Discovery of COVID-19 Vaccine by Using Acaciades as a Phytomedicine Improving Science and Technology Communication Applications- An Ideas
Subhas Chandra Datta*
Headmaster, Secretary and Researcher, Kanchannagar, Burdwan Municipality, India
Received Date: 07/06/2020; Published Date: 24/06/2020
*Corresponding author: Subhas Chandra Datta, PhD, Headmaster of Kanchannagar D.N.Das High School (HS), C/O- Rajendra Nath Nag, House No.-430A, Bajeprotappur (Katwa Road), Opposite to entry of SBI Bajeprotappur, P.O.- Bajeprotappur, Burdwan Municipality, 24 no. Ward, Bardhaman, Purba Bardhaman, Burdwan- 713101, West Bengal, India
Cite this article: Subhas Chandra Datta, Discovery of COVID-19 Vaccine by Using Acaciades as a Phytomedicine Improving Science and Technology Communication Applications- An Ideas. Op Acc J Bio Sci & Res 1(5)-2020.
The spread of novel pandemic coronavirus COVID-19, is now likely to become puzzle for the leaping from animals to human of fifth endemic coronavirus, may collapse or change human civilizations, and the world economy as well as communities. Though needs, but lockdown badly affects our economic growth down to 2.5 % in India and 2.2 % of world. In current situations, no antiviral drugs are to treat COVID-19 coronaviruses. To move forward, it will require new and more efficient solutions, science and technology applications, products, and it has to fulfill all requirements. Present pretreatment- investigations have revealed that Acaciasides (A&B) prepared from the funicles of Acacia uriculiformis A. Cunn., may use as potential phytmedicine against various pathogens like root-knot-, leaf spot-, powdery mildew-, mosaic- and tukra- diseases of mulberry by inducing their natural defense response. And it is reported that the saponins of Acaciaside is being used traditionally to overcome various medical complications like sore eyes, aches, rheumatism, allergy, itching, and rashes etc. And it has also been reported that many pharmacological activities like central nervous system depressant activity, antioxidant, antimicrobial, antimalarial, antifilarial, anticestocidal, antimutagenic, antihelmintic, chemopreventive, spermicidal, wound healing, hepatoprotective and antidiabetic activity due to its low toxicity and high efficacy at very low dose. Recently there is no effective treatment against COVID-19. So, the effect of the virus is likely to be seen long after medical science offers a cure or at least a vaccine for COVID-19. To overcome the situation, two ideas for discovery of ‘Vaccine’ or ‘Treatment’ by using biomedicines;-Acaciasides as a Phytomedicine, which may kill the COVID-19 by boosting our immune system, and the whole world may retain in normal forms by fighting against this war.
Keywords: Novel Coronavirus COVID-19; Acaciades, Phytomedicine; Ideas; Vaccine; Science and Technologies Applications
On New Year eve 2019, a cluster of cases of pneumonia of unknown cause detected in Wuhan City, Hubei Province of China, was informed by WHO. The coronavirus disease (COVID-2019) was identified as the causative virus by Chinese authorities on 7 January . Our normal life is on hold due to escalating novel coronavirus COVID-19 emergency. The ongoing spread of the COVID-19 pandemic is having profound effects on our private and professional life, the world economy, and the social organization of the communities we live in . The recent emergence of this virus, which is causing an outbreak of unusual viral pneumonia in patients and different animals which will help to understand the biology and potential risk of coronavirus that exist in richness in wildlife such as bats . The novel coronavirus SARS-CoV-2 that causes COVID-19 is now likely to become the fifth endemic coronavirus in human population. Recently, scientists are working to decipher its genome to help us stop other coronaviruses entering the human population.
They don’t know “the procedure of transfer of the novel coronavirus which causes COVID-19 made the leap from animals to humans” is a puzzle and scientists are trying to solve as humanity comes to grip with the deadly pandemic sweeping the globe and it is concluded that "corona viruses clearly have the capacity to jump species boundaries and adapt to new hosts, making it straightforward to predict for the more emergence in the future." How we respond to that will require more research to assist develop public health policy. They point to policy and other measures to help prevent other coronaviruses becoming a health danger to humans . It is reported that wildlife contains many coronaviruses which could potentially emerge in humans in the future. A very important lesson from this pandemic to help prevent the next one is that humans must not exposed them to wildlife, for example by banning 'wet markets' and the trade in wildlife and the pangolins play a vital role in the emergence of SARS-CoV-2 which causes the COVID-19, is still unknown.
It is also striking is that the pangolin viruses contain some genomic regions that are very closely related to the human virus and the receptor binding domain is the most important which dictates how the virus is able to attach and infect human cells "- said by Professor Holmes . A new coronavirus (CoV) identified as COVID-19 virus is the etiological agent responsible for the 2019-2020 viral pneumonia outbreak that commenced in Wuhan [6-9]. For this, WHO responses to the outbreak, the Research and Development Blueprint has been activated to accelerate diagnostics, vaccines and therapeutics for this novel coronavirus .
Every time when turn on television news, some expert is saying “There is currently no effective treatment for coronavirus.” Currently there are no targeted therapeutics and effective treatment options remain very limited which plays a pivotal role in mediating viral replication and transcription, making it an attractive drug target for this virus [10-11]. It is reported in a chapter entitled “Genetic Resistance to Coronavirus Infection- A Review” . In another study, it is reported that 2019-nCoV is sufficiently divergent from SARS-CoV to be considered a new human-infecting betacoronavirus and our phylogenetic analysis suggests that bats might be the main host of this virus, an animal which is used at the seafood market in Wuhan might represent an intermediate host facilitating the emergence of the virus in humans. Specially, structural analysis suggests that 2019-nCoV might be able to bind to the angiotensin converting enzyme 2 receptor in humans. The future evolution, adaptation, and spread of this virus warrant urgent investigation .
Recently, in ‘India Today’ discuss regarding the “Trade in the time of COVID-19: The economic impact of coronavirus on India and beyond” shows the estimates of India’s aggressive 21-day lockdown could bring the country’s growth down to 2.5 % from the 4.5 %. The effect of coronavirus is likely to be seen long after medical science offers a cure or at least a vaccine [14-15]. In a joint report from the World Health Organization (WHO) and the World Bank estimates the impact of such a pandemic at 2.2 per cent to 4.8 per cent of global GDP (US$3 trillion) [14-15]. That was well before the world knew of Covid-19. So, The effect of the virus is likely to be seen long after medical science offers a cure or at least a vaccine for Covid-19 (the disease caused by the new coronavirus). That’s because the economic cost of shutting down large swathes of the world is going to be steep [14-15].
