from the plant. The word Aspirin is derived from spirin, based on Meadowsweet's synonym name Spiraea ulmaria.

Glycyrrhiza glabra Liquorice

Hydrastis canadensis

Goldenseal Antimicrobial

Hypericum perforatum

St. John's wort Antidepressant

Kaempferia galanga

Galanga resurrectionlily, Shannai

Marrubium vulgare Horehound Expectorant

Matricaria recutita (Chamomilla recutita)

Chamomile Relaxant/Calmative

Mentha × piperita Peppermint Irritable Bowel Syndrome/Peristalsis

Nepeta cataria Catnip Soothes coughs

Panax Ginseng

Papaver somniferum Opium Poppy

Pain relief. Morphine made from the refined and modified sap is used for pain control in terminal patients. Dried sap was used as a traditional medicine until the 19th century.


26

Passiflora spp. Passion-flower Insomnia

Phytolacca spp. Pokeweed

Topical: acne

Internal: tonsilitis

Plantago spp. Plantain and Psyllium Astringent

Salvia stenophylla Blue Mountain Sage

Poppiocious seediouphylla

Poppy seeds Helps sleeping/relieves pain

Rosmarinus officinalis

Rosemary

Salix alba White willow

Ancient medicine, already described by Greek pharmacologist Dioscorides. Bark contains salicylic acid, which name is derived from Salix.

Symphytum officinale

Comfrey Stops infection

Salvia officinalis Sage Improves cognitive function in mild to moderate Alzheimer's disease.

Tanacetum parthenium (Chrysanthemum parthenium)

Feverfew Relieves Migranes, helps fevers and chills.

Taraxacum officinale

Dandelion Digestive

Tilia spp. Lime Blossom

Urtica dioica Stinging Nettle

Valeriana officinalis Valerian Sedative

Verbascum thapsus Mullein

boosts the Immune system, antispasmodic, diuretic, anodyne, and demulcent Used to treat coughs, (protracted) colds, hemoptysis, catarrh, dysentery, diarrhoea and as a general tonic (like ginseng) to boost the immune system

Zingiberis rhizoma Ginger can help ease nausea from chemotherapy


27

There are hundreds of plants used all over the world, which are used in herbal medicine

as treatments for viral infections.

Here are some of the most accessible and reliable natural antiviral herbs:

Astragalus (astragalus membranaceus) : has been in the Chinese Materia Medica for

centuries and has shown to have immune boosting properties. This classic energy tonic

has a warming and toning effect. Although not used as a treatment for acute illnesses,

astragalus is believed to be very useful as an herbal remedy for treating viral infections,

including those that cause the common cold and flu.

Echinacea (eucalyptus globulus): has long been used as an antiviral remedy for colds

and flu. It appears to work by boosting production of interferon, the body’s own antiviral

fighter, as well as, stimulating infection-fighting white blood cells. Echinacea has three

compounds, which are chicoric acid, caffeic acid, and echinacin. These three compounds

have specific antiviral properties that can resist viruses.

Forsythia (Forsythia suspensa) and Honeysuckle (Lonicera japonica) are the top choice in

the Chinese Materia Medica for addressing heat toxins. Often used together, these

flowers are the main ingredient in a formula called Yin Qiao San for treating viruses

causing upper respiratory infections.

Garlic (allium sativum): Compounds that are rich in sulfur content are found in

abundance in garlic and are active against the virus responsible for flu. These sulfur-

based compounds have shown to be effective in clinical studies. A similar but less

effective antiviral action is also found in the common onion, which is a close relative of

the garlic plant.

Ginger (zingiber officinale): Phenolic compounds are responsible for relaxing the

muscles of the stomach, and explains ginger‘s effect in easing motion sickness. Fresh or

dried, the root has been shown to minimize vomiting. In addition, the phenolic

ingredients act within the stomach as a sedative and painkiller, which helps to reduce

over-activity of the gut. In stomach infections, the oil acts as an antiseptic and an anti-


28

inflammatory. The gingerols alone are thought to be responsible for ginger‘s action as a

liver protective.

Goldenseal (hydrastis canadensis): Berberine is the active compound in goldenseal that

stimulates the immune system. douche of goldenseal is excellent for reducing yeast

infection. Berberine increases blood flow to the spleen and stimulate the activity of

macrophages, blood cells that are an important part of the immune system.

Licorice (glycyrrhiza glabra): also used for centuries and found in the Chinese Materia

Medica, this herb’s potent antiviral action works against a wide range of viral agents.

With eight active antiviral compounds, the most recognized being glycyrrhizin, these

compounds inhibit as well as block the viral penetration of the body’s cells and the

multiplication of genetic material from the viral particle.

Essential Oils with Antiviral Properties

Use these oils externally by blending a few drops into a base oil of vegetable, sunflower,

safflower or canola oils and massage into pulse points, chest, tops of feet and wrists. Do

not digest these oils internally. Tea tree can be added to a half a cup of warm water and

gargled then spit out for treating sore throats.