It is reported that the novel coronavirus infection became a pandemic because it thrives on asymptomatic patients of Covid-19 and different health agencies world over have been firm in dealing with symptomatic Covid-19 patients. It will be faced when the real challenge has been to identify those Covid-19 patients who don't show any symptoms which has forced affected countries to revise their death toll because novel coronavirus is still an unknown enemy to humans. It may be possible for the symptoms which are currently atypical for Covid-19 may evolve into key indicators of novel coronavirus infection in months and years to come because, as the World Health Organization warns, this virus is going to stay among humans for a long time . In theory, challenge trials could enroll volunteers who are at low risk of harm from the virus that causes COVID-19-dubbed SARS-CoV-2—such as young adults who rarely develop serious symptoms after becoming infected naturally. “Now this pandemic situation is analogous to war, in which there is a long tradition of volunteers risking their health and lives on dangerous missions for which they know about the risks and are willing to do so in order to help save the lives of others,” the letter states. “The delay of every week in the deployment of a vaccine to the seven billion humans on earth will cost thousands of lives.” Foster counters that testing a vaccine in health care workers who are needed in a hot spot also has risks, as it might take them out of work for a time. He hopes the letter “just makes the FDA a little less scared,” stressing that “it’s pretty clear that the only way we’re really finally going to get out of this [pandemic] is when we have an effective vaccine” . Antimalarial drags widely used against COVID-19 heighten risk of cardiac arrest. How can doctors minimize the danger? .
In the Drug Target Review’s hub for COVID-19 research hub, reported that no vaccine currently available for this coronavirus which has spread swiftly across the globe, resulting in devastating effects to the world’s economy and an ever-increasing number of fatalities. To support collaboration in this field, they have gathered the latest news and updates relating to COVID-19 drug discovery efforts . More than 100 treatments and vaccines are in development to stem the COVID-19 pandemic, and some onlookers have worried that this sprawling and potentially duplicative effort is wasting time and resources. Hoping to bring order to the chaos, the National Institutes of Health (NIH) and major drug companies today announced a plan to stage carefully designed clinical trials of the drugs and vaccines they have decided are the highest priorities for testing and development . On 30 January 2020, Global research and innovation forum: towards a research roadmap; following the recommendations of the Emergency Committee, the Director-General of WHO declared that the outbreak constitutes a Public Health Emergency of International Concern (PHEIC). The world scientists on COVID-19 met at the WHO’s Geneva headquarters from 11 to 12 February 2020 to assess the current level of knowledge regarding the new virus, agree on critical research questions that need to be answered immediately, and ways to work together to accelerate and fund priority research that can contribute to curtail this outbreak and prepare for the future outbreaks .
To move forward, it will require new and more efficient solutions, science and technology applications, products, and it has to fulfill all requirements. To meet the challenge of the problems; it is already reported that the Biomedicine- Acaciasides (A&B) is being used traditionally to overcome various medical complications. It has been used to treat several medical ailments due to its low toxicity and high efficacy and presence of effective bioactive phytoconstituents of Biomedicine- Acaciasides (A&B) [21,22]. It is already reported from previous experiments that Acaciasides or Acacia auriculiformis-extract or Aakashmoni, prepared from the funicles (fruits) of Acacia auriculiformis A. Cunn, is highly effective in ameliorating different diseases like root-knot, leaf spot, powdery mildew, mosaic and tukra diseases in different plants and root callous by using their defense-response against pathogen infection and leaving no residual toxicity in the leaves to affect the growth [22-46]. Recently published a paper entitled “Enriched Science and Technology Communication Economy in Agriculture by Use of Acaciasides as Potential Bio-Agents Against Various Pathogens” [35,47].
Phytomedicine (any biological medicines- or agents- of biological origin derived from plants source) provide a new class of biological compounds which stand as a suitable and useful alternative to conventional but hazardous methods of chemical control against pathogens [22,25,35]. Our main aim was to investigate new and more efficient solutions, technologies and products for controlling disease, by using Biomedicine- Acaciasides (A&B) as Phytomedicine with Pretreatment, which directly influence the defense mechanism for preventing diseases and resource productive economies applications. The main purpose of the present investigation is to see the efficacy of Pretreatment effects with the Acaciasides (A&B) using as Phytomedicine at very low dose, prepared or isolated from the funicles of Acacia uriculiformis A. Cunn., against various pathogens like root-knot-, leaf spot-, powdery mildew-, mosaic- and tukra- diseases of mulberry (Morus alba L., cv. S1) by inducing their natural defense response.
Recently there are no targeted therapeutics and effective treatment options remain very limited. So, the effect of the virus is likely to be seen long after medical science offers a cure or at least a vaccine for COVID-19. Now it is planned to publish as a proposal for current outcomes and Therapies on Coronavirus disease (COVID-19) outbreak that helps the readers as well as a scientific community to take measures or treatment opportunity or discover vaccine to avoid new coronavirus. Our main goal is to limit infections. To overcome the situation, a proposal for clinical study may be arranged in near future or as early as possible by the researcher for discovery of ‘Vaccine’ or at least treatment by using Biomedicine- Acaciasides as a Phytomedicine, which may kill the COVID-19 by boosting our immune system which may significantly solve or help or advance the current problem of the COVID-19 infections and clinical practice and acts as indispensable source to access the pharmacological developments (Figure 1).
Material and Methods
Preparation of crude Acacia auriculiformis extract
Air-dried and powdered funicles or fruits of Acacia auriculiformis A. Cunn. (Figure 2) were extracted with 90% ethanol at room temperature (25 + 2 °C) for 15 days and were filtered for collecting extract. Later, the ethanol from the extract was removed by evaporation at room temperature (25 + 2 °C). The residue, obtained after removal of the solvent under reduced pressure, was dried in a desicator over anhydrous calcium chloride [22,23,32,35,46-48].
Isolation of Acaciasides
Crude residue of Acacia auriculiformis- extract from the funicles of Acacia uriculiformis A. Cunn. Were again successively extracted with petrol (60-80°C) and 90% ethanol. The ethanol extract, on removal of the solvent under reduced pressure, yielded a viscous dark brown mass. The extract was chromatographed on silica gel with petrol, petrol-chloroform (1:1), chloroform, chloroform-methanol (9:1, 7:3, 3:2, 1:1 and 2:3) as successive eluents. The chloroform-methanol (7:3 and 3:2) eluates were then combined fraction was found to be composed mainly of two compounds which were separated by repeated preparative HPLC employing Spherisorb S-10-ODS reversed phase column with the solvent system methanol-water (7:3) at a flow rate of 4 ml / min and refractive index detector as white amorphous solids. These two solid compounds, designated as acaciaside A and acaciaside B (Figure 1) according to their increasing order of polarity, were found to be triterpenoid saponins by Liebermann-Burchard, molisch and forth tests [22,23,32,35,46-48].
Ten sets of cavity block with 1ml sterile tap water containing 50 larvae (J2) of Meloidogyne incognita were taken; five set was treated as control and other five were treated as treatment set. The Acaciasides (A&B) were dissolved in sterile tap water at 0.0125 mg / ml forming acaciasides-test solution. To assess the direct effect of acaciasides- test solution, the water was removed by pipette and in all the treatment sets, immediately replaced by 1ml of test solutions (0.0125 mg acaciasides / ml concentration) were added, except the control and observed with every one hour interval for a period of 24 hours exposure period at room temperature (25±2°C). Immediately after observation of each block, nematodes were transferred to sterile tap water again to see if any recovery occurred after 4 hours [22,25-29,30,31,35-37,40-44,49].
Preparation of Acaciasides- Pretreatment test solution
In our experiment, the mixture of acaciaside A and acaciaside B (3:2) was dissolved in sterile tap water at 0.0125 mg / ml forming acaciasides-Pretreatment test solution and used for Pretreatment plots [25,26,32,35,46].
Site of the Experimental Plots
The field experiment was carried out at the Sriniketan Sericultural Composite Unit, Government of West Bengal, India where temperature was 28 + 5°C and relative humidity was 75 + 5 % (Figure 2) [11-12]. Throughout year, the whole mulberry field was naturally infected with root-knot disease of mulberry plants (Morus alba L., cv. S1) caused by Meloidogyne incognita (Kofoid & White) Chitwood root-knot nematodes pathogens. And every year in September-October occurrence of four foliar diseases were seen and these were: leaf spot disease caused by Cercosporam moricola (Cooke) fungus pathogens, powdery mildew disease caused by Phyllactinia corylea (Pers.) Karst fungus pathogens, mosaic disease caused by mosaic virus pathogens and tukra disease caused by Maconellicoccus hirsutus (Green) mealy bug pathogens [22,25,35].
Figure 2: Funicles of Acacia auriculiformis.
Estimation of the nematode pathogen population
Soil and root samples [22,25,50,51] were taken at random from a sericulture field spreading over an area of 5.6 acre of land with a view to determining the extent and intensity of Meloidogyne incognita (Kofoid & White) Chitwood nematode pathogen infestation. Later, two areas (in the same locality and climatic condition) each measuring 0.02 ha; one naturally root-knot disease infected Untreated field and other naturally root-knot disease infected Pretreated field, were demarcated in the mulberry field where there were no soil differences as well as environmental factor.
Preparation of fields
The first 0.02 ha nematode infected (2863 + 55 J2 /1 kg of soil) sandy soil area (18889.76 x 1066.80 x 45.72 cm3) was mixed with yard manure (2:1 vol / vol). Every day, at least 40 random sampling of moist rhizospheric soil (200g of soil i.e., each sample collected by making a hole of 1.8 cm wide and 6 cm deep) were done in the nematode infected area for 30 days and were assessed the M. incognita population [43,44,22,24,25,27-28,32,36,50,51-56] and this naturally infected soil-filled area, demarking Untreated Field, was replicated thrice. The other 0.02 ha (18889.76 x 1066.80 x 45.72 cm3) naturally M. incognita infected sandy soil field was also prepared by mixing yard manure (2 : 1 vol / vol), removing weeds, irrigating water and interchanging among the soil for uniform distribution of manure and nematodes in the naturally infected field which was estimated by regular soil sampling like a same process of previous one. This naturally infected soil-filled area, demarking Pretreated field, was also replicated thrice [25,32,35].
Plantation of mulberry cutting
Mature three years old mulberry cutting, Morus alba L., cv. S1 (average 25cm length and 20g fresh weight) collected from same sericulture field, were planted with a gap of 45cm throughout the experimental fields where there were no soil difference and climatic conditions. The planted mulberry cuttings were allowed to grow for a period of three months. Regular rhizospheric soil and root sampling (at random) were done for estimation of nematode population during this three month growth period of mulberry in all fields [25-29,32,35,50,51]. At least 80 number at random rhizospheric soil sampling (200g in each sample) were collected from rhizospheric root-soil area of root (10-15cm X 10-15cm) and at least 40 number at random root sampling (2g fresh root in each sample) were collected from newly formed roots (or gall roots) for determining the intensity or presence of nematodes in all the Pretreatment experimental fields.
Division of groups and plots
After three months growth of mulberry, M. incognita population were estimated in the rhizospheric soil as well as roots [25-29,32,35,50,51] (at least 40 at random sampling in each area) of mulberry plants in each areas of mulberry field. The M. incognita infected mulberry plants were achieved growth of 50-60 cm in height. The infected mulberry plants were divided in to 16 plots (Figure 3), each measuring the area of 472.44cm X 533.4cm X 45.72cm. The mulberry plants divided into two plant groups; Untreated Plants Groups and Pretreated Plants Groups and each group has 8-plots (20 plants / plot). At first all the plants were pruned, manured with NPK and irrigated every 7 days. Rhizospheric soil was interchanged among the plants to keep the nematode infestation as uniform as possible in the naturally infected field. After pruning, the plants were allowed to grow for a period of 137 days when their root-knot, leaf spot, powdery mildew, viral and tukra diseases were assessed [32,33,38,39,56,57]. The field trial was replicated three times.
Figure 3: Field grown with mulberry plants naturally infected with M. incognita.