Eucalyptus (eucalyptus globulus): this oil has compounds which include quercitrin,

hyperoside, and tannic acid, which help eliminate viruses.

Juniper (juniperus): has an antiviral agent. Juniper contains a potent antiviral compound

called deoxypodophyllotoxi n. The herpes viruse, flu and many other types of viruses

seem to be inhibited by extracts from juniper .

Lemon Balm (melissa officinalis) : is a potent inhibitor on the herpes virus, among other

viruses that lead to infection.

Tea Tree Oil (melaleuca alternifolia) : used for everything from earache to athlete’s

foot, as well as gum inflammation and skin infections, this antiviral oil can be applied

directly to an infected area, three times daily. Tea tree can be added to a half a cup of

warm water and gargled then spit out for treating sore throats.


29

� Some important antiviral activity of plants

Antiviral drugs are a class of medication used specifically for treating viral infections.

Like antibiotics, specific antivirals are used for specific viruses. They are relatively

harmless to the host, and therefore can be used to treat infections. They should be

distinguished from viricides, which actively deactivate virus particles outside the body.

Most of the antiviral now available are designed to help deal with HIV; herpes viruses,

best known for causing cold sores and genital herpes, but actually causing a wide range

of diseases; the hepatitis B and C viruses, which can cause liver cancer, and influenza A

and B viruses. Researchers are now working to extend the range of antivirals to other

families of pathogens.

Antiviral drugs work by inhibiting the virus before it enters the cell, stopping it from

reproducing, or, in some cases, preventing it from exiting the cell. However, like

antibiotics, viruses may evolve to resist the antiviral drug.

VIRUS: AN INTRODUCTION

Virus are obligate intracellular organism, contain DNA or RNA within a cylindrical or

spherical protein coat or capsid, which may be surrounded by a lipid bilayer (envelope).

Viruses are organisms that can be characterized as having two distinct phases in their life

cycle - an intracellular and an extracellular phase.

Intracellular phase is the replicative phase during which the virus multiplies in the

infected cell. There it borrows the metabolic machinery of the cell to direct the synthesis

of proteins coded by the viral genome. The structural and nonstructural virion

components are synthesized independently, and the structural proteins are assembled into

whole virions during the final stage of reproduction. When virions leave the cell, they are

particles of uniform size, shape and chemical composition that in some cases can

crystallize. This is the extracellular phase of the virus. The viral particles can initiate the

infectious process of new cells, and hence they constitute the infectious form of the virus.


30

Virus life cycle

Viruses consist of a genome and sometimes a few enzymes stored in a capsule made of

protein (called a capsid), and sometimes covered with a lipid layer (sometimes called an

'envelope'). Viruses cannot reproduce on their own, and instead propagate by subjugating

a host cell to produce copies of themselves, thus producing the next generation.

Researchers working on such "rational drug design" strategies for developing antivirals

have tried to attack viruses at every stage of their life cycles. Some species of mushrooms

have been found to contain multiple antiviral chemicals with similar synergistic effects.[3]

Viral life cycles vary in their precise details depending on the species of virus, but they

all share a general pattern:

• Attachment to a host cell.

• Release of viral genes and possibly enzymes into the host cell.

• Replication of viral components using host-cell machinery.

• Assembly of viral components into complete viral particles.

• Release of viral particles to infect new host cells.

Limitations of vaccines

Vaccines bolster the body's immune system to better attack viruses in the "complete

particle" stage, outside of the organism's cells. They traditionally consist of an attenuated

(weakened or killed) version of the virus. These vaccines can, in rare cases, harm the host

by inadvertently infecting the host with a full-blown viral occupancy. Recently "subunit"

vaccines have been devised that consist strictly of protein targets from the pathogen.

They stimulate the immune system without doing serious harm to the host. In either case,

when the real pathogen attacks the subject, the immune system responds to it quickly and

blocks it.

Vaccines are very effective on stable viruses, but are of limited use in treating a patient

who has already been infected. They are also difficult to successfully deploy against


31

rapidly mutating viruses, such as influenza (the vaccine for which is updated every year)

and HIV. Antiviral drugs are particularly useful in these cases.

Anti-viral targeting

The general idea behind modern antiviral drug design is to identify viral proteins, or parts

of proteins, that can be disabled. These "targets" should generally be as unlike any

proteins or parts of proteins in humans as possible, to reduce the likelihood of side

effects. The targets should also be common across many strains of a virus, or even among

different species of virus in the same family, so a single drug will have broad

effectiveness. For example, a researcher might target a critical enzyme synthesized by the

virus, but not the patient, that is common across strains, and see what can be done to

interfere with its operation.

The target proteins can be manufactured in the lab for testing with candidate treatments

by inserting the gene that synthesizes the target protein into bacteria or other kinds of

cells. The cells are then cultured for mass production of the protein, which can then be

exposed to various treatment candidates and evaluated with "rapid screening"

technologies.