Pretreatment with Phytomedicine- Acaciasides (A&B)
Seventy six days after pruning of mulberry plants, all the plots (Pretreated groups and Untreated groups) were done by foliar spray and soil drench @ 20ml / plant in each type of Pretreatment (0.0125 mg Acaciasides / ml concentration in case of Pretreated groups) twice at an interval of 15 days with Acaciasides- pretreatment test solution and sterile tap- water respectively before the onset of diseases. Two pretreatments were given in such a way that all the leaves and rhizospheric soil of the plants were completely drenched with Pretreatment test-solutions and tap-water. During spraying, the soil surface underneath each plant was covered with polyethylene sheet. All the Acaciasides- pretreated groups were received 80 ml / plant test solutions (1 mg / plant) and other Untreated- plant groups were similarly received 80 ml / plant sterile tap water respectively [25-26,32,35,46]. It was told about untreated (control); these controls were only pretreated with the sterile tap water (i.e. without Acaciasides- pretreatment test solution). At thirty days after the second Pretreatment all the parameters of diseases were assessed again for each group [22,25-27,30-32,35,36,46]. All the data were used for statistical analysis by student’s t-test.
Analysis of Residue
Mulberry leaves, collected fifteen days after second treatment were homogenized in a blender and extracted with ethanol. The residue run in thin layer chromatography plate (TLC) with the standard from the Acaciasides- pretreatment test substances. The test substances were Acaciasides- pretreatment test solution [22,25-28,30-32,35,36,46].
Plant pathogens caused Mulberry Diseases
Rhizospheric soil and root sample were taken at random from all the infected plots. Meloidogyne incognita populations (10 samples / plot in each plant group) were estimated in the rhizospheric soil as well as roots [22,25-28,30-32,34-36,46,50] of infected mulberry plants. Total number and surface area of leaves of all plant groups were counted [22,25-28,30-32,34-37,46]. Total number of root-galls/plant were counted in the infected roots of mulberry plants [22,46,51-58]. The total protein content of the leaf and root samples (10 at random sampling / plot) from each of the 16 plots was determined [57-59]. All the data from experiments were counted for statistical analysis by student’s t- test. In this field trial, sacrifices of mulberry plants were not done due to well reported pathological characters from our previous experiments [22,25-28,30-32,34-37,46].
Every year in September-October occurring, the main foliar diseases, observed in the sericulture field, were: leaf spot disease caused by Cercosporam moricola (Cooke) fungus pathogens, powdery mildew disease caused by Phyllactinia corylea (Pers.) Karst fungus pathogens, mosaic disease caused by mosaic virus pathogens and tukra disease caused by Maconellicoccus hirsutus (Green) mealy bug pathogens. All the disease identified according to their characteristic symptoms by the experts concerned [22,25-29,30,31,56,60-62]. Diseased leaves of each type were counted in each plots [22,25-29,30,31,56,60-62]. The percentage of disease infection based on diseased leaf surface area [22,25-31,33,59,63,64].
Rearing of silkworms
The eggs of a mother moth of the multivoltine ‘Nistari’ race (Bombyx mori L.) supplied by Regional Sericultural Research and Training Institute, Berhampore-742101, India, after hatching (93 % hatching rate) and brushing 1st stage silk worm larvae in the rearing tray, the larvae were divided into two batches (180 silkworm larvae / batch) and reared [22,25-31-35,59,63-65]. The larvae of infected untreated batch (control) were fed with the leaves of pathogens infected diseased leaves of mulberry plants from infected untreated (control) plots and the larvae of infected pretreated batch were fed with the leaves of Acaciasides-Pretreated leaves of mulberry plants from infected treated (control) plots. Fresh leaves were given to the larvae 4- times daily. Mulberry leaves were used for feeding fifteen days after the last Pretreatment with Acaciasides. The larvae were kept inside the rearing chamber at 27±2°C and 70 + 15% RH. The fresh weight of the larvae and that of the leaves served were recorded daily for each batch until the larvae started spinning. The consumption of fresh leaves [(Fresh leaves served – Dry leaves residues - Fresh leaves initially consumed) X Moisture loss], number of feeding and number of feeding day to cocoon formation, number of escaping feeding during moulting, moulting span days and mortality rate were recorded. The fresh silk gland weight of mature 5th instar larvae (before start spinning), starting time to spinning span of spinning, fresh cocoon weight, fresh shell weight, silk layer ratio (SR % = Shell weight / Cocoon weight X 100), effective rate of rearing (ERR % = Number of cocoon harvested / Number of silk worm hatched X 100), sex ratio percentage (Number of male adult emerged / Number of female adult emerged X 100) and egg laying capacity of mother moth were determined [9-15,17,18,46]. For statistical analysis by student’s t- test, ten mature 5th instar silkworm larvae for fresh silk gland weight and ten cocoons for fresh shell weight were dissected out in each batches including replica of all batches [22,25-31-35,59,63,64]. All the data from rearing trial were used for statistical analysis by student’s t- test.
Science and Technology Communication Applications
The activity of students, researchers, teachers, staff, community, photographers, visitors, different scientist, administrators, institutions, farmers, NGO and media personnel, -campaign or -aware or -make news or -publication regarding the importance of “Biomedicine- Acaciasides (A&B) use as potential phytomedicine against various pathogens: Enriching Science and Technology Communication Applications and Healthcare-, defense response- and immunity- as well as Biodiversity Conservation- issues” in different audio visual media (TV channels), social media, web pages, news papers and journals is recorded [22-26,30-32,37,45,47,52,65].
Future approach in research
It will achieved from typical analysis or justifications of literature review, research articles, specifies hypotheses, backgrounds, problems, brief review of the key literature, reports of clinical research trials or fields, note of any relevant controversies or disagreements in the trials or field, important references and data or conclusions from the work, extensive discussion of relevant literature as well as present investigation results. Specially, emphasize the new and important aspects of the study and the conclusions that follow from them. For experimental studies it is useful to clarify the main findings, then explore possible mechanisms or explanations for these findings, compare and contrast how the research is different from previous reporting and how the observations will significantly advancement of the current problem or knowledge of the subject, state the limitations of the study. Emphasis of claiming priority of work that has not been completed. Then new hypotheses will be arise and clearly label them as such trials [22-26,30-32,37,45,47,52,65].
Toxicity test on mortality
It was observed that Biomedicine- Acaciasides (A&B) had direct toxic effects on nematodes mortality within the exposure period of 24 hours because all the nematodes died and any recovery of nematodes occurred after 4 hours and no mortality occurred in the control.
Analysis of residues toxicity
Mulberry leaves collected fifteen days after the second treatment, did not contain any toxic residue of the Acaciasides- pretreated test substance.
Table1 shows the Pretreatment effects of Acaciasides (A&B) on Meloidogyne incognita pathogens infected mulberry plants in a field trial replicated thrice (P<0.01 by ‘t’- test). All naturally infected plants (Pretreated plant group) pretreated with Acaciasides (A&B) showed increase number and surface area of leaves, and higher protein content in leaves and root than infected Untreated (control) plants (Untreated plant group). In all infected Acaciasides- Pretreated plants, the population of root-knot nematodes decreased significantly in rhizospheric soil and as well as in roots than infected untreated (control) plants. The number of root galls also decreased significantly after acaciasides - Pretreatment.
Table 1: Effects of pretreatment with phytomedicines- Acaciasides (A&B) on root-knot diseases infected mulberry plants in a field trial replicated thrice.
Table 2 shows the pretreatment effects of acaciasides on leaf spot, powdery mildew, mosaic viral and tukra diseases of mulberry plants in a field trial replicated thrice assessed initially (Day- 0) and after a period of 30 days (Day -30) by ‘t’- test (P<0.01). Acaciasides (A&B) with Pretreatment significantly reduced the number of leaves infected with leaf spot, powdery mildew, mosaic viral and tukra (Figure 4) as compared to the untreated condition (Day- 0). The percentage of control achieved were 62.08% for leaf spot, 77.89% for powdery mildew (Figure 5), 64.91% for mosaic virus (Figure 6) and 38.42% for tukra infection as compared to the untreated level (Day- 0). In case of infected untreated plots leaf spot, powdery mildew, mosaic viral and tukra diseases showed naturally 27.80 %, 17.76 %, 29.37 % and 21.20 % reduction respectively, in 30 days (Day -30), and it is noted that there is no difference between previous-posttreatment-effects with present-pretreatment-effects.
Table 2: Effects of pretreated with phytomedicines- Acaciasides (A&B) on leaf spot, powdery mildew, mosaic and tukra diseases of mulberry plants in a field replicated thrice assessed initially (Day-0) and after a period of 30 days (Day-30).
Figure 4: Mulberry leaf infected with "Tukra Disease”.
Figure 5: Mulberry leaf infected with "powdery mildew'.
Figure 6: Mulberry leaf infected with "Mosaic Virus”.
Effects on feeding silkworms
Table 3 shows the Pretreatment effects of Acaciasides on diseased infected mulberry plants in a silkworm rearing and field trial replicated thrice on the feeding, growth and mortality of silkworms (P<0.01 by ‘t’-test). The average consumption of leaves by the 5th instars (Figure 7), average number of feeding to cocoon formation (Figure 8), average number of feeding day to cocoon formation, average number of escaping– feeding during moulting and average moulting span days were less for Acaciasides (A&B)- Pretreated plants than for infected Untreated (control) ones. The average mortality rate percent (%) was nil with Acaciasides- Pretreated plants groups and 56% with infected untreated (control) one. However, the average fresh weight of the 5th instars larvae were higher with acaciasides- Pretreated plants than with infected untreated (control) one, and it is noted that there is no difference between previous-posttreatment-effects with present-pretreatment-effects.
Table 3: Effects of disease-infected and Acaciasides- pretreated mulberry plants in a field on the feeding and growth of silkworms in the silkworms rearing trials (replicated thrice).
Figure 7: 5th instar larvae of Bombyx mori L.
Figure 8: Cocoon formation by 5th instar larvae.
Effects on silk production and rearing practices
Table 4 shows the effects of feeding Acaciasides- pretreated mulberry leaves on silk production, spinning characters and rearing practices in a silkworm rearing (Figure 8) and field trial replicated thrice (P<0.01 by ‘t’-test). The average fresh silk gland weight, average fresh cocoon weight, average fresh shell weight and average shell ratio (SR %) were higher with Acaciasides- pretreated plants than with infected untreated (control) one. It is notable that average starting time to spinning day and average span of spinning day (i.e. duration of span) were fewer with the Acaciasides- pretreated plants than with infected untreated (control) ones. Average effective rate of rearing (ERR%), average sex ratio percentage and average egg laying capacity were significantly higher with all Acaciasides- pretreated groups, and it is noted that there is no difference between previous-posttreatment-effects with present-pretreatment-effects.
Table 4: Effects of disease-infected and acaciasides- pretreated mulberry plants in a field on the growth of silk gland, spinning time, cocoon, shell, rearing, sex ratio and egg laying capacity in the silkworms rearing trials (replicated thrice).
Here, key findings
a. Here, pretreated with Acaciasides (A&B), isolated from the funicles of A. auriculiformis, were used against root-knot, leaf spot, powdery mildew, mosaic viral and tukra diseases caused by pathogens in a mulberry field trial. It was also observed the responses of silkworms feeding on mulberry leaves.
b. Acaciasides were soluble in water and applied to 6 plots twice at an interval of 15 days @ 1 mg / plant infected with above mentioned pathogens by foliar spray and soil drench using 20 ml solution for each type of pretreatment serving pretreatment plots. The remaining 6 plots treated with tap water serving Control plots.
c. Pretreated with Acaciasides, were highly effective in ameliorating mulberry diseases and were increased the protein content of mulberry leaves. Silkworm’s larvae feeding on the leaves of treated plants showed improved growth, increased cocoon weight and shell weight, fewer feeding to cocoon formation, zero mortality rate and increased the effective rate of rearing.
d. The results showed that Pretreated with acaciasides, use as potential eco-friendly phyomedicine against various pathogens of plants enriching agriculture significantly without disturbing biosphere.
Future approach in Research Proposals
The results fulfill the goal of a research proposal because the present pretreatments with phytomedicines; Acaciasides (A&B) need to justify future research and to present the practical ways in which the proposed study should be conducted by the future researcher for conducting the research consistent with requirements of the professional or academic field and a statement on anticipated outcomes and or benefits derived from the study's completion.
Science and Technology Communication Applications
The students, researchers, teachers, staff, community, photographers, visitors, different scientist, administrators, institutions, farmers, NGOs and media personnel campaign, aware, discuss, arrange workshops and seminars, make news and publish as abstract regarding the importance of “Pretreatment with Biomedicines; Acaciasides, use as Potential Phytomedicine Against Various Plant Pathogens: Enriching Science and Technology Communication Applications in Agriculture-, and Healthcare-, Defense Response-, Vaccinations- and Immunity- as well as Biodiversity Conservation- Issues” in different national- and local- audio visual media (TV channels), different social media, web pages, newspapers and different -national and –international Journals as well as Congress Proceedings also.