Approaches by life cycle stage

Before cell entry

One anti-viral strategy is to interfere with the ability of a virus to infiltrate a target cell.

The virus must go through a sequence of steps to do this, beginning with binding to a

specific "receptor" molecule on the surface of the host cell and ending with the virus

"uncoating" inside the cell and releasing its contents. Viruses that have a lipid envelope

must also fuse their envelope with the target cell, or with a vesicle that transports them

into the cell, before they can uncoat.


32

This stage of viral replication can be inhibited in two ways:

• Using agents which mimic the virus-associated protein (VAP) and bind to the

cellular receptors. This may include VAP anti-idiotypic antibodies, natural

ligands of the receptor and anti-receptor antibodies.

• Using agents which mimic the cellular receptor and bind to the VAP. This

includes anti-VAP antibodies, receptor anti-idiotypic antibodies, extraneous

receptor and synthetic receptor mimics.

This strategy of designing drugs can be very expensive, and since the process of

generating anti-idiotypic antibodies is partly trial and error, it can be a relatively slow

process until an adequate molecule is produced.

Entry inhibitor

A very early stage of viral infection is viral entry, when the virus attaches to and enters

the host cell. A number of "entry-inhibiting" or "entry-blocking" drugs are being

developed to fight HIV. HIV most heavily targets the immune system's white blood cells

known as "helper T cells", and identifies these target cells through T-cell surface

receptors designated "CD4" and "CCR5". Attempts to interfere with the binding of HIV

with the CD4 receptor have failed to stop HIV from infecting helper T cells, but research

continues on trying to interfere with the binding of HIV to the CCR5 receptor in hopes

that it will be more effective.

Uncoating inhibitor

Inhibitors of uncoating have also been investigated.Amantadine and rimantadine, have

been introduced to combat influenza. These agents act on penetration/uncoating.

Pleconaril works against rhinoviruses, which cause the common cold, by blocking a

pocket on the surface of the virus that controls the uncoating process. This pocket is

similar in most strains of rhinoviruses and enteroviruses, which can cause diarrhea,

meningitis, conjunctivitis, and encephalitis.

During viral synthesis

A second approach is to target the processes that synthesize virus components after a

virus invades a cell.


33

Reverse transcription

One way of doing this is to develop nucleotide or nucleoside analogues that look like the

building blocks of RNA or DNA, but deactivate the enzymes that synthesize the RNA or

DNA once the analogue is incorporated. This approach is more commonly associated

with the inhibition of reverse transcriptase (RNA to DNA) than with "normal"

transcriptase (DNA to RNA).The first successful antiviral, acyclovir, is a nucleoside

analogue, and is effective against herpesvirus infections. The first antiviral drug to be

approved for treating HIV, zidovudine (AZT), is also a nucleoside analogue.

An improved knowledge of the action of reverse transcriptase has led to better nucleoside

analogues to treat HIV infections. One of these drugs, lamivudine, has been approved to

treat hepatitis B, which uses reverse transcriptase as part of its replication process.

Researchers have gone further and developed inhibitors that do not look like nucleosides,

but can still block reverse transcriptase.Another target being considered for HIV

antivirals include RNase H - which is a component of reverse transcriptase that splits the

synthesized DNA from the original viral RNA .

Integrase

Another target is integrase, which splices the synthesized DNA into the host cell genome.

Transcription

Once a virus genome becomes operational in a host cell, it then generates messenger

RNA (mRNA) molecules that direct the synthesis of viral proteins. Production of mRNA

is initiated by proteins known as transcription factors. Several antivirals are now being

designed to block attachment of transcription factors to viral DNA.

Translation / antisense

Genomics has not only helped find targets for many antivirals, it has provided the basis

for an entirely new type of drug, based on "antisense" molecules. These are segments of

DNA or RNA that are designed as complementary molecule to critical sections of viral

genomes, and the binding of these antisense segments to these target sections blocks the

operation of those genomes. A phosphorothioate antisense drug named fomivirsen has


34

been introduced, used to treat opportunistic eye infections in AIDS patients caused by

cytomegalovirus, and other antisense antivirals are in development. An antisense

structural type that has proven especially valuable in research is morpholino antisense.

Morpholino oligos have been used to experimentally suppress many viral types:

• caliciviruses

• flaviviruses (including WNV)

• dengue

• HCV

• coronaviruses

Translation / ribozymes

Yet another antiviral technique inspired by genomics is a set of drugs based on

ribozymes, which are enzymes that will cut apart viral RNA or DNA at selected sites. In

their natural course, ribozymes are used as part of the viral manufacturing sequence, but

these synthetic ribozymes are designed to cut RNA and DNA at sites that will disable

them.

A ribozyme antiviral to deal with hepatitis C has been suggested, and ribozyme antivirals

are being developed to deal with HIV.[13] An interesting variation of this idea is the use of

genetically modified cells that can produce custom-tailored ribozymes. This is part of a

broader effort to create genetically modified cells that can be injected into a host to attack

pathogens by generating specialized proteins that block viral replication at various phases

of the viral life cycle.