The pure pytocompounds Acaciasides (A&B), isolated from the funicles of A. auriculiformis, phytomedicine not only reduced root-knot, leaf spot, powdery mildew, viral and tukra diseases but also improved the nutritive value (specially protein) of the treated leaves of naturally infected plants [22,25-26,32,35,46]. Acaciasides-pretreatments directly influences on the consumption of leaves, number of feeding and number of feeding day to cocoon formation, and indirectly effects on molting stage in the infected treated groups from this trial. And due to ill development of infected Untreated (control) batches silkworm larvae took more time to molt which is proved from the number of escaping feeding during molting. Higher nutritive value specially protein of treated plants contributed to higher growth of silkworm larvae, silk gland weight, cocoon weight and shell weight which increase silk production significantly for commercial purpose [22,25-26,32,35,46]. The improved health of the larvae, cocoon weight, silk gland and shell weight from the Acaciasides- pretreated groups of infected plants might have resulted in the fewer starting time to spinning and span of spinning day and the total elimination of the mortality rate. Or, the Acaciasides might have infused in to mulberry leaves a substance that has conferred disease resistance on growing silkworm larvae by releasing defense-related natural products by plants [22,25,26,32,35-39,54,55,66]. For this, the effective rate of rearing (ERR%) is very high in all Acaciasides- pretreated treatment batches which enriches the sericulture industry in many ways, especially for commercial purpose. The mulberry leaves did not contain any toxic residues of the Acaciasides- Pretreated test substances by the thin layer chromatography (TLC). Rather, the acaciasides might have induced natural defense response in the test plants against all above mentioned pathogens and has conferred defense response on growing larvae [22,25,26,32,35-39,54,55,66-68].
The present study clearly showed that Acaciasides (A&B) were pretreated as effective or potential phytomedicine or biomedicine and it had no direct toxic effect on plants but to the pathogens of mulberry plants. The phytomedicines- acaciasides (A&B), could induced some resistance in mulberry against pathogens infection. It can be assumed that Acaciaside (A&B) could induce synthesis of some antagonistic substance in the pretreated plants. Lectins accumulated in gall regions of root of Hibiscus esculentus infected with M. incognita [32,35,69]. Systemic acquired resistance can be induced by in different crop plants by localized virus infection, non-pathogenic and pathogenic microorganisms or their culture filtrates or by salicylic acid [22,25-44,54,68-72]. Plant-derived natural products have important functions in ecological interactions. In some cases these compounds are deployed to sites of pathogen challenged by vesicle-mediated trafficking. Polar vesicle trafficking of natural products, proteins and other, as yet uncharacterized, cargo is emerging as a common theme in investigations of diverse disease resistance mechanisms in plants . Sequestration implies the involvement of specific transport process. For example, the defence-related triterpene glycoside avenaacin A-1 is synthesized .
It is reported that a plant plasma membrane ATP binding cassette-type transporter is involved in antifungal terpenoid secretion . Functional analysis has confirmed a role for this transporter in disease resistance . Though, M. incognita is known to share common antigens with its host plants . It appeared that during natural infection with the nematode, host plants showed minimal defense responses to the nematode because of this antigenic similarity. Acaciasides (A&B) contains two triterpenoids saponins [25,32,35,48,54] and these saponins provide defense to the test plants against pathogens [77-81]. Acaciasides must be responsible for defense resistance of the mulberry. Acaciasides (A&B) may synthesis various antigens particularly (low molecular weight proteins; 5kd to 25kd) and induce defense responses involving a number of pathogenesis related protein in which the naturally infected plant pathogens fails to tolerate [25,32,35,48,53-54]. It is observed that in lady’s finger plants treated with NE (nematode extract) i.e. Pretreated showed the highest number of root proteins (no. 24) but in inoculated untreated root was 18 number and uninoculated untreated root was 11 number [25,32,35,37]. Those showed that NE served as a stimulus for the expression of may proteins particularly the defense– related proteins which later provide resistance to pathogen- nematodes infection. However, in the test plant were treated with NE after inoculation i.e. Posttreated with live nematodes did not show much increase in number of proteins (23) in root [25,32,35,37].
Those showed that nematode pathogens infestation somehow serve as a repressor for the expression of defense gene in plant [25,32,35,37]. From this point of view, It can be assumed that Acaciasides serve as a stimulus for the expression of many new induced defense-related PR-proteins by systemic acquired resistance which provides defense-resistance to various pathogens causing major diseases of plants. It can be told that Acaciasides (A&B) acquiring systemic resistance could serve very effective eco-friendly Phytomedicine and promoted growth of test plants by inducing their defense responses of the host plants by expression of some new proteins against many plant pathogens infection causing major diseases and this Phytomedicine conserved our biodiversity and makes pollution free environment.
Now the key question is, whether plant-derived natural products (Biomedicine- Acaciasides- A&B), can be used as potential phytomedicine by inducing defense- response against various plant pathogens causing major mulberry diseases in a field trial and silkworms rearing. It is surprising that all naturally infected Acaciasides- pretreated plants not only are less affected by pathogens but also have a better growth than infected untreated (control) plants. The positive effects of growth may be responsible for defense resistance against pathogens. Acaciasides (A&B) might have induced synthesis of many new proteins which have stimulated increase photosynthesis rate, stomata- activity and water retention capacity of Acaciasides- Pretreated plants [30-44,82]. The positive effects of growth on disease-infected Pretreated plants might not only be responsible for defense resistance to pathogens but also improved growth of silkworm larvae and silk gland weight, cocoon weight, shell weight, effective rate of rearing (ERR%), sex ratio percentage and egg laying capacity of mother moth with zero mortality rate were higher with all Acaciasides- Pretreated groups which increase silk production for commercial purpose. It is proved from the results that silk production is higher in all Acaciasides- Pretreated plants than infected Untreated treated (control) plants. Now the answer is, Phytomedicine- Acaciasides was not only highly effective in ameliorating mulberry diseases but also enriched sericulture industry as well as agriculture industry.
Future Approach in Research Proposals
We’re not completely human, at least when it comes to the genetic material inside our cells. We all may harbor, as many as, one hundred forty five genes which have jumped from bacteria, other unicellular organisms, and viruses and made themselves at home in the human genome. All, the scientists indicated hundreds of genes that appeared to have been transferred from bacteria, archaea, fungi, other microorganisms, and plants to animals, they report online today in Genome Biology. In the case of humans, they found one hundred forty five genes that seemed to have jumped from simpler organisms, including 17 that had been reported in the past as possible horizontal gene transfers . The genomic sequencing showed that this pathogenic coronavirus is 96.2% identical to a bat coronavirus and shares 79.5% sequence identify to SARS-CoV [84-86].
The genomics of plant and animal, is a vast area of research with respect to the biological issues covered because it continues to deal with the structure and function of genetic material underpinning all organisms . Approximately, ten percent of the human genome is made of bits of virus- DNA. Mostly, this viral DNA is not always harmful. In some cases, researchers have found that actually it has a beneficial impact. When viruses infect us, they can embed small chunks of their genetic material in our DNA . The viral content of human genomes is more variable beyond our imagination. Millions of years ago, into the primordial genetic material of our progenitors, parts of human DNA are of viral origin were inserted and have been inherited by successive generations. Thus, the genomes of modern humans are not thought to vary much. HERV (Human endogenous retroviruses) are by far the most common virus-derived sequences in human genome and mobile DNA shows a mechanism that has introduced more inter-individual variation in HERV content between humans than previously appreciated . Ben L. Callif informs, “The Human Genome Is Full of Viruses and Your body requires viruses, but viruses don’t always require a body” . It is known that the “Human Genome Is Full of Viruses and Our body requires viruses, but viruses don’t always require a body” .
It is reported in a chapter entitled “Genetic Resistance to Coronavirus Infection- A Review” where researchers have organized their review of genetic resistance to coronaviruses according to those three host resistance mechanisms: genetic control at the level of the, -cellular receptors, -macrophage and -acquired immunity. However, they would like to stress that those ‘levels’ are purely operational boundaries. In reality, a host can be infected with a virus several times during its lifetime, and thus all available innate and immune resistance mechanisms will be called into play at once. In addition, they have included a general outline of the methods used to identify host resistance genes in mouse models of infection . SARS-CoV-2 is the etiological agent responsible for the pandemic COVID-19 outbreak and the main protease (Mpro) of SARS-CoV-2 is a key enzyme which plays a important role helping in viral replication and transcription. Structure-based design of antiviral drug candidates targeting the SARS-CoV-2 main protease .
Once the virus infects the host cell, it takes over the host cell’s machinery to produce more viruses. The host cell essentially becomes a virus factory. When the human body is attacked by germs, the immune system kicks into gear to fight of the assault. Germ fighting white blood cells in the body are called up to destroy the intruder. These cells target specific sites on the virus, working to destroy the infection. Also, a healthy person’s immune system creates a blueprint of the attacking agent. With this blueprint, the body effectively remembers the germ - enabling a person to fight of reinfection by the same or similar viruses .
In the evolution of human history shows the evidence that people are using traditional medicine for therapeutic purpose. The reports from the World Health Organization (WHO) claim that 70%- 80% population is primarily dependent on animals and plant-based medicines because of limited or no access to medical services. The drugs obtained from wild plants and animals are not only used as traditional medicines but also as raw materials in the formulation of modern allopathic and herbal preparations [1,14]. It is reported that as an internal treatment, the innate response of the patient's immune system to the presence of an invading microorganism has been studied, highlighting anti-microbial peptides as the host's own defense molecules. This work shows a compilation of the most relevant and current antimicrobial peptides that could be used as potential therapeutic agents against microorganisms located in the skin and related to acne disease . But in a case report of a congenital immune deficiency disease –WHIM syndrome, is a rare primary immunodeficiency disorder and it is an acronym for some of the characteristic symptoms of the disorder; warts, hypogammaglobulinemia, infections, and myelokathexis. This case report provides data of a patient with recurrent respiratory and cutaneous infection who was diagnosed with WHIM syndrome was presented with chronic productive cough, fever, pleuritic chest pain, chills and sweating also .
It is reported that the Biomedicine or Phytomedicine- Acaciasides (A&B) is being used traditionally to overcome various medical complications like sore eyes, aches, rheumatism, allergy, itching, and rashes. Besides, it has also been proven for many pharmacological activities like central nervous system depressant activity, antioxidant, antimicrobial, antimalarial, antifilarial, anticestocidal, antimutagenic, chemopreventive, spermicidal, wound healing, hepatoprotective and antidiabetic activity due to its low toxicity (LD50 = 3741.7 mg/kg) and high efficacy and the various phytochemical investigations reveal the presence of chief constituents as flavonoids and triterpenoid saponin glycosides (Acaciasides- A&B). The low toxicity and the presence of major bioactive phytoconstituents like flavonoids and triterpenoid saponin glycosides are responsible for a therapeutic remedy for various diseases and pharmacological activities respectively. It has been used to treat several medical ailments due to its low toxicity and the presence of bioactive phytoconstituents . Isolated saponins (acaciasides -A&-B acylated triterpenoid bisglycoside) from fruits (funicles) of A. auriculiformis were screened for their antifilarial activity and results were found to be significant . A US patent claimed the potential of acaciasides (A&B) isolated from A. auriculiformis for the prevention of HIV infection and as a vaginal contraceptive .
Recently, in the Drug Target Review 2020, NovavaxInc, which contributed to the development of other epidemic vaccines, has announced it is currently in pre-clinical animal trials for several multiple nanoparticle COVID-19 vaccine candidates. The biotechnology company has announced its efforts to help in creating a vaccine against SARS-CoV-2. The company stated they have used their recombinant protein nanoparticle technology platform to generate antigens derived from the coronavirus spike protein and their previous experience working with other coronaviruses, including both MERS and SARS, allowed them to mobilize quickly against COVID-19 and successfully complete the critical preliminary steps to engineer viable vaccine candidates, as said by Stanley Erck, President and Chief Executive Officer of Novavax which adjuvant is saponin-based and it has shown a “potent and well-tolerated effect” [98,99].
For successful vaccination requires four components; knowing the vaccine target, what kind of immune response, how to generate that response, and understanding responses in the people who we want to vaccinate. Human Immunomics Initiative (HII) aims to decode the underlying mechanisms and rules of how the human immune system fights disease with advances in computing and artificial intelligence, genomics, systems biology, and bioinformatics . And should follow guide line of WHO entitled “Vaccine-preventable diseases and vaccines” .