Protease inhibitors

Some viruses include an enzyme known as a protease that cuts viral protein chains apart

so they can be assembled into their final configuration. HIV includes a protease, and so

considerable research has been performed to find "protease inhibitors" to attack HIV at

that phase of its life cycle.[14] Protease inhibitors became available in the 1990s and have


35

proven effective, though they can have unusual side effects, for example causing fat to

build up in unusual places. Improved protease inhibitors are now in development.

Protease inhibitors have also been seen in nature. A protease inhibitor was isolated from

the Shiitake mushroom (Lentinus edodes).The presence of this may explain the Shiitake

mushrooms noted antiviral activity in vitro.

Assembly

Rifampicin acts at the assembly phase.

Release phase

The final stage in the life cycle of a virus is the release of completed viruses from the host

cell, and this step has also been targeted by antiviral drug developers. Two drugs named

zanamivir (Relenza) and oseltamivir (Tamiflu) that have been recently introduced to treat

influenza prevent the release of viral particles by blocking a molecule named

neuraminidase that is found on the surface of flu viruses, and also seems to be constant

across a wide range of flu strains.

Immune system stimulation

A second category of tactics for fighting viruses involves encouraging the body's immune

system to attack them, rather than attacking them directly. Some antivirals of this sort do

not focus on a specific pathogen, instead stimulating the immune system to attack a range

of pathogens.One of the best-known of this class of drugs are interferons, which inhibit

viral synthesis in infected cells.[19] One form of human interferon named "interferon

alpha" is well-established as part of the standard treatment for hepatitis B and C,[20] and

other interferons are also being investigated as treatments for various diseases.

A more specific approach is to synthesize antibodies, protein molecules that can bind to a

pathogen and mark it for attack by other elements of the immune system. Once

researchers identify a particular target on the pathogen, they can synthesize quantities of

identical "monoclonal" antibodies to link up that target. A monoclonal drug is now being


36

sold to help fight respiratory syncytial virus in babie and antibodies purified from

infected individuals are also used as a treatment for hepatitis B.

AAAAidsidsidsids

The acquired immune deficiency syndromes (AIDS) is the state of profound immuno-

suppression produced by chronic infection with the human immune deficiency virus

(HIV). It is characterized by profound immunosuppression that leads to opportunistic

infections, secondary neoplasm and neurologic manifestations.

Plants Used in Treating HIV

While new drugs are constantly being produced and studied for the treatment of HIV, or

human immunodeficiency virus, the use of plants is not out of the question. Plants are

still being used in developing nations in the hopes of treating, and one day curing, HIV.

Herbs

1. Herbal medicinal remedies have been used for centuries to treat a variety of ailments in

various cultures. Aloe vera, St. John's Wort, echinacea and ginseng have been used to

boost the immune system. Ginkgo, garlic and astragalus also help revive a failing

immune system.

Licorice

2. Licorice, or deglycyrrhizinated licorice, has a soothing effect on the mouth and throat for

HIV and AIDS patients suffering from ulcers in these places.

Astragalus Root

3. A Chinese herb, the astragalus root has been used in the East to prevent and slow the

shortening of telomere in immune cell chromosomes. Since HIV attacks the body's

immune cells, the use of the root is now being turned toward this virus. Experts believe


37

the root can be added successfully to anti-retroviral therapies for HIV-positive persons or

perhaps replace them entirely for a lower cost.

Cat's Claw

4. A woody Peruvian vine used by the Ashanica Indians has been proven to reduce the side

effects of AZT therapy for HIV and AIDS patients. Known as cat's claw, it stimulates the

immune system's response to the disease.

South African Plant

5. Known as Sutherlandia frutescens, the South Africa native plant is being used as

an immune and energy booster for sufferers of HIV and AIDS. It is also an

effective anti-depressant.

Materials and methods

Plant material Seventeen plants (Table 3.2) which are used to treat, HIV- infections in

immunocompromised patients were collected from different areas in Mozambique

Preparation of plant extracts

Dried powdered plant materials wer e extracted with acetone .Fifty grams of powdered

plant material was extracted with 500 ml of solvent over two days under reflux.

The extracts were then filtered and concentrated to dryness under reduced pressure

and the residues freshly dissolved in an ap propriate solvent on the day that the bioassay

was done.

HIV-1 Reverse tra nscriptase (RT) assay The effect of plant extracts on RT activity in vitro was evaluated with a non-

radioactive HIV-RT colorimetric ELISA kit (Roche, Germany). The assay was carried out

in triplicate. Adriamycin, an anticancer drug and also an inhibitor of viral reverse


38

transcriptase (Goud etal ., 2003) was used as a positive control.In each test well,

2µl of diluted recombinant HIV-1 reverse transcriptase (4-6 n g), 20µl o f diluted

extract, and 20µl o freaction mixture was dispensed. The final concentration of each

extract in each well was 20 µg/ml. Since this part of the experiment was not

conducted at the University of Pretoria, but at Nelson Mandela Metropolitan

University; due to cost implications, only one concentration was selected. Negative

control wells contained 40µl of lysis buff er and 20µl of reaction mixture. The

concentration of positive drug control (Adriamycin) was 100µ g/ml. Positive control

wells contained 20µl diluted recombinant HIV-1 Reverse transcriptase (4-6 ng), 20µl of

lysis buffer containing 10% DMSO, and 20µl o f reaction mixture. The wells of the

micro titer plate modules were washed five times with 250µl of washing buffer per well

for 30 seconds each. The washing buffer was then carefully removed and 200 µ l of anti-

DIG-POD working solution was dispensed into each well. Incubation at 37oC followed

once again for 1 hour after the micro titer plate modules were covered with foil. The

wells were then washed in the same manner as before, the washing buffer was

carefully removed from the wells, and 200µl of ABTS substrate was dispensed into

the wells. Incubation then commenced for 10-30 min at room temperature (15-25 C). The

absorbencies of the samples were measured at 405nm( reference wavelength: 492

nm). The percentage inhibitory activity of the extracts samples were then calculated,

with reference to the positive control.

Herpes simplexHerpes simplexHerpes simplexHerpes simplex

Plant Extracts Antiviral Activity against Herpes simplex

PLANT EXTRACTS ANTIVIRAL ACTIVITY AGAINST HERPES SIMPLEX VIRUS

TYPE 1 AND AFRICAN SWINE FEVER VIRUS

ABSTRACT


39

Twenty-eight extracts prepared from plants used in African traditional medicine and from

Rhamnus glandulosa Ait. of the Portuguese flora, were screened in order to assay their

antiviral activity against Herpes simplex virus type 1 (HSV-1) and African swine fever

virus (ASFV). Twelve of these extracts revealed virucidal activity against HSV-1

whereas only six have the same activity against ASFV. Further studies showed that

thirteen of the tested extracts inhibited HSV-1 infection, some of which had a significant

activity against this virus such as Senna podocarpa (Guill. & Pert.) Lock, Cassia

sieberiana DC., Guiera senegalensis J.F. Gmel., Piliostigma thonningii (Schum.) Milne-

Redhead, Rhamnus glandulosa Ait. and Uvaria chamae P. Beauv. Four of the twenty-one

tested extracts inhibited ASFV infection

MATERIALS AND METHODS

Plant Materials

Plants were collected in Guinea-Bissau and identified by Dr. A. Diniz; voucher

herbarium specimens have been preserved in LISC Herbarium of the "Centro de Botânica

Tropical", Lisbon, Portugal; Rhamnus glandulosa leaves were collected by Mr. N¢brega

from a plant specimen of "Jardim Botânico do Funchal", Madeira Island, Portugal.

Extracts Preparation

Different parts of the dried plants were extracted with ethanol at room temperature and

concentrated under reduced pressure (temperature < 40øC). The dried extracts were

dissolved in dimethylsulfoxide (DMSO) at a concentration of 100 mg/ml.

Cells Used

Vero cells were obtained from the American Type Culture Collection and were grown as

monolayers in Dulbecco's Modified Eagle Medium (DME) (Gibco, Scotland)

supplemented with 10% of New-born Calf Serum (NCS) (Gibco, Scotland) and

gentamicin (50 æg/ml).

Viruses


40

African swine fever virus (ASFV), strain Lisbon 60, was originally obtained in 1960

from an infected pig in Lisbon (Ribeiro and Azevedo, 1961), adapted to grow in monkey

cells and was cloned by four successive plaque purification in Vero cells, as described by

Enjuanes et al. (1976). Herpes simplex virus type 1 (HSV-1, clinical strain) stocks were

prepared in Vero cells in DME supplemented with 2% NCS. Only the extracellular

viruses were used, after low speed centrifugation of cell debris. When necessary, vires

suspensions were concentrated by centrifugation at 11000 G for 6 h at 4øC. Plaque

titrations were done as described by Enjuanes et al. (1976).

Antiviral Assays

Cytotoxicity assays -- As a first step, the cytotoxicity of the extracts was evaluated in

order to determine their maximum tolerated concentration (MTC) to Vero cells. Vero

Cell monolayers in 96-well plates (Nunc, England) were incubated with decreasing

concentrations of the extracts (dissolved in DMSO) for 48 h at 37øC. After 48 h, a dye

uptake assay (Finter, 1969) was performed to determine the percentage of living cells in

relation to the control (cells incubated with DMSO).

Virucidal activity of plant extracts -- The direct effect of plant extracts on HSV-1 and

ASFV was determined by incubating both viruses for 1 h at 37øC with the extracts at the

MTC. The extracts were removed by centrifugation (53,000 G or 100,000 G) and

virucidal activity was determined by plaque titration. Controls were performed by

incubating the viruses with Phosphate Buffered Saline-A (PBS-A) solution (Dulbecco

and Vogt, 1954).