It is already reported that the presence of chief constituents as flavonoids and two acylated triterpenoidbisglycoside saponins present in Acaciasides (A&B) [14,22,25,37,39,41,48,53,77,99,102-107]. In medicine, it may be used in vaccine formulations to regulate immune function because it acts as antioxidants and scavenge oxidative stress. The adjuvant Acaciasides (A&B) may be used with recombinant protein nanoparticle antigens derived from the coronavirus spike protein and combine these antigens with its adjuvant Acaciasides (A&B) for the final formulation of the vaccine and it may be shown a “potent and well-tolerated effect” through stimulating the entry of antigen-presenting cells into the injection site and enhancing antigen presentation in local lymph nodes, boosting immune responses [32,35,98,99]. In a letter as an e-mail, the Science Advisory Board Net, at Express Cells, for their business of creating better knock-in cell lines for drug discovery, toxicology, and other biologic research and add for purchase SARS-CoV-2 Spike Protein (NC_045512.2), SARS-CoV-2 Nucleocapsid Protein (NC_045512.2), TMPRSS2 (NM_001135099.1), ACE2 (NM_021804.3), BSG (CD147) (NM_001728.3), SARS-CoV Nucleocapsid Protein (MK062179.1), SARS-CoV Spike Protein (MK062179.1), MERS-CoV Nucleocapsid Protein (NC_019843.3), MERS-CoV Spike Protein (NC_019843.3)  and the readily available coronavirus spike proteins may be helped to use for vaccine preparation which may fight against “Covid Toes among kids: New symptom of novel coronavirus infection “ and it is obligatory to inform the concerned authority- ‘ClinicalTrials.gov.’ to maintain all applicable laws and regulations, in the study-schedules and emphasis on poor-developing countries for cost-effective-emergency-vaccine [109-111].
The adjuvant -Acaciasides (A&B) may be used with anti-Human antibodies like IgG (A80-104A, A80-105A), IgM (A80-100A, A80-101A), & IgA (A80-102A, A80-103A) and offer treatments or vaccine preparation of COVID-19 (SARS-CoV-2) and it may also be accelerated the discovery to improve lives . After achieving successful clinical trials, the concerned authority- ClinicalTrials.gov, may be permitted for the use as a vaccine, for treatments for cost-effective-emergency-vaccine [110-111].
Here, the results and discussion fulfill the goal of a research ideas because the present Pretreatments with phytomedicine- Acaciasides (A&B) need to justify future research and to present the practical methods in which the proposed study should be conducted. The plans for conducting research are governed by standards of the results in which the solutions or problems resides, therefore, the guidelines for research proposals are more exacting and less formal than a general project proposal [32,35,47]. Research ideas contain extensive literature reviews. It is amazed for use of pretreated with Acaciasides - directly or -indirectly help the society in various ways and may also provide a unique platform for showcasing the research across the globe and progress the further advanced research in Agriculture, Sericulture, Horticulture and Entomology deals with applications economy. Common people also realize the meaning of new and more efficient solutions, technologies, products and it has to improve Science and Technology Communication Applications Economy forming joyful environment and fulfill its food and nutrition requirement which resist any kinds of thresholds or natural infections for the climate change and resource productive application economies enriching quality of environment.
Now it is planned to publish as a proposal for current outcomes and therapies on coronavirus disease (COVID-19) outbreak that helps the readers as well as a scientific community to take measures or treatment opportunity or discovery of vaccine to avoid new coronavirus. Our main goal is to limit infections. Let us all take this basic information’s as proposal and also educate people, help them to fight against this war, the normal life of everyone is on hold due to this escalating coronavirus emergency, which in a way helps all the scientist, readers, authors and editors to take necessary and respective steps to save or avoid this dangerous disease. I request all to support this initiative and help to reach the targeted audience. At this critical time in the global response to the COVID-19 pandemic, I am proud to be working alongside the scientific community as a coworker that accelerate research, discovery, and testing to contain the SARS-CoV-2 outbreak. And it also focus the future Trends in Medicine which serves as a evidence-based resource covering various disciplines of medicine and clinical practice and acts as indispensable source to access the pharmacological developments. And it also deals with articles related to the translational research or investigations in clinical practice, epidemiological studies and general topics of interest to the biomedical research community.
The cost effective, non-phytotoxic, non-pollutant, easily available biomedicines; Acaciasides (A&B), not only use as potential phytomedicine against various plant pathogens like root-knot disease, leaf spot disease, powdery mildew disease, mosaic disease and tukra disease of mulberry- and improves growth of silk worms- at an extremely low dose enriching sericulture industry as well as agriculture sector commercially, but also efficient phytomedicine against various diseases of animal as well as human by inducing their natural immunity. And it would go a long way in tackling various pests of crops in a safe way by inducing their defense-responses of host plants against pathogens. In near future, it is expected that a clinical study or trials or treatments may be arranged for discovery of ‘Vaccine’ by using phytomedicine- Acaciasides which may kill the novel Coronavirus COVID-19 by -improving immune function or by -stimulating the production of T-cells, and -generating superoxide anions, -initiating lipid peroxidation and -dissolving the fatty coats of coronaviruses and the whole world may retain in normal forms by fighting against this war, and acts as indispensable source to access the pharmacological developments for clinical practice, epidemiological studies and biomedical research community by focusing the future trends in medicine.
The work described here has been supported by Rtd. Prof. N.C.Sukul and Prof. S.P.Sinha Babu, Department of Zoology, Visva-Bharati and Joint Director, Sriniketan Sericultural Composite Unit, Govt. of West Bengal. I am thankful to the eminent Senior Consultant Physician & Cardiologist Dr. Tushar Kanti Batabyal, M.B.B.S., M.D., Ex-Clinical Tutor of Burdwan Medical College & Hospital, for inspiration and guidance. I also like to congratulates Mr. Rakesh Khan, Secretary and Mr. Subhendu Bose, President with all Young Green-Members of “NGO named Burdwan Green Haunter and Students’ Goal” for arranging several awareness programme on “Health Care, Vaccination, Biodiversity Conservation and Enriching Science and Technology Communication Economy Application Issues”. Last but not the least, I am thankful to eminent educationist Sri Tapaprakash Bhattacharya for continuous supports & helping in writing manuscripts.
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