Effect of plant extracts on viral replication -- Vero cell monolayers were incubated for 30

min at 37øC with the extracts at MTC. The extracts were removed and the cells washed

with PBS-A or DME2 (DME supplemented with 2% NCS) at the same temperature. Cell

monolayers were infected with HSV-1 or ASFV at a multiplicity of infection (m.o.i.) of 1

in the presence of the extracts. After adsorption of the viruses for 2 h at 37øC, the

remaining viruses were removed and cells were again washed with PBS-A. The plates

were incubated at 37øC until an extensive cytopathic effect was observed in control


41

(without extract) wells. The media from all infected wells were recovered and titrated for

virus.

Effect of plant extracts on viral adsorption -- Vero cell monolayers were pre-cooled at

4øC for 15 min after which they were infected with HSV-1 in presence of the extracts.

After viral adsorption for 2 h at 4øC, the remaining virus was removed and cells washed

with PBS-A. They were then incubated for a further 24 h at 37øC with DME2 and later

the progeny virus was plaque titrated.

All the experiments were effectuated in duplicate.

Chikungunya Treatment in PlantsChikungunya Treatment in PlantsChikungunya Treatment in PlantsChikungunya Treatment in Plants

Ayurveda is the traditional indian medicine and is a popular alternative medicine in India.

Ayurveda medicines are usually vegetable drugs and hence side effects usually are

minimal. But patients have to be very careful since there are a large number of unlicensed

ayurveda practitioners and some of them are known to add steroids in medicines for

quick benefit.

In Ayurveda Chikungunya is known as Sandhijwara which literally means "fever of the

joints". The symptoms of Sandhijwara and Chikungunya are very similar and hence

Ayurveda treatment provides relief for the disease. The medicines usually prescribed for

fighting the fever are Vilvadi Gulika, Sudarsanam Gulika and Amritarishta. Some

practitioners claims that Chikungunya can be cured faster with Ayurveda treatment. Since

there is no treatment in modern medicine, people consult Ayurveda practitioners for fever

relief in many parts of India. In ayurveda, Chikungunya is considered as a ' vata doksha'

disorder. Diet changes usually prescribed include increased intake of fluid food,

increased intake of vegetables, reduced use of oil and avoiding intake of tea and coffee.

Some practitioners of Ayurveda claim that a medicine called Panchathikta Kashayam can

cure Chikungunya. It is a very bitter drug. Ayurveda massage is also administered for

joint pain relief.


42

Herbs such as Sacred Basil (Tulsi), Carrot and Grapes are usually recommended as a

relief for the pain and fever. Since these are natural herbs there is little risk in trying them

out.

Natural plantNatural plantNatural plantNatural plants s s s ffffor Chicken Poxor Chicken Poxor Chicken Poxor Chicken Pox

Chicken pox is a highly contagious disease affecting mostly children and sometimes

adults also. The rash appears on the body 2 days after the person gets infected. It mainly

occurs during winter and spring season. The duration of this disease ranges from ten to

twenty-one days but is usually between fourteen and seventeen days.

Symptoms of Chicken Pox

Following are the major chicken pox symptoms:

1. Low grade fever

2. Mild headache and weakness

3. Rash appears on the upper chest or back

Causes of Chicken Pox

Following are the major chicken pox causes:

1. Virus infection

2. Wrong feeding of children

Chicken Pox Treatment and Advice

1. Take Raw fruit, vegetable juices and lemon juice

2. Keep the patient in a well-ventilated room

3. Lukewarm water baths can be given every day to relieve itching. For better results,

neem leaves can be added to this water.


43

Herbal Remedies for Chicken Pox

1. Prepare herbal tea by any of the herbs like chamomile (babunah), basil (tulsi),

marigold (zergul) and lemon balm (billilotan). A little cinnamon (dalchini), honey, and

lemon may be added to this tea. It should be sipped slowly several times a day. It is an

effective herbal remedy for chicken pox.


44

Antiviral activity

Of

Medicinal plants

Of

Nilgiris


45

There is an increasing need for search of new compounds with antiviral activityas the treatment

of viral infections with the available antiviral drugs is often unsatisfactory due to the problem of

viral resistance2 coupled with the problem of viral latency and conflicting efficacy in recurrent

infection in immunocompromised patients. Ethnopharmacology provides an alternative approach

for the discovery of antiviral agents, namely the study of medicinal plantswith a history of

traditional use as a potential source of substances with significant pharmacological and biological

activities4. The Indian subcontinent is endowed with rich and diverse local health tradition, which

is equally matched with rich and diverse plant genetic source5. A detailed investigation and

documentation of plants used in local health traditions and ethnopharmacological evaluation to

verify their efficacy and safety can lead to the development of invaluable herbal drugs or isolation

of compounds of therapeutic value. A number of compounds extracted from various species of

higher plants have shown antiviral activity; Examples included tannins7, flavones8, alkaloids, that

displayed in vitro activity against numerous viruses. It has been suggested that selection of plant

on the basis of ethnomedical considerations gives a higher hit rate than screening programmes of

general synthetic products10. Bacopa monneri has been used in conditions like epilepsy, insanity,

nervous disorders11, Hypercicum hookerianum in anxiety and inflammation11, Usnea complanta

and Tagetes minuta for bacterial infections, Santolina chamaecyparissus as a stimulant,

vermifuge and a stomachic.A number of plant extracts reported in traditional medicine to have

antiinfective properties were studied in our laboratory15-19 and were also screened for antiviral

activity. Herpes simplex viruses (HSV) are ubiquitous agents which cause a variety of diseases

ranging in severity from mild to severe, and in certain cases, these may even become life

threatenings, especially in immunocompromised patients. After primary infection, HSV persists

in the host for the lifetime. HSV infection is thus considered lifelong infection. Nucleoside

analogues such as aciclovir (ACV), penciclovir etc., are the only approved drugs for the treatment

of HSV infections. However, the widespread use of nucleoside based drugs has led to the

emergence of resistance in HSV especially among immunocompromised patients. In a recent

survey from Taiwan, the incidence of ACVresistant HSV strains was found to be around 5 per

cent among immunocompromised patients and 14 per cent among bone marrow transplant

recipients. This indicates the need for search of newer antiviral agents to treat such infections.

The present study was undertaken to test the extracts of 18 plants for their antiviral activity

against herpes simplex virus type I (HSV-1, a DNA virus).


46

Material & Methods Plant materials, reagents, cell line and virus: The plant materials were collected from in and

around Ootacamund, Tamil Nadu, India and were authenticated by the Botanical Survey of India,

Government Arts College, Ootacamund where sample specimens were deposited. Extracts of

different plants were prepared by using Soxhlet extraction unit (Borosil, Mumbai) as per the

standard procedure21. The essential oils from different parts of plants were isolated by water

distillation using Clavenges apparatus (Borosil, Mumbai)22. Eagle's minimum essential medium

(EMEM), trypsin, penicillin, streptomycin and amphotericin B were purchased from Hi-media

Labs, Mumbai, India. 3-(4, 5- dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and

trypan blue dye were purchased from Sigma, USA. New born calf serum (NBCS) was procured

from PAA Labs, Austria. Vero cells (African green monkey kidney cell) were obtained from

Pasteur Institute of India, Coonoor. Vero cells were grown in EMEM supplemented with Earle's

salts and 10 per cent heat inactivated NBCS, 100 IU/ml penicillin, 100µg/ml streptomycin and

5µg/ml amphotericin B. The cells were maintained at 37ºC in a humidified atmosphere with 5 per

cent CO2 and were subcultured twice a week. HSV-1 was from the collection of the Christian

Medical College and Hospital, Vellore. The virus was propagated in Vero cells and the infective

titre of the stock solution was 10-7 TCID50/ml (50% tissue culture infective dose).

Cytotoxicity assay: Each extract was separately dissolved in 1 ml of distilled dimethyl sulphoxide

(DMSO) and volume was made up to 10 ml with maintenance medium to obtain a stock solution

of 1 mg/ml concentration, sterilized by filtration and further dilutions were made from the stock.

The cytotoxicity assays were carried out using 0.1ml of cell suspension, containing 10,000 cells

seeded in each well of a 96-well microtitre plate (Tarsons India Pvt. Ltd., Kolkata). Fresh

medium containing different concentrations of the test sample was added after 24 h of seeding.

Control cells were incubated without test sample and with DMSO. The little percentage of

DMSO present in the wells (maximal 0.2%)

was found not to affect the experiment. The microtitre plates were incubated at 37ºC in a

humidified incubator with 5 per cent CO2 for a period of 72 h. Sixteen wells were used for each

concentration of the test sample. The morphology of the cells was inspected daily and observed

for microscopically detectable alterations, i.e., loss of monolayer, granulation and vacuolization

in the cytoplasm. The cytopathogenic effect (CPE) was scored. The 50 per cent cytotoxic

concentration (CTC50), was determined by the standard MTT assay23,24, trypan blue dye exclusion

method25, cell metabolic function by protein estimation26, and total cellular DNA content by H

thymidine labeling.


47

Antiviral assay: Different nontoxic concentrations of test drugs, i.e., lower than CTC50 were

checked for antiviral property by cytopathic effect (CPE) inhibition assay28 and virus yield

reduction assay29 against different virus challenge doses of 2, 10 and 100 TCID50. In CPE

inhibition assay, cells were seeded in a 96-well microtitre plate with 10,000 cells per well,

incubated at 37ºC in a humidified incubator with 5 per cent CO2 for a period of 48 h. The plates

were washed with fresh MEM and challenged with different virus challenge doses and incubated

at 37ºC for 90 min for adsorption of the virus. The cultures were treated with different dilutions

of plant extracts in fresh maintenance medium and incubated at 37ºC for five days. Every 24 h the

observation was made and cytopathic effects were recorded. Anti-HSV-1 activity was determined

by the inhibition of cytopathic effect compared with control, i.e., the protection offered by the test

samples to the cells was scored. In virus yield assay, reduction in the yield of virus when cells

were treated with the plant extracts was determined.


48

Antiviral Activity

of

Brazilian Plant


49

Unlike antimicrobial drugs against bacteria and fungi, only a few effective antiviral drugs

are available. One of the most important reasons for the lack of success in developing

antiviral drugs is due to the nature of the infectious viral agents, which totally depend

upon the cell they infect for their multiplication and survival, so that many compounds

that may cause the death of viruses also are very likely to injure the host cell that harbour

them .

On of the possible method which can be used for the discovery of active substances is the

screening of plant extracts for antiviral activity followed by bioassay guided fractionation

of active extracts to identify the active substance.

In searching for natural products as potential antiviral agents, several medicinal plants

from the South American folk medicine showed in vitro antiviral activity 2-4. In an effort

to discover new compounds, with relevant anti-herpes virus activity, the aim of our study

was to examine the potential antiviral activity of native species from South Brazil. The

species were selected mainly on the,basis of their ethnopharmacological indications for

treatment of viral infections such as influenza virus, bronchopulmonary infections and

cold, but some others species were collected randomly. In this study, hydroethanolic and

aqueous crude extracts from fifty species were screened for in vitro antiviral activity

against herpes simplex virus type I (HSV-1) in Vero cell line.

MATERIALS AND METHODS

Plant material

The plants tested were collected from the Rio Grande do Sul State area, Brazil.

Herbarium specimens are deposited in the Herbarium of the Federal University of Rio

Grande do Sul, Brazil (ICN). The air-dried plant materials were

grounded and extracted by maceration in 50% ethanol. An aqueous extract was also

prepared with dried plant material (100 g) in hot distilled water not exceeding 60 °C (200

ml). The extracts were filtered, the ethanol removed, lyophilized and stored at –20 °C

until tested.

Cells and viruses

African green monkey kidney cells (Vero cellline ATCC CCL-81) were grown in Eagle’s

minimum essential medium (MEM) supplemented with 10 % newborn calf serum, 2 µg.

ml–1 of amphotericin B and 10 mg.ml–1 of enrofloxacin. A virus stock of herpes simplex


50

virus type I, strain KOS (University of Rennes/France), ATCC-VR733 and 29R-acyclovir

resistant were prepared on Vero cells infected at a low multiplicity of infection, incubated

for 1 to 2 days, frozen/thawed three times, before clearing the preparation by

centrifugation at low speed to remove the cell debris. Virus stocks were maintained in

liquid nitrogen until use. Virus titration was performed by Kärber method 5 using a 96-

well microtitre plates. The virus titre was estimated from cytopathogenicity and

expressed as

50% tissue culture infectious doses (TCID50.ml–1). It was 10–6 TCID50.ml–1 for strain

KOS; 10–4 TCID50.ml–1 for strain ATCC and 10–4 TCID50.ml–1 for strain 29R-

acyclovir resistant.

Evaluation of cytotoxicity

To assess the effect of extracts on uninfected Vero cells, dilutions ranging from 40

mg.ml–1 to 0.15 µg.ml–1 in the MEM medium, were added to Vero monolayer’s (using a

96-well micro plate with 4.0 x 104 cells per well). After 72 h of incubation at 37 °C,

cytotoxicity was determined by microscopic examination of cell morphology in treated

and untreated cultures. The concentration of the extract at which the cell number was

reduced to 50% of the controls, was taken as the 50% cytotoxic concentration (CC50).

The maximum concentration at which no reduction on cell numbers was observed

(compared to controls) was considered as the maximum tolerated concentration (MTC) 6-

9. The MTC was determined for each extract before proceeding to make antiviral activity

assays. All assays were

carried out in quadruplicate.

Antiviral activity

Dilutions of the extracts were prepared starting from the previously determined MTC.

Extracts from MTC, MTC/2, MTC/4, MTC/8 and MTC/16 were added on confluent 24 h

old monolayers of Vero cells grown in microtitre tissue

culture plates just before virus inoculation. One hundred tissue culture infection doses per

50 ml (TCID50) of the HSV-1 KOS strain were added to each of the wells. Toxicity

controls, cell and virus controls titration were rum simultaneously. All assays were

carried out in quadruplicate. When some antiviral activity was detected

on HSV-1 strain KOS, the extracts were also tested against strain ATCC-VR733 and 29

Racyclovir resistant. Plates were incubated for 72 h at 37 °C, and then examined for the


51

presence of cytopathic effects (CPE). The concentration of extract which inhibited 100%

of the viral CPE (compared to the controls) was considered as the active concentration.

Discussion


52

Conclusion


53

